Using 3D image registration to maximize the reproducibility of longitudinal bone strength assessment by HR-pQCT and finite element analysis.

We developed and validated a finite element (FE) approach for longitudinal high-resolution peripheral quantitative computed tomography (HR-pQCT) studies using 3D image registration to account for misalignment between images. This reduced variability in longitudinal FE estimates and improved our ability to measure in vivo changes in HR-pQCT studies of bone strength. INTRODUCTION: We developed and validated a finite element (FE) approach for longitudinal high-resolution peripheral quantitative computed tomography (HR-pQCT) studies using 3D rigid-body registration (3DR) to maximize reproducibility by accounting for misalignment between images. METHODS: In our proposed approach, we used the full common bone volume defined by 3DR to estimate standard FE parameters. Using standard HR-pQCT imaging protocols, we validated the 3DR approach with ex vivo samples of the distal radius (n = 10, four repeat scans) by assessing whether 3DR can reduce measurement variability from repositioning error. We used in vivo data (n = 40, five longitudinal scans) to assess the sensitivity of 3DR to detect changes in bone strength at the distal radius by the standard deviation of the rate of change (σ), where the ideal value of σ is minimized to define true change. FE estimates by 3DR were compared to estimates by no registration (NR) and slice-matching (SM). RESULTS: Group-wise comparisons of ex vivo variation (CVRMS, %) found that FE measurement precision was improved by SM (CVRMS < 0.80%) and 3DR (CVRMS < 0.62%) compared to NR (CVRMS~2%), and 3DR was advantageous as repositioning error increased. Longitudinal in vivo reproducibility was minimized by 3DR for failure load estimates (σ = 0.008 kN/month). CONCLUSION: Although 3D registration cannot negate motion artifacts, it plays an important role in detecting and reducing variability in FE estimates for longitudinal HR-pQCT data and is well suited for estimating effects of interventions in in vivo longitudinal studies of bone strength.

Longitudinal bone microarchitectural changes are best detected using image registration.

Longitudinal studies of bone using high-resolution medical imaging may result in non-physiological measurements of longitudinal changes. In this study, we determined that three-dimensional image processing techniques best capture realistic longitudinal changes in bone density and should therefore be used with high-resolution imaging when studying bone changes over time. INTRODUCTION: The purpose of this study was to determine which longitudinal analysis technique (no registration (NR), slice-match (SM) registration, or three-dimensional registration (3DR)) produced the most realistic longitudinal changes in a 3-year study of bone density and structure using high-resolution peripheral quantitative computed tomography (HR-pQCT). METHODS: We assessed HR-pQCT scans of the distal radius and tibia for men and women (N = 40) aged 55-70 years at baseline and 6, 12, 24, and 36 months. To evaluate which longitudinal analysis technique (NR, SM, or 3DR) best captured physiologically reasonable 3-year changes, we calculated the standard deviation of the absolute rate of change in each bone parameter. The data were compared between longitudinal analysis techniques using repeated measures ANOVA and post hoc analysis. RESULTS: As expected, both SM and 3DR better captured physiological longitudinal changes than NR. At the tibia, there were no differences between SM and 3DR; however, at the radius where precision was lower, 3DR produced better results for total bone mineral density. CONCLUSIONS: At least SM or 3DR should be implemented in longitudinal studies using HR-pQCT. 3DR is preferable, particularly at the radius, to ensure that physiological changes in bone density are observed.

Basedow paraplegia: A possible misnomer.

