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Biomedical subjects

D H Turnbull

Publications and source records attributed to D H Turnbull.

At least 19 recordsLinked to original sources
PubMed2001

The HD mutation causes progressive lethal neurological disease in mice expressing reduced levels of huntingtin.

Huntingtin is an essential protein that with mutant polyglutamine tracts initiates dominant striatal neurodegeneration in Huntington's disease (HD). To assess the consequences of mutant protein when huntingtin is limiting, we have studied three lines of compound heterozygous mice in which both copies of the HD gene homolog (Hdh) were altered, resulting in greatly reduced levels of huntingtin with a normal human polyglutamine length (Q20) and/or an expanded disease-associated segment (Q111): Hdh(neoQ20)/Hdh(neoQ20), Hdh(neoQ20)/Hdh(null) and Hdh(neoQ20)/Hdh(neoQ111). All surviving mice in each of the three lines were small from birth, and had variable movement abnormalities. Magnetic resonance micro-imaging and histological evaluation showed enlarged ventricles in approximately 50% of the Hdh(neoQ20)/Hdh(neoQ111) and Hdh(neoQ20)/Hdh(null) mice, revealing a developmental defect that does not worsen with age. Only Hdh(neoQ20)/Hdh(neoQ111) mice exhibited a rapidly progressive movement disorder that, in the absence of striatal pathology, begins with hind-limb clasping during tail suspension and tail stiffness during walking by 3-4 months of age, and then progresses to paralysis of the limbs and tail, hypokinesis and premature death, usually by 12 months of age. Thus, dramatically reduced huntingtin levels fail to support normal development in mice, resulting in reduced body size, movement abnormalities and a variable increase in ventricle volume. On this sensitized background, mutant huntingtin causes a rapid neurological disease, distinct from the HD-pathogenic process. These results raise the possibility that therapeutic elimination of huntingtin in HD patients could lead to unintended neurological, as well as developmental side-effects.

Animals↗
PubMed2001

Telencephalic neural progenitors appear to be restricted to regional and glial fates before the onset of neurogenesis.

The contribution of early cell lineage to regional fate in the mammalian forebrain remains poorly understood. Previous lineage-tracing studies using retroviral methods were only begun at mid-neurogenesis and have suffered from region-specific retroviral silencing. We have been able to study cell lineage in the telencephalon from the onset of neurogenesis by using ultrasound backscatter microscopy to label the forebrain neuroepithelium and a modified retroviral lineage library to overcome regional silencing. Our studies suggest that by embryonic day 9.5, forebrain clones are primarily restricted to territories within anatomically demarcated regional boundaries, such as the cortex, striatum and hypothalamus. In addition, we observed a subset of clones that appeared to be composed entirely of glia. These observations suggest that both regional and cell-type restrictions exist within progenitor populations before the first forebrain cells become postmitotic.

Animals↗
PubMed2001

Sensitivity and performance time in MRI dephasing artifact reduction methods.

Although shimming can improve static field inhomogeneity, local field imperfections induced by tissue susceptibility differences cannot be completely corrected and can cause substantial signal loss in gradient echo images through intravoxel dephasing. Dephasing increases with voxel size so that one simple method of reducing the effect is to use thin slices. Signal-to-noise ratio (SNR) can then be increased by averaging over the subslices to form the final, thick slice. We call this method subslice averaging or SSAVE. Alternatively, a range of different amplitude slice select rephase gradients can be used to compensate for different susceptibility induced gradient offsets. The final image can then be formed by combining individual images in a variety of ways: summation, summation of the squares of the images, forming the maximum intensity projection of the image set, and Fourier transformation followed by summation. We show here that, contrary to previous claims, the theoretical sensitivity (i.e., SNR divided by the square root of the imaging time) of all these alternative methods is very similar. However, performance time (i.e., minimum-imaging time) of the simplest method, SSAVE, is much shorter than that of alternatives. This is confirmed experimentally on phantoms and anesthetized mice. Magn Reson Med 45:470-476, 2001.

Animals↗
PubMed2001

In utero fate mapping reveals distinct migratory pathways and fates of neurons born in the mammalian basal forebrain.

