The 1.4-Mb CMT1A duplication/HNPP deletion genomic region reveals unique genome architectural features and provides insights into the recent evolution of new genes.

Duplication and deletion of the 1.4-Mb region in 17p12 that is delimited by two 24-kb low copy number repeats (CMT1A-REPs) represent frequent genomic rearrangements resulting in two common inherited peripheral neuropathies, Charcot-Marie-Tooth disease type 1A (CMT1A) and hereditary neuropathy with liability to pressure palsy (HNPP). CMT1A and HNPP exemplify a paradigm for genomic disorders wherein unique genome architectural features result in susceptibility to DNA rearrangements that cause disease. A gene within the 1.4-Mb region, PMP22, is responsible for these disorders through a gene-dosage effect in the heterozygous duplication or deletion. However, the genomic structure of the 1.4-Mb region, including other genes contained within the rearranged genomic segment, remains essentially uncharacterized. To delineate genomic structural features, investigate higher-order genomic architecture, and identify genes in this region, we constructed PAC and BAC contigs and determined the complete nucleotide sequence. This CMT1A/HNPP genomic segment contains 1,421,129 bp of DNA. A low copy number repeat (LCR) was identified, with one copy inside and two copies outside of the 1.4-Mb region. Comparison between physical and genetic maps revealed a striking difference in recombination rates between the sexes with a lower recombination frequency in males (0.67 cM/Mb) versus females (5.5 cM/Mb). Hypothetically, this low recombination frequency in males may enable a chromosomal misalignment at proximal and distal CMT1A-REPs and promote unequal crossing over, which occurs 10 times more frequently in male meiosis. In addition to three previously described genes, five new genes (TEKT3, HS3ST3B1, NPD008/CGI-148, CDRT1, and CDRT15) and 13 predicted genes were identified. Most of these predicted genes are expressed only in embryonic stages. Analyses of the genomic region adjacent to proximal CMT1A-REP indicated an evolutionary mechanism for the formation of proximal CMT1A-REP and the creation of novel genes by DNA rearrangement during primate speciation.

Initial sequencing and analysis of the human genome.

The human genome holds an extraordinary trove of information about human development, physiology, medicine and evolution. Here we report the results of an international collaboration to produce and make freely available a draft sequence of the human genome. We also present an initial analysis of the data, describing some of the insights that can be gleaned from the sequence.

Dynamic modeling of interstitial laser photocoagulation: implications for lesion formation in liver in vivo.

BACKGROUND AND OBJECTIVE: Interstitial Laser Photocoagulation (ILP) is a minimally invasive cancer treatment technique, whereby optical energy from implanted optical fibers is used to therapeutically heat small, solid tumors. In this work, the potential of ILP without tissue charring is investigated. STUDY DESIGN/MATERIALS AND METHODS: Optical diffusion and bio-heat transfer equations were used to develop dynamic models of interstitial laser heating in liver in vivo. Modifications in the optical properties due to tissue coagulation (T > or = 60 degrees C) were incorporated into the physical description. In addition, the effect of three different blood perfusion patterns on temperature distributions was explored. Model-predicted temperatures were used as an index for thermal damage based on an accumulated temperature injury (Arrhenius) model. Thermal damage dimensions were determined with tissue temperatures constrained to remain below 100 degrees C, so as to minimize the potential for tissue charring and smoke production. RESULTS: The model predicts that increases in scattering due to coagulation and choice of perfusion pattern affect substantially thermal damage dimensions. The results indicate that, for single fiber ILP at 2.55 W for 600 s, the maximum achievable thermal damage diameter in liver, without charring, is 9.6 mm. In addition, ILP performed with high-low power ramping may have an advantage over constant power treatments, in that, larger volumes of thermal damage can be realized earlier in an irradiation. CONCLUSIONS: For ILP performed with a single spherical emitting fiber, optimal irradiation parameters exist such that thermal lesions in liver up to approximately 10 mm in diameter can be induced while the maximum tissue temperature remains below 100 degrees C, avoiding tissue charring.