Thyrotoxic myopathy frequently occurs in clinical practice; however, the association of hyperthyroidism with a flaccid, areflexic paraplegia, so-called Basedow paraplegia, appears to represent a controversial and doubtful entity. An 18-year-old female with undiagnosed and untreated Graves' disease presented with acute onset of global weakness predominantly in the lower limbs, but also affecting the upper limbs. The weakness was accompanied by hypotonia and areflexia. Clinically, the patient had a goitre and signs of thyroid ocular disease. Laboratory testing confirmed the presence of hyperthyroidism, and thyroid-stimulating hormone receptor antibodies were positive. The cerebrospinal fluid protein level was raised. The electroneuronographic and needle examinations were compatible with a clear denervation process, such as acute motor axonal neuropathy, a variant of Guillain-Barre syndrome. Intravenous immunoglobulin therapy, carbimazole and propranolol were administered. The occurrence of hyperthyroidism with a flaccid, areflexic paraplegia appears to represent more of a fortuitous than a causative association. It is important to consider and treat other causes, such as acute idiopathic polyneuritis.

Plasma cytokine levels and human papillomavirus infection at the cervix in rural Nigerian women.

INTRODUCTION: We conducted a study to test the hypothesis that systemic dysregulation of Th1/Th2 cytokine levels was associated with detection of carcinogenic or overall human papillomavirus (HPV) at the cervix among 964 women residing in a rural village in Nigeria. METHODS: Levels in plasma were measured for 19 cytokines, including Th1-like cytokines IL-2, IL-12 (p40), TNF-a, IFN-g; Th2-like cytokines IL-4, IL-5, IL-6, IL-10, IL-13; innate/inflammation cytokines IL-1a, IL-1b, IL-8, eotaxin, MCP-1, MIP-1a, and IL-7; and cell development cytokines G-CSF, VEGF, and IL-17. Analysis was restricted to 5 cytokines, TNF-α (Th1), IL-8 (Th2), eotaxin and MCP-1 (innate/inflammation), and G-CSF (cell development), whose levels were detected in 80% or more of the samples measured as well as had a coefficient of variation of <30%. RESULTS: Strong correlations were noted between levels of eotaxin and TNF-α (r=0.75), IL-8 and MCP-1 (r=0.60), eotaxin and G-CSF (r=0.44), and G-CSF and IFN-γ (r=0.43). Detection of carcinogenic or non-carcinogenic HPV DNA was unrelated to cytokine levels, except for levels of eotaxin and TNF-α, which were inversely correlated, albeit weakly, with detection of any carcinogenic HPV (P=0.048 and P=0.067, respectively). In analyses stratified by age group, levels of eotaxin were inversely correlated with detection of any HPV DNA (P=0.026) and carcinogenic HPV (P=0.042) in older, but not younger, women. CONCLUSIONS: Our results do not support the hypothesis of association between systemic cytokine dysregulation and detection of HPV at the cervix in Nigerian women, but subgroup analyses raise questions about inverse associations between eotaxin and TNF-α in older women.

Circulating levels of immune and inflammatory markers and long versus short survival in early-stage lung cancer.

BACKGROUND: Some patients diagnosed with early-stage lung cancer and treated according to standard care survive for only a short period of time, while others survive for years for reasons that are not well understood. Associations between markers of inflammation and survival from lung cancer have been observed. MATERIALS AND METHODS: Here, we investigate whether circulating levels of 77 inflammatory markers are associated with long versus short survival in stage I and II lung cancer. Patients who had survived either <79 weeks (~1.5 years) (short survivors, SS) or >156 weeks (3 years) (long survivors, LS) were selected from a retrospective population-based study. Logistic regression was used to calculate adjusted odds ratios (ORs) and corresponding 95% confidence intervals (CIs). The false discovery rate was calculated to adjust for multiple testing. RESULTS: A total of 157 LS and 84 SS were included in this analysis. Thirteen markers had adjusted OR on the order of 2- to 5-fold when comparing the upper and lower quartiles with regard to the odds of short survival versus long. Chemokine CCL15 [chemokine (C-C motif) ligand 15] was the most significant marker associated with increased odds of short survival (ORs = 4.93; 95% CI 1.90-12.8; q-value: 0.042). Smoking and chronic obstructive pulmonary disease were not associated with marker levels. CONCLUSIONS: Our results provide some evidence that deregulation of inflammatory responses may play a role in the survival of early-stage lung cancer. These findings will require confirmation in future studies.