Recent studies suggest that neurons born in the developing basal forebrain migrate long distances perpendicularly to radial glia and that many of these cells reach the developing neocortex. This form of tangential migration, however, has not been demonstrated in vivo, and the sites of origin, pathways of migration and final destinations of these neurons in the postnatal brain are not fully understood. Using ultrasound-guided transplantation in utero, we have mapped the migratory pathways and fates of cells born in the lateral and medial ganglionic eminences (LGE and MGE) in 13.5-day-old mouse embryos. We demonstrate that LGE and MGE cells migrate along different routes to populate distinct regions in the developing brain. We show that LGE cells migrate ventrally and anteriorly, and give rise to the projecting medium spiny neurons in the striatum, nucleus accumbens and olfactory tubercle, and to granule and periglomerular cells in the olfactory bulb. By contrast, we show that the MGE is a major source of neurons migrating dorsally and invading the developing neocortex. MGE cells migrate into the neocortex via the neocortical subventricular zone and differentiate into the transient subpial granule neurons in the marginal zone and into a stable population of GABA-, parvalbumin- or somatostatin-expressing interneurons throughout the cortical plate.

Animals↗
PubMed2000

Two lineage boundaries coordinate vertebrate apical ectodermal ridge formation.

Proximal-distal outgrowth of the vertebrate limb bud is regulated by the apical ectodermal ridge (AER), which forms at an invariant position along the dorsal-ventral (D/V) axis of the embryo. We have studied the genetic and cellular events that regulate AER formation in the mouse. In contrast to implications from previous studies in chick, we identified two distinct lineage boundaries in mouse ectoderm prior to limb bud outgrowth using a Cre/loxP-based fate-mapping approach and a novel retroviral cell-labeling technique. One border is transient and at the limit of expression of the ventral gene En1, which corresponds to the D/V midline of the AER, and the second border corresponds to the dorsal AER margin. Labeling of AER precursors using an inducible Cre showed that not all cells that initially express AER genes form the AER, indicating that signaling is required to maintain an AER phenotype. Misexpression of En1 at moderate levels specifically in the dorsal AER of transgenic mice was found to produce dorsally shifted AER fragments, whereas high levels of En1 abolished AER formation. In both cases, the dorsal gene Wnt7a was repressed in cells adjacent to the En1-expressing cells, demonstrating that signaling regulated by EN1 occurs across the D/V border. Finally, fate mapping of AER domains in these mutants showed that En1 plays a part in positioning and maintaining the two lineage borders.

Animals↗
PubMed2000

40 MHz Doppler characterization of umbilical and dorsal aortic blood flow in the early mouse embryo.

Physiological study of the developing mouse circulation has lagged behind advances in molecular cardiology. Using an innovative high-frequency Doppler system, we noninvasively characterized circulatory hemodynamics in early mouse embryos. We used image-guided 43 MHz pulsed-wave (PW) Doppler ultrasound to study the umbilical artery and vein, or dorsal aorta in 109 embryos. Studies were conducted on embryonic days (E) 9.5-14.5. Heart rate, peak blood flow velocities, and velocity time integrals in all vessels increased from E9.5-14.5, indicating increasing stroke volume and cardiac output. Heart rate, ranging from 192 bpm (E9.5) to 261 bpm (E14.5), was higher than previously reported. Placental impedance, assessed by the time delay between the peaks of the umbilical arterial and venous waveforms and by venous pulsatility, decreased with gestation. Acceleration time, a load-independent Doppler index of cardiac contractility, remained constant but seemed sensitive to heart rate. High-frequency PW Doppler is a powerful tool for the quantitative, noninvasive investigation of early mouse circulatory development.

Animals↗
PubMed2000

Advances in ultrasound biomicroscopy.

The visualisation of living tissues at microscopic resolution is attracting attention in several fields. In medicine, the goals are to image healthy and diseased tissue with the aim of providing information previously only available from biopsy samples. In basic biology, the goal may be to image biological models of human disease or to conduct longitudinal studies of small-animal development. High-frequency ultrasonic imaging (ultrasound biomicroscopy) offers unique advantages for these applications. In this paper, the development of ultrasound biomicroscopy is reviewed. Aspects of transducer development, systems design and tissue properties are presented to provide a foundation for medical and biological applications. The majority of applications appear to be developing in the 40-60-MHz frequency range, where resolution on the order of 50 microm can be achieved. Doppler processing in this frequency range is beginning to emerge and some examples of current achievements will be highlighted. The current state of the art is reviewed for medical applications in ophthalmology, intravascular ultrasound, dermatology, and cartilage imaging. Ultrasound biomicroscopic studies of mouse embryonic development and tumour biology are presented. Speculation on the continuing evolution of ultrasound biomicroscopy will be discussed.