Evaluating patients for return to work.

The family physician is often instrumental in the process of returning a patient to the workplace after injury or illness. Initially, the physician must gain an understanding of the job's demands through detailed discussions with the patient, the patient's work supervisor or the occupational medicine staff at the patient's place of employment. Other helpful sources of information include job demand analysis evaluations and the Dictionary of Occupational Titles. With an adequate knowledge of job requirements and patient limitations, the physician should document specific workplace restrictions, ensuring a safe and progressive reentry to work. Occupational rehabilitation programs such as work hardening may be prescribed, if necessary. If the physician is unsure of the patient's status, a functional capacity evaluation should be considered. The family physician should also be familiar with the Americans with Disabilities Act as it applies to the patient's "fitness" to perform the "essential tasks" of the patient's job.

Mutational analysis of the Drosophila sister-chromatid cohesion protein ORD and its role in the maintenance of centromeric cohesion.

The ord gene is required for proper segregation of all chromosomes in both male and female Drosophila meiosis. Here we describe the isolation of a null ord allele and examine the consequences of ablating ord function. Cytologically, meiotic sister-chromatid cohesion is severely disrupted in flies lacking ORD protein. Moreover, the frequency of missegregation in genetic tests is consistent with random segregation of chromosomes through both meiotic divisions, suggesting that sister cohesion may be completely abolished. However, only a slight decrease in viability is observed for ord null flies, indicating that ORD function is not essential for cohesion during somatic mitosis. In addition, we do not observe perturbation of germ-line mitotic divisions in flies lacking ORD activity. Our analysis of weaker ord alleles suggests that ORD is required for proper centromeric cohesion after arm cohesion is released at the metaphase I/anaphase I transition. Finally, although meiotic cohesion is abolished in the ord null fly, chromosome loss is not appreciable. Therefore, ORD activity appears to promote centromeric cohesion during meiosis II but is not essential for kinetochore function during anaphase.

A source localization principle for linear shift-invariant systems with application to point optical and radioactive sources.

The source localization principle is an inequality between the means of scalar fields produced by different sources in any linear shift-invariant system. This principle is presented here as a pair of conditions (spatial and temporal) under which a point source produces a greater mean field over finite source-centered regions than all other sources. Biomedical applications involving point optical sources and radioactive sources are discussed.

Comparison of 810 nm and 1064 nm wavelengths for interstitial laser photocoagulation in rabbit brain.

BACKGROUND AND OBJECTIVE: This laboratory animal study is a comparison of Nd:YAG 1064 nm and diode 810 nm laser wavelengths in brain interstitial laser photocoagulation (ILP). Specific goals were to identify potential complications and physical characteristics of the thermal damage at both wavelengths prior to undertaking a clinical trial in humans. STUDY DESIGN/MATERIALS AND METHODS: A total of 41 ILP illuminations were performed in vivo in the brains of 33 anesthetized rabbits using plane-cut fiber tips implanted directly or through catheters, and diffusing fiber tips. Delivered powers ranged from 1.1 to 4.2 W. Exposures ranged from 300 to 900 s. Survival ranged from 0 to 48 h. Experiments were performed in animals with and without VX-2 brain tumors. RESULTS: Thermal damage from 1.1 W at 810 nm was similar to that from 1.6 W at 1064 nm, but more pronounced. With plane-cut fiber tips, there was a greater propensity for severe physical effects (smoke, charring, bubbling, surface damage) at 810 nm than at 1064 nm, yet hemorrhage, thrombosis and vapor dissemination were observed at both wavelengths, in both normal brain and tumor. CONCLUSIONS: For ILP in brain, 1064 nm may be better suited than 810 nm, although both are questionable with plane-cut-fiber tips. Compactness and portability may be the only valid reasons for using laser diodes operating around 810 nm. At 1064 nm, the power delivered from plane-cut fiber tips should be less than 1.5 W, necessitating long exposures, or else an open catheter should be used. Fiber tips with distributed emission may be preferred, provided structural integrity can be maintained.