Thermometers for low temperature Magic Angle Spinning NMR.

The measurement of temperature in a Magic Angle Spinning NMR probe in the temperature range 85-300K is discussed. It is shown that the shift of the (119)Sn resonance of Sm(2)Sn(2)O(7) makes a good thermometer with shift being given by delta=223 - 9.54x10(4)/Tppm and a potential precision of better than 0.5K over the entire temperature range. The sensitivity is such (e.g. 4.2ppm/K at 150K) that small temperature gradients across the sample can readily be measured. Furthermore, since the spin-lattice relaxation time is very short, measurements can be made in approximately 1s enabling relatively rapid temperature changes to be followed. Values for the chemical shift of (207)Pb in Pb(NO(3))(2) down to approximately 85K are also presented. Although the (207)Pb shift variation is approximately linear near room temperature (we find a slope 0.725+/-0.002ppm/K over the range 293-153K), it clearly deviates from linearity below approximately 130K.

Analogues of Y27632 increase gap junction communication and suppress the formation of transformed NIH3T3 colonies.

BACKGROUND: Constitutive activation of RhoA-dependent RhoA kinase (ROCK) signalling is known to promote cellular transformation and the ROCK inhibitor Y-27632 has the ability to suppress focus formation of RhoA transformed NIH3T3 cells. METHODS: Sixty-four novel structural analogues of Y27632 were synthesised and tested for their ability to persistently inhibit the transformation of NIH3T3 cells by Rho guanidine exchange factor 16 (ARHGEF16) or Ras. In vitro kinase inhibitor profiling, co-culture of transformed cells with non-transformed cells and a novel Lucifer yellow/PKH67 dye transfer method were used to investigate their mode of action. RESULTS: Four Y27632 analogues inhibited transformed focus formation that persisted when the compound was withdrawn. No toxicity was observed against either transformed or non-transformed cells and the effect was dependent on co-culture of these two cell types. In vitro kinase inhibitor profiling indicated that these compounds had reduced activity against ROCK compared with Y27632, targeting instead Aurora A (AURKA), p38 (MAPK14) and Hgk (MAP4K4). Dye transfer analysis showed they increased gap junction intercellular communication (GJIC) between transformed and non-transformed cells. CONCLUSIONS: These data are the first to suggest that transient blockade of specific kinases can induce a persistent inhibition of non-contact inhibited transformed colony formation and can also remove pre-formed colonies. These effects could potentially be mediated by the observed increase in GJIC between transformed and non-transformed cells. Selection of kinase inhibitors with this property may thus provide a novel strategy for cancer chemoprevention.

QuadFit--a new cross-platform computer program for simulation of NMR line shapes from solids with distributions of interaction parameters.

A new Java computer program called QuadFit has been written to simulate NMR line shapes from solid materials. The program takes into account the major interactions, with a key feature that distributions of isotropic chemical shift and quadrupolar interaction parameters can be calculated, which are often encountered in amorphous and disordered materials. The quadrupolar interaction can be simulated for all the transitions for both half-integer and integer spins. The utility of the program is demonstrated with examples of (27)Al (nuclear spin I=5/2) in an atomically disordered aluminoborate mullite, (65)Cu (I=3/2) in CuInSe(2) and (10)B (I=3) in amorphous B(2)O(3). The program has good cross-platform compatibility and is written for high stability. The program has been designed with an easy to use graphical interface. It can be run efficiently on any reasonably powerful PC and is freely available from the Warwick website (http://go.warwick.ac.uk/quadfit).

A natural abundance (77)Se solid-state NMR study of inorganic compounds.