Animals↗
PubMed1999

2D multislice and 3D MRI sequences are often equally sensitive.

A simple theoretical model was developed to compare the sensitivities (i.e., signal-to-noise ratios per unit imaging time) of two-dimensional (2D) multislice and 3D imaging sequences. The model shows that the sensitivities of 3D and 2D multislice MRI sequences are usually similar. Sensitivities are identical in T2-weighted sequences when the T(R)s of the two sequences are the same. In T1-weighted gradient-echo sequences, sensitivities are very similar when Ernst angle excitation is used and the T(R) of the 2D sequence is less than T1. The predictions of the model are confirmed in phantom and animal experiments.

Animals↗
PubMed1999

A method for rapid gain-of-function studies in the mouse embryonic nervous system.

We used ultrasound image-guided injections of high-titer retroviral vectors to obtain widespread introduction of genes into the mouse nervous system in utero as early as embryonic day 8.5 (E8.5). The vectors used included internal promoters that substantially improved proviral gene expression in the ventricular zone of the brain. To demonstrate the utility of this system, we extended our previous work in vitro by infecting the telencephalon in vivo as early as E8. 5 with a virus expressing Sonic Hedgehog. Infected embryos showed gross morphological brain defects, as well as ectopic expression of ventral telencephalic markers characteristic of either the medial or lateral ganglionic eminences.

Abnormalities, Multiple↗
PubMed1999

Neonatal cardiomyopathy in mice homozygous for the Arg403Gln mutation in the alpha cardiac myosin heavy chain gene.

Heterozygous mice bearing an Arg403Gln missense mutation in the alpha cardiac myosin heavy chain gene (alpha-MHC403/+) exhibit the histopathologic features of human familial hypertrophic cardiomyopathy. Surprisingly, homozygous alpha-MHC403/403 mice die by postnatal day 8. Here we report that neonatal lethality is caused by a fulminant dilated cardiomyopathy characterized by myocyte dysfunction and loss. Heart tissues from neonatal wild-type and alpha-MHC403/403 mice demonstrate equivalent switching of MHC isoforms; alpha isoforms in each increase from 30% at birth to 70% by day 6. Cardiac dimensions and function, studied for the first time in neonatal mice by high frequency (45 MHz) echocardiography, were normal at birth. Between days 4 and 6, alpha-MHC403/403 mice developed a rapidly progressive cardiomyopathy with left ventricular dilation, wall thinning, and reduced systolic contraction. Histopathology revealed myocardial necrosis with dystrophic calcification. Electron microscopy showed normal architecture intermixed with focal myofibrillar disarray. We conclude that 45-MHz echocardiography is an excellent tool for assessing cardiac physiology in neonatal mice and that the concentration of Gln403 alpha cardiac MHC in myocytes influences both cell function and cell viability. We speculate that variable incorporation of mutant and normal MHC into sarcomeres of heterozygotes may account for focal myocyte death in familial hypertrophic cardiomyopathy.

Animals↗
PubMed1999

Dilated cardiomyopathy in homozygous myosin-binding protein-C mutant mice.

To elucidate the role of cardiac myosin-binding protein-C (MyBP-C) in myocardial structure and function, we have produced mice expressing altered forms of this sarcomere protein. The engineered mutations encode truncated forms of MyBP-C in which the cardiac myosin heavy chain-binding and titin-binding domain has been replaced with novel amino acid residues. Analogous heterozygous defects in humans cause hypertrophic cardiomyopathy. Mice that are homozygous for the mutated MyBP-C alleles express less than 10% of truncated protein in M-bands of otherwise normal sarcomeres. Homozygous mice bearing mutated MyBP-C alleles are viable but exhibit neonatal onset of a progressive dilated cardiomyopathy with prominent histopathology of myocyte hypertrophy, myofibrillar disarray, fibrosis, and dystrophic calcification. Echocardiography of homozygous mutant mice showed left ventricular dilation and reduced contractile function at birth; myocardial hypertrophy increased as the animals matured. Left-ventricular pressure-volume analyses in adult homozygous mutant mice demonstrated depressed systolic contractility with diastolic dysfunction. These data revise our understanding of the role that MyBP-C plays in myofibrillogenesis during cardiac development and indicate the importance of this protein for long-term sarcomere function and normal cardiac morphology. We also propose that mice bearing homozygous familial hypertrophic cardiomyopathy-causing mutations may provide useful tools for predicting the severity of disease that these mutations will cause in humans.