Identification of ORD, a Drosophila protein essential for sister chromatid cohesion.

Attachment between the sister chromatids is required for proper chromosome segregation in meiosis and mitosis, but its molecular basis is not understood. Mutations in the Drosophila ord gene result in premature sister chromatid separation in meiosis, indicating that the product of this gene is necessary for sister chromatid cohesion. We isolated the ord gene and found that it encodes a novel 55 kDa protein. Some of the ord mutations exhibit unusual complementation properties, termed negative complementation, in which particular alleles poison the activity of another allele. Negative complementation predicts that protein-protein interactions are critical for ORD function. The position and nature of these unusual ord mutations demonstrate that the C-terminal half of ORD is essential for sister chromatid cohesion and suggest that it mediates protein binding.

Investigations of large vessel cooling during interstitial laser heating.

Interstitial laser heating of tissues is influenced by blood flow in the treatment region. Temperature gradients around large blood vessels may result in local underheating of tissues. A three-dimensional, time-dependent finite difference model of interstitial laser heating around large vessels is presented. A thermal conduction model was developed using a transport theory approximation for the energy distribution from an optical line source. Calculated transient temperature profiles and temperature reductions around 0.144 and 0.400 cm diam vessels show qualitative agreement with those measured in a series of tissue phantom studies. Experiments and calculations for a large vessel located approximately 1.0 cm from the optical source indicate that temperature reductions are less than 1 degree C at distances greater than approximately 1.0 cm from the vessel surface. The model also indicates that significant reductions in the extent of a thermal coagulation boundary can occur if a large vessel is situated inside the normal coagulation zone.

Basic optothermal diffusion theory for interstitial laser photocoagulation.

A theoretical basis for interstitial laser photocoagulation (ILP) practiced with point-emitting fiber tips has been established by solving the bioheat transfer equation, using basic Green's function methods, for steady and instantaneous point sources of both optical energy and direct heat. Three combination optical and thermal parameters have been identified that strongly influence temperature distributions during ILP. These are defined here as optothermal heat capacities and an optothermal diffusion length, all of which characterize how a thermal diffusion temperature profile is flattened and reduced when optical diffusion is added. Relevance and limitations of this theory for practical ILP are discussed. A useful result is a mathematical verification of previous empirical observations that point optical sources heat tissues less than point heat sources of the same power. A comparison of normalized theoretical temperature transients with published measurements suggests that in normal liver, blood perfusion cooling may exceed thermal conduction by a factor of 5.6 +/- 1.7.

Hepatic interstitial laser photocoagulation. An investigation of the relationship between acute thermal lesions and their sonographic images.

OBJECTIVES: The relationship between hepatic interstitial laser photocoagulation (ILP) lesions and their acute ultrasound images was evaluated. In addition, the natural history of ILP lesions in normal pig liver was documented. METHODS: Eighteen pigs underwent laparotomy and ultrasound-monitored ILP. In part 1 of the study, 12 pigs each had four separate exposures (1.50 W for 60, 100, 300, and 500 seconds) and were divided into four groups according to when they were killed (0, 3, 7, and 21 days). In part 2 of the study, six pigs each had two sequential exposures (1.60 W for 1,000 and then 500 seconds) at separate hepatic sites. Survival time was 3 days. Necropsy and histologic examination were performed in all animals. In 0- and 3-day survivors, actual thermal lesions were compared with "early" (immediately after ILP) and "late" (1 hour after ILP) ultrasound images. RESULTS: In the 300-, 500-, and 1,000-second exposures of parts 1 and 2, thermal lesions were overestimated or approximated by early ultrasound and were underestimated or approximated by late ultrasound. Analysis of variance showed statistically significant differences between thermal lesions and their early and late ultrasound images (F = 18.6, P < .001, no interactions). Time-growth characteristics of ILP lesions were reasonably consistent on ultrasound; exceptions were identifiable 200 seconds into the exposure. In part 2, ultrasound changes were minimal in five of six 500-second (second sequential) technically satisfactory exposures. Thermal lesions were seen at necropsy. All lesions healed by formation of granulation tissue and collagen. CONCLUSIONS: During ILP, early ultrasound images frequently overestimate actual thermal lesions. Ultrasound-monitored ILP of tumors may be most effective if, on early ultrasound, echogenic changes extend beyond the tumor margins. Late ultrasound images underestimate or approximate thermal lesions. Their value in clinical ILP should be investigated. It is unclear why ultrasound images of proven thermal lesions were not seen during 5 of 6 otherwise satisfactory 500-second ILP exposures performed immediately after 1,000-second exposures.