Various inorganic selenium-based compounds were analysed by (77)Se solid-state NMR, and a distinct difference in chemical shift ranges for compounds where selenium is present as selenide (Se(2-)) ionically and covalently bonded systems was observed. The selenides exhibit a shift range of approximately -700 to -100ppm, as opposed to 700 to 1600ppm for the compounds where there tends to be more direct covalent bonding to the selenium. The anisotropic hyperfine shift observed in NbSe(2) is shown to be axially symmetric, where the H(11) component is found to be normal to the Se3-trigonal plane.

Long-term effects of a plant-based dietary portfolio of cholesterol-lowering foods on blood pressure.

OBJECTIVE: To determine the effect on blood pressure of dietary advice to consume a combination of plant-based cholesterol-lowering foods (dietary portfolio). METHODS: For 1 year, 66 hyperlipidemic subjects were prescribed diets high in plant sterols (1.0 g/1000 kcal), soy protein (22.5 g/1000 kcal), viscous fibers (10 g/1000 kcal) and almonds (22.5 g/1000 kcal). There was no control group. Seven-day diet record, blood pressure and body weight were monitored initially monthly and later at 2-monthly intervals throughout the study. RESULTS: Fifty subjects completed the 1-year study. When the last observation was carried forward for non-completers (n=9) or those who changed their blood pressure medications (n=7), a small mean reduction was seen in body weight 0.7+/-0.3 kg (P=0.036). The corresponding reductions from baseline in systolic and diastolic blood pressure at 1 year (n=66 subjects) were -4.2+/-1.3 mm Hg (P=0.002) and -2.3+/-0.7 mm Hg (P=0.001), respectively. Blood pressure reductions occurred within the first 2 weeks, with stable blood pressures 6 weeks before and 4 weeks after starting the diet. Diastolic blood pressure reduction was significantly related to weight change (r=0.30, n=50, P=0.036). Only compliance with almond intake advice related to blood pressure reduction (systolic: r=-0.34, n=50, P=0.017; diastolic: r=-0.29, n=50, P=0.041). CONCLUSIONS: A dietary portfolio of plant-based cholesterol-lowering foods reduced blood pressure significantly, related to almond intake. The dietary portfolio approach of combining a range of cholesterol-lowering plant foods may benefit cardiovascular disease risk both by reducing serum lipids and also blood pressure.

The concept of correlated progression as the basis of a model for the evolutionary origin of major new taxa.

Understanding the evolutionary processes responsible for the long treks through morphospace associated with the origin of new higher taxa is hampered by the lack of a realistic and usable model that accounts for long-term phenotypic evolvability. The systems-related concept of correlated progression, in which all the traits are functionally linked and so constrained to evolve by small increments at a time in parallel with each other, provides the basis for such a model. Implications for the process of evolution at high taxonomic level are that: the evolving traits must be considered together as a system, and the exact sequence of incremental changes in characters is indeterminable; there are no identifiable key innovations; selection acts on the phenotype as a whole rather than on individual traits; and the selection force is therefore multidimensional. Application of the model to the pattern of evolution of traits and trait states as revealed by the fossil record of the stem groups of such taxa as mammals, turtles and tetrapods generates realistic testable hypotheses about how such groups evolved.

Effect on hematologic risk factors for coronary heart disease of a cholesterol reducing diet.

BACKGROUND: A dietary portfolio of cholesterol-lowering ingredients has proved effective in reducing serum cholesterol. However, it is not known whether this dietary combination will also affect hematologic risk factors for coronary heart disease (CHD). Reductions in hematocrit and polymorphonuclear leukocytes have been reported to improve cardiovascular risk. We, therefore, report changes in hematological indices, which have been linked to cardiovascular health, in a 1-year assessment of subjects taking an effective dietary combination (portfolio) of cholesterol-lowering foods. METHODS: For 12 months, 66 hyperlipidemic subjects were prescribed diets high in plant sterols (1.0 g/1000 kcal), soy protein (22.5 g/1000 kcal), viscous fibers (10 g/1000 kcal) and almonds (23 g/1000 kcal). Fifty-five subjects completed the study. RESULTS: Over the 1 year, data on completers indicated small but significant reductions in hemoglobin (-1.5+/-0.6 g/l, P=0.013), hematocrit (-0.007+/-0.002 l/l, P<0.001), red cell number (-0.07+/-0.02 10(9)/l, P<0.001) and neutrophils (-0.34+/-0.13 10(9)/l, P=0.014). Mean platelet volume was also increased (0.16+/-0.07 fl, P=0.033). The increase in red cell osmotic fragility (0.05+/-0.03 g/l, P=0.107) did not reach significance. CONCLUSIONS: These small changes in hematological indices after a cholesterol-lowering diet are in the direction, which would be predicted to reduce CHD risk. Further research is needed to clarify whether the changes observed will contribute directly or indirectly to cardiovascular benefits beyond those expected from reductions previously seen in serum lipids and blood pressure.

