Doyle L. Hawkins

Regression of Lung Cancer by Hypoxia-Sensitizing Ruthenium Polypyridyl Complexes

The ruthenium (II) polypyridyl complexes (RPC), Δ-[(phen)2Ru(tatpp)]Cl2 (Δ-[3]Cl2) and ΔΔ-[(phen)2Ru(tatpp)Ru(phen)2]Cl4 (ΔΔ-[4]Cl4, are a new generation of metal-based antitumor agents. These RPCs bind DNA via intercalation of the tatpp ligand, which itself is redox-active and is easily reduced at biologically relevant potentials. We have previously shown that RPC 44+ cleaves DNA when reduced by glutathione to a radical species and that this DNA cleavage is potentiated under hypoxic conditions in vitro. Here, we show that 32+ also exhibits free radical–mediated DNA cleavage in vitro and that 32+ and 44+ both exhibit selective cytotoxicity toward cultured malignant cell lines and marked inhibition of tumor growth in vivo. The murine acute toxicity of RPCs 32+ and 44+ (maximum tolerable doses ∼ 65 μmol/kg) is comparable with that for cisplatin (LD50 ∼ 57 μmol/kg), but unlike cisplatin, RPCs are generally cleared from the body unchanged via renal excretion without appreciable metabolism or nephrotoxic side effects. RPCs 32+ and 44+ are shown to suppress growth of human non–small cell lung carcinoma (∼83%), show potentiated cytotoxicity in vitro under hypoxic conditions, and induce apoptosis through both intrinsic and extrinsic pathways. The novel hypoxia-enhanced DNA cleavage activity and biologic activity suggest a promising new anticancer pharmacophore based on metal complexes with aromatic ligands that are easily reduced at biologically accessible potentials. Mol Cancer Ther; 12(5); 643–53. ©2013 AACR.

Intraoperative bile duct visualization using near-infrared hyperspectral video imaging

BACKGROUND Current methodologies for imaging the biliary system during cholecystectomy are cumbersome and do not eliminate the risk of bile duct injury. We describe an approach to intraoperative biliary imaging that will enable surgeons to see through the hepatoduodenal ligament and visualize the anteriorly placed biliary system. METHODS A laparoscopic-capable, near-infrared, hyperspectral imaging system was built. Reflected light passes through a liquid crystal filter that is continuously tunable in the near-infrared spectrum (650-1,100 nm). Spectroscopic image data are collected from laparoscopic surgery images onto array detectors formatted into a 3-dimensional hyperspectral data cube having spatially resolved images in the x-y plane and wavelength data in the z plane. Deconvoluting and color-coding the spatial and spectral information provides an image representative of inherent chemical properties to the imaged tissue. RESULTS Images of porcine biliary structures were obtained. The common duct-reflected spectra displayed a characteristic lipid shoulder at 930 nm and a strong water peak at 970 nm. Venous structures had absorption peaks at 760 nm (deoxyhemoglobin), 800 nm (oxyhemoglobin), and 970 nm (water). Arterial vessels had absorption peaks at 800 nm and 970 nm that would be expected for oxyhemoglobin and water. CONCLUSIONS We have designed and constructed a device to significantly enhance intraoperative biliary imaging. This system should enable surgeons to see through the hepatoduodenal ligament and image the anteriorly placed biliary system without the need for dissection of the cystic duct, as is needed with intraoperative cholangiography. Because the biliary system can be seen before any dissection is performed, this dimensional imaging technology has the potential for eradicating bile duct injury.

Exploration of structure-antifouling relationships of capsaicin-like compounds that inhibit zebra mussel (Dreissena polymorpha) macrofouling.

Abstract Macrofouling of aquatic man-made structures by zebra mussels (Dreissena polymorpha) poses significant economic burdens on commercial freshwater shipping and facilities utilising raw water. The negative environmental impact of some current antifouling technologies has limited their use and prompted investigation of non-organometallic and non-oxidising antifoulants as possible environment-friendly alternatives. The plant-derived natural product capsaicin and 18 other compounds with one or more capsaicin-like structural features were tested for their potential to inhibit zebra mussel byssal attachment at a single high concentration of 30 μM. Of these, three compounds displaying the highest levels of attachment inhibition where selected for further concentration-response testing. This testing revealed that capsaicin (8-methyl-N-vanillyl-trans-6-nonenamide), N-vanillylnonanamide, and N-benzoylmonoethanolamine benzoate all inhibited byssal attachment with potency values (EC50) in the micromolar range. None of these compounds were lethal to adult specimens of the water flea, Daphnia magna, at concentrations that inhibited mussel byssal attachment.