Hepatic interstitial laser photocoagulation: demonstration and possible clinical importance of intravascular gas.

PURPOSE: To investigate gas formation during hepatic interstitial laser photocoagulation (ILP). MATERIALS AND METHODS: In vitro, ILP was performed with a neodymium yttrium aluminum garnet laser on a piece of liver in a water bath. In vivo, nine pigs underwent 24 ultrasound (US)-monitored ILP procedures. Fiber tips were more than 1 cm from (n = 16) or adjacent to (n = 8) intrahepatic veins. The gas production seen on US images was graded on a scale of 0 to 4. Precordial Doppler US was performed in all cases. RESULTS: In vitro, smoke bubbles emanated from the vessels during ILP. In vivo, US showed intravascular gas production during nine of 15 exposures of at least 500 seconds duration. Gas production scores of 2 or more were recorded for nine exposures. Intracardiac gas was identified on eight precordial Doppler US recordings. All animals survived. CONCLUSION: Gas was detected in the heart during some ILP exposures. Patients with a probe-patent foramen ovale (24% prevalence) could be at risk for paradoxic air embolus during ILP.

Dependence of laser photocoagulation on interstitial delivery parameters.

Photocoagulation was performed ex vivo between tissue slabs by delivering continuous-wave laser energy from an optical fiber either directly, or by depositing the energy into a 2.4 mm diameter steel sphere at the fiber tip. The dependence of photocoagulation lesions on the following variables was assessed: (1) energy source: Nd:YAG-532 nm, 1,064 nm +/- steel sphere, (2) tissue type: porcine muscle (light), bovine muscle (dark), (3) delivered power: P = 1.5-3.0 W (porcine), 1.0-2.5 W (bovine), (4) exposure duration: T = 300-1500 s. The resulting cross-sectional photocoagulation lesions are summarized as follows: 532 nm: elongated; central charring in all cases; 1,064 nm: circular; central charring only in bovine for P > or = 2.0 W, T > or = 500 s; sphere: circular; central charring in bovine for P > or = 1.5 W and porcine for P > or = 2.0 W. These experiments suggest photocoagulation lesion size decreases as optical penetration increases. The results indicate that interstitial laser photocoagulation lesions > 10 mm diameter can be made without charring in both lightly and heavily pigmented tissues ex vivo by delivering 1,064 nm laser energy at sufficiently low power for at least 1,000 s from well-polished optical fibers.

Selecting source locations in multifiber interstitial laser photocoagulation.

A theoretical basis and practical algorithm are described for selecting optimal fiber source locations in multifiber interstitial laser photocoagulation (ILP). By analyzing the shape of overlapping coagulation patterns, the optimal separation of adjacent point heat sources inside a flat target volume boundary is shown to be d* = 2 square root of 2rc, where rc is the coagulation radius of a single source. Against a curved boundary, the algorithm specifies how d* should be altered. To assess the validity of the theory, ILP was conducted in bovine muscle by delivering 1064 nm laser energy through two or four plane-cut optical fibers simultaneously. Delivered power, exposure duration, and source separation were varied. The observed coagulation patterns matched the theory-predicted patterns at delivered powers of 1.60W and 1.85W, but not at 2.40W. Also, the experiments indicate that reciprocity of delivered power and exposure duration is invalid for ILP.

Comparison of magnetic resonance images and the histopathological findings of lesions induced by interstitial laser photocoagulation in the brain.