The origin and early radiation of the therapsid mammal-like reptiles: a palaeobiological hypothesis.

The replacement of the basal synapsid pelycosaurs by the more 'mammal-like' therapsids in the Permian was an important event in the history of tetrapods because it initiated the eventual transition to the mammals. It is also an example of taxon replacement in the fossil record that is unusually amenable to explanation, based on a combination of analysis of the biological significance of the inferred character changes, with the stratigraphic, palaeogeographic and palaeoecological circumstances of the time. An hypothesis is presented in which the origin of the therapsids resulted from a correlated progression of character evolution leading to higher levels of metabolic activity and homeostatic regulation of the body. It was a response to the availability of a seasonally arid, savanna-like biome. The subsequent explosive radiation of therapsids was associated with habitat expansion made possible by the Mid-Permian development of geographical continuity between that biome and the temperate biomes. The final extinction of the pelycosaurs was a case of incumbent replacement by the new therapsid lineages.

Functional trade-offs in the limb bones of dogs selected for running versus fighting.

The physical demands of rapid and economical running differ from the demands of fighting in ways that may prevent the simultaneous evolution of optimal performance in these two behaviors. Here, we test an hypothesis of functional trade-off in limb bones by measuring mechanical properties of limb bones in two breeds of domestic dog (Canis lupus familiaris L.) that have undergone intense artificial selection for running (greyhound) and fighting (pit bull) performance. The bones were loaded to fracture in three-point static bending. To correct for the effect of shear, we estimated the shear stress in the cross section and added energy due to shear stress to the tensile energy. The proximal limb bones of the pit bulls differed from those of the greyhounds in having relatively larger second moments of area of mid-diaphyseal cross sections and in having more circular cross-sectional shape. The pit bulls exhibited lower stresses at yield, had lower elastic moduli and failed at much higher levels of work. The stiffness of the tissue of the humerus, radius, femur and tibia was 1.5-2.4-fold greater in the greyhounds than in the pit bulls. These bones from the pit bulls absorbed 1.9-2.6-fold more energy before failure than did those of the greyhounds. These differences between breeds were not observed in the long bones of the feet, metacarpals and metatarsals. Nevertheless, the results of this analysis suggest that selection for high-speed running is associated with the evolution of relatively stiff, brittle limb bones, whereas selection for fighting performance leads to the evolution of limb bones with relatively high resistance to failure.

Solid-state NMR characterisation of the thermal transformation of a Hungarian white illite.

(1)H, (27)Al, (29)Si and (39)K solid-state NMR are reported from a Hungarian illite 2:1 clay for samples heated up 1600 degrees C. This single-phase sample has a small amount of aluminium substitution in the silica layer and very low iron-content ( approximately 0.4wt%). Thermal analysis shows several events that can be related to features in the NMR spectra, and hence changes in the atomic scale structure. As dehydroxylation occurs there is increasing AlO(4) and AlO(5)-contents. The silica and gibbsite layers become increasingly separated as the dehydroxylation progresses. Between 900 and 1000 degrees C the silica layer forms a potassium aluminosilicate glass. The gibbsite-layer forms spinel/gamma-Al(2)O(3) and some aluminium-rich mullite. Then on heating to 1600 degrees C changes in the (29)Si and (27)Al MAS NMR spectra are consistent with the aluminosilicate glass increasing its aluminium-content, the amount of mullite increasing probably with its silicon-content also increasing, and some alpha-Al(2)O(3) forming.