Alleles↗
PubMed1998

Noninvasive, in utero imaging of mouse embryonic heart development with 40-MHz echocardiography.

BACKGROUND: The increasing number of transgenic and targeted mutant mice with embryonic cardiac defects has resulted in the need for noninvasive techniques to examine cardiac structure and function in early mouse embryos. We report the first use of a novel 40-MHz ultrasound imaging system in the study of mouse cardiac development in utero. METHODS AND RESULTS: Transabdominal scans of mouse embryos staged between 8.5 and 13.5 days of gestation (E8.5 to E13.5) were obtained in anesthetized mice. Atrial and ventricular contractions could be discerned from E9.5, and changes in cardiac morphology were observed from E9.5 to E13.5. Hyperechoic streaming patterns delineated flow through the umbilical, vitelline, and other major blood vessels. Diastolic and systolic ventricular areas were determined by planimetry of the epicardial borders, and fractional area change was measured as an index of contractile function. Significant increases in ventricular size were documented at each stage between E10.5 and E13.5, and the ability to perform serial imaging studies over 3 days of embryonic development is described. Finally, the detection of vascular cell adhesion molecule 1 (VCAM-1) homozygous null mutant embryos demonstrates the first example of noninvasive, in utero analysis of cardiac structure and function in a targeted mouse mutant. CONCLUSIONS: We used 40-MHz echocardiography to identify key elements of the early mouse embryonic cardiovascular system and for noninvasive dimensional analysis of developing cardiac ventricles. The ability to perform serial measurements and to detect mutant embryos with cardiac defects highlights the usefulness of the technique for investigating normal and abnormal cardiovascular development.

Animals↗
PubMed1998

40-MHZ echocardiography scanner for cardiovascular assessment of mouse embryos.

Congenital heart disease results from genetic defects that are manifested at early stages of embryogenesis. The mouse is the preferred animal model for studies of mammalian embryonic development and for an increasing number of human disease models. A number of genes identified in the mouse are critical for normal cardiovascular development, but an understanding of the underlying mechanisms regulating heart development is still incomplete, in part because of the lack of methods to measure hemodynamics in live mouse embryos. We describe the development of a 40-MHz ultrasound scanner, which allows image-guided continuous-wave and pulsed Doppler blood flow measurements in mouse embryos, in utero, at the critical early developmental stages. Doppler waveforms acquired from mouse embryonic umbilical vessels, descending aorta, and cardiac ventricles are presented to demonstrate the utility of the method. By combining image-guided ultrasound Doppler with the many available mouse mutants, this approach should lead to new insights into embryonic cardiovascular structure-function relationships.

Animals↗
PubMed1998

Alteration of limb and brain patterning in early mouse embryos by ultrasound-guided injection of Shh-expressing cells.

A basic limitation of the study of development in the mouse is the inaccessibility of the embryos, which are encased in the maternal uterus. We demonstrate the first use of ultrasound backscatter microscopy for guiding injections of cells and other agents into early stage mouse embryos. Cells were injected into the mouse neural tube cavity as early as 9.5 days post coitus (E9.5), and into the developing limb buds as early as E10.5. Furthermore, a cell-line engineered to express the secreted factor Sonic Hedgehog (Shh) was injected into early developing mouse brains or limbs. The Shh-expressing cells were found to induce ectopic expression of the Shh target genes Patched and Hnf3beta in the dorsal brain, and to alter digit patterning in the anterior limb bud. These results show that gene misexpression studies can be performed in mouse embryos using ultrasound-guided injection of transfected cells or retroviruses. In combination with the many available mouse mutants, this method offers a new approach for analyzing genetic interactions through gain-of-function studies performed in mutant mouse backgrounds.