Interest has developed in using magnetic resonance (MR) imaging to monitor the volume of tissue destroyed by interstitial laser photocoagulation (ILP). In these experiments, ILP was induced in the normal brains of 9 anesthetized cats by delivering 1.5 W of continuous-wave Nd:YAG laser energy (1,064 nm) from a single 400-microns core optical fiber for 1,000 s. The irradiations were monitored using proton spin-echo MR imaging during and immediately after ILP and at postirradiation survival times of 2, 5, and 14 days. At 2 days postirradiation, the necrotic thermal lesion consisted of a central cavity surrounded by 2 concentric zones of coagulative necrosis, one dense and the other dispersed. The lesion shrank and the zonal appearance became less obvious over the 14 day survival period. An enhancing halo on contrast-enhanced T1-weighted images acquired immediately postirradiation best approximated the total lesion diameter at 2 days. These images also indicated that the volume of tissue destroyed during ILP corresponded better to the necrotic volume determined at 2 days than at 5 days and 14 days postirradiation. T2-weighted images acquired during and immediately after ILP consistently underestimated the total lesion diameter at 2 days.

Sonographic changes during hepatic interstitial laser photocoagulation. An investigation of three optical fiber tips.

RATIONALE AND OBJECTIVES: Interstitial laser photocoagulation (ILP) destroys tumors thermally, using laser energy delivered from implanted optical fibers. The objectives of the study are to identify a fiber tip/delivered energy combination which produces lesions of useful size, visible on ultrasound (US) during ILP, and to compare ILP lesions and their US images. METHODS: Hepatic ILP was performed at laparotomy in six pigs, using three different fiber tips (cylindrical diffusing, spherical diffusing, plane-cut). US images were obtained during ILP, immediately after ("early" images), and before the animals were killed (2-2.5 hours, "late" images). Actual lesions were assessed histopathologically. RESULTS: Few US changes were seen around cylindrical diffusing and spherical diffusing tips until tip destruction. Plane-cut tips, at 1.5 to 2.0 W, produced prominent US images of the 1- to 2-cm thermal lesions. Early images tended to overestimate necrosis. Late images approximated necrosis. CONCLUSION: For US-controlled ILP, plane-cut tips are better than currently available cylindrical diffusing or spherical diffusing tips. Lesion image growth periods might enable control of lesion size. Further studies are needed to determine the consistency of the described relationship between lesion images and actual lesions.

Interstitial laser photocoagulation: Nd:YAG 1064 nm optical fiber source compared to point heat source.

Interstitial laser photocoagulation (ILP) was performed in vitro in lean bovine and chicken muscle by delivering 1.6 W of continuous-wave Nd:YAG laser energy (1064 nm) from a 400-microns core optical fiber for 300s. The resulting thermal coagulation lesion was consistently larger when the delivered energy was deposited into a small steel sphere than when it was delivered freely into the tissue. Mathematical modelling confirms this result. This preliminary study suggests that a point heat source produces a larger volume of thermal coagulation than a point optical source (1064 nm) delivering the same power.

Magnetic resonance imaging of interstitial laser photocoagulation in brain.

Magnetic resonance (MR) imaging can be used to monitor the development of thermal lesions induced in tissue using interstitial laser photocoagulation (ILP). A potential application for ILP is the treatment of surgically inaccessible brain tumors. For the successful clinical application of MR-monitored ILP, it is necessary to relate MR images of ILP lesions to the actual induced lesions. In this preliminary study we performed ILP in the normal brains of anesthetized cats by delivering interstitially 1.0, 1.5, and 2.0 W of continuous-wave Nd:YAG laser energy (1,064 nm) for 1,000 s via a plane-cut 400 microns core optical fiber. At 48 h post-irradiation the lesions consisted of four sharply demarcated concentric zones of thermal damage. Lesion diameter increased linearly with delivered power. T2-weighted proton spin-echo images acquired during ILP showed a region of complete or near signal loss that underestimated the actual lesion at 48 h. Gadolinium-enhanced T1-weighted spin-echo images acquired immediately post-irradiation showed the actual lesion precisely.