Compensating for the effect of inlet gas temperature on heated humidifier performance.

The humidity output of heated humidifiers may be compromised by inlet gas temperatures exceeding approximately 26 degrees C, with humidity dropping below the recommended levels for intubated patients. A new version of the Fisher & Paykel MR850 humidifier claims to deal with this problem by offering a humidity compensation option. The present study tested this feature by measuring humidity output using the gravimetric method and a hygrometer at different inlet gas temperatures (16.6 degrees C to 40.0 degrees C) with compensation on and off. It was found that the compensation is effective in maintaining humidity levels despite high inlet gas temperatures.

Tetracycline-inducible transgene expression mediated by a single AAV vector.

Regulated gene delivery systems are usually made of two elements: an inducible promoter and a transactivator. In order to optimize gene delivery and regulation, a single viral vector ensuring adequate stoichiometry of the two elements is required. However, efficient regulation is hampered by interferences between the inducible promoter and (i) the promoter used to express the transactivator and/or (ii) promoter/enhancer elements present in the viral vector backbone. We describe a single AAV vector in which transcription of both the reverse tetracycline transactivator (rtTA) and the transgene is initiated from a bidirectional tetracycline-responsive promoter and terminated at bidirectional SV40 polyadenylation sites flanking both ITRs. Up to 50-fold induction of gene expression in human tumor cell lines and 100-fold in primary cultures of rat Schwann cells was demonstrated. In addition an 80-fold induction in vivo in the rat brain has been obtained. In vitro, the autoregulatory vector exhibits an induced expression level superior to that obtained using the constitutive CMV promoter. Although extinction of the transgene after removal of tetracycline was rapid (less than 3 days), inducibility after addition of tetracycline was slow (about 14 days). This kinetics is suitable for therapeutic gene expression in slowly progressive diseases while allowing rapid switch-off in case of undesirable effects. As compared to previously described autoregulatory tet-repressible (tetOFF) AAV vectors, the tet-inducible (tetON) vector prevents chronic antibiotic administration in the uninduced state.

Induction of necrotic tumor cell death by TRAIL/Apo-2L.

A great deal of enthusiasm is being generated for TRAIL (TNF-related apoptosis-inducing ligand)/Apo-2L as a tumor therapeutic agent because it is cytotoxic to a variety of tumor cell types but not normal cells. Moreover, it is well documented that TRAIL/Apo-2L-induced tumor cell death is a caspase-dependent apoptotic process. Through the use of a transfected cell line expressing murine TRAIL/Apo-2L and a recombinant adenovirus encoding the murine TRAIL/Apo-2L cDNA (Ad5-mTRAIL) against two murine tumor cell lines [TRAMP-C2 (prostate adenocarcinoma) and Renca (renal adenocarcinoma)], we found that mTRAIL/Apo-2L also can kill tumor cells by inducing necrosis. Specifically, we observed the default method of mTRAIL/Apo-2L-induced death in TRAMP-C2 cells was via a necrotic process, characterized by the complete lack of an annexin V(+)/PI(-) population, SAPK/JNK phosphorylation, caspase activation, Bid cleavage, or cytochrome c release. Moreover, the inclusion of zVAD-fmk, an inhibitor of caspase activation, markedly enhanced mTRAIL/Apo-2L-mediated killing of TRAMP-C2. In contrast, apoptosis was induced in TRAMP-C2 using TNF, as measured by the criteria listed above, as was Renca by mTRAIL/Apo-2L. These results demonstrate the natural occurrence of both TRAIL/Apo-2L-induced apoptotic and necrotic signaling mechanisms within tumor cells.