Animals↗
PubMed1997

Specification of mouse telencephalic and mid-hindbrain progenitors following heterotopic ultrasound-guided embryonic transplantation.

We have demonstrated the utility of ultrasound backscatter microscopy for targeted intraparenchymal injections into embryonic day (E) 13.5 mouse embryos. This system has been used to test the degree of commitment present in neural progenitors from the embryonic ventral telencephalon and mid-hindbrain region. Many E13.5 ventral telencephalic progenitors were observed to integrate and adopt local phenotypes following heterotopic transplantation into telencephalic or mid-hindbrain targets, whereas mid-hindbrain cells of the same stage were unable to integrate and change fate in the telencephalon. In contrast, many mid-hindbrain cells from an earlier developmental stage (E10.5) were capable of integrating and adopting a forebrain phenotype after grafting into the telencephalon, suggesting that mouse mid-hindbrain progenitors become restricted in their developmental potential between E10.5 and E13.5.

Animals↗
PubMed1996

Ultrasound backscatter microscope analysis of mouse melanoma progression.

The incidence and mortality rate of cutaneous melanoma continue to increase throughout the world, making the study of melanoma biology an important area of current research. While recent breakthroughs in transgenic mouse technology have led to promising mouse skin models of melanoma, there is presently no technique available for quantitatively studying subsurface melanoma progression, in vivo. We demonstrate the first application of an imaging method called ultrasound backscatter microscopy (UBM) for imaging early murine melanomas with spatial resolution of 30 microns axial and 60 microns lateral. Murine B16 F10 melanomas have been imaged from their earliest detection, over several days, until they are 2 to 5 mm in diameter. Melanoma dimensions measured by UBM were found to be in excellent agreement with those determined histopathologically on the excised tumours. The relative rms errors in UBM-determined melanoma height and width were found to be 8.7% and 4.2%, respectively. The mean rate of increase in tumour height of early murine melanoma was found to be 0.37 +/- 0.06 mm/day. Computer-generated volumetric renderings of melanomas have been produced from three-dimensional image data, allowing quantitative comparisons of tumour volumes to be made. Using a priori assumptions of ellipsoid tumour shape, the relative error in UBM-determined volume was shown to be less than 17%. These results should be of considerable interest to investigators studying melanoma biology using mouse skin models, and have implications in the use of high frequency ultrasound imaging for the clinical assessment of cutaneous melanoma.

Animals↗
PubMed1996

High frequency 40-MHz ultrasound. A possible noninvasive method for the assessment of the boundary of basal cell carcinomas.

BACKGROUND: Ultrasound imaging systems operating close to 20 MHz in frequency have been used to image skin tumors. Ultrasound imaging at 20 MHz has been used to determine the boundaries of basal cell carcinomas (BCCs). An inherent shortcoming of imaging systems operating at these frequencies is their limited resolution. OBJECTIVE: We investigated whether 40-MHz ultrasound imaging could provide higher resolution compared with the lower frequency systems and thus be a superior, noninvasive method of assessing the boundaries of BCCs. METHODS: Nine BCCs from six individuals were examined clinically and ultrasonographically, and then biopsied to confirm diagnosis. The depth of BCCs measured on histological sections was compared with the corresponding value obtained using ultrasound. For this study we required a nonsurgical, nondestructive means of treating BCCs that would allow repeated ultrasound imaging, and therefore topical 5-flurouracil (5-FU) was chosen. Following 5-FU therapy a biopsy was obtained from the site of the treated BCC after ultrasound imaging had been performed. Clinical, ultrasonic and histopathologic evaluation of each BCC was carried out independently by different individuals. At the end of the study all the BCC sites were treated surgically be electrodesiccation and curettage or completely excised. RESULTS: High resolution ultrasound images of BCCs were obtained with agreement between histology and ultrasound findings in all none lesions prior to therapy and in eight of none lesions posttherapy. There was a significant correlation between the depth of BCCs measured histologically and using ultrasound (P = 0.0004, r = 0.92). CONCLUSIONS: This study suggests that 40-MHz ultrasound may provide an estimate of the boundary of a BCC in vivo. High frequency 40-MHz ultrasound imaging may be an adjunct to clinical and histologic evaluation but does not replace the need to obtain tissue for microscopic examination.

Aged↗