Relationships among steroid hormone levels, vocal effort and body condition in an explosive-breeding toad

Mutually reinforcing hormone–behaviour relationships may drive temporal changes in steroid hormone levels and vocal effort in anuran amphibians. Recent models propose that chorus activity stimulates the production of androgens in signallers and receivers, thereby mediating an increase in vocal effort. The energetic demands associated with high vocal effort should, in turn, elevate circulating corticosterone levels to promote the mobilization of energy reserves. High threshold corticosterone levels, however, are expected to negatively affect androgen levels and, hence, vocal effort. Steroid hormone levels and vocal effort are thus expected to be low at the onset of chorus activity, increase over sequential nights of calling until peak levels are reached, and subsequently decline; energy reserves should show inverse temporal trends. To test these predictions, we examined temporal changes in androgen (dihydrotestosterone and testosterone) levels, corticosterone levels, vocal effort and body condition in Woodhouses toads, Bufo woodhousii . Contrary to model predictions, androgen levels, vocal effort and body condition remained relatively unchanged and circulating corticosterone levels tended to decrease within and across nights of chorus activity. Furthermore, although vocal effort was not correlated with circulating androgen levels, it was positively correlated with circulating corticosterone levels, supporting the prediction that high vocal effort promotes the production of corticosterone. Yet despite this correlation, poor body condition was not associated with high corticosterone levels, suggesting that factors other than energetic state influence corticosterone levels in B. woodhousii . We discuss alternative hypotheses regarding relationships among the measured parameters in this explosive-breeding species.

Detecting shifts in functions of multivariate location and covariance parameters

Abstract Procedures are studied for monitoring for shifts in functions of the mean vector or covariance matrix of an arbitrary multivariate distribution. The procedures provide the capabilities, among others, to monitor a linear function (e.g. a contrast) of the mean vector, or to monitor the covariance matrix for shifts in variances, correlations, generalized variance or for shifts away from the conditions of sphericity or intra-class correlation structure. Two sampling schemes are considered: the retrospective and sequential settings. Test statistics are quadratic forms in split-sample mean and covariance estimates. The asymptotic theory of U-statistics is used to control null error rates. A small Monte Carlo study investigates non-null power and reaction time. A total of eight procedures are studied.

Sex-biased gene expression in a ZW sex determination system.

Studies of the transcriptome have shown that a substantial fraction of interspecific differences in gene expression is the result of sex-biased gene expression. These results suggest that sex-dependent selection may be an important force in generating differences between species but to date all studies have focused on Drosophila. We examined a sample of the transcriptome in the gonads of two species of Xenopus to provide an additional test of how sex-biased gene expression may contribute to differences in gene expression between species. In contrast to Drosophila, Xenopus provides an example of a ZW system with morphologically indistinguishable sex chromosomes. About 81% of the transcriptome was differentially expressed between X. laevis and X. muelleri and there were more genes that were male-biased compared to the number of genes that were female-biased or non-sex-biased. While there were more genes up-regulated in males of Xenopus, the largest magnitude difference in expression between species occurred in female-biased genes, and male-biased genes were proportionally more abundant for the smallest changes in expression between species. Our results suggest that more genes have a role in the function of the testis compared to the ovary and female-biased genes play a principle role in expression divergence between species. These results differ from those in the Drosophila XY system in that more female-biased genes had >4-fold difference of expression between species than male-biased genes, suggesting that ZW sex chromosomes may facilitate enhanced gene expression divergence between species.

A test of the Energetics-Hormone Vocalization model in the green treefrog

Male courtship displays may be regulated by, and affect the production of, circulating hormones. The Energetics-Hormone Vocalization (EHV) model, for example, posits that interactions among chorusing male anuran amphibians stimulate androgen production that then mediates an increase in vocal effort. Increased vocal effort is expected to deplete energy reserves and increase glucocorticoid levels that, in turn, negatively affect androgen levels and vocalization. Androgen levels, glucocorticoid levels, and vocal effort are thus expected to increase across and within nights of chorus activity and should be positively correlated in calling males; energy reserves should decline temporally and be inversely related to glucocorticoid levels. We tested predictions of the EHV model in the green treefrog, Hyla cinerea. Consistent with the model, both testosterone and dihydrotestosterone levels increased across the breeding season in calling males. However, testosterone levels decreased and dihydrotestosterone levels did not change within nights of chorus activity, suggesting that chorusing behavior did not drive the seasonal elevation in androgens. Corticosterone (CORT) level remained relatively stable across the breeding season and decreased within nights of chorus activity, contrary to model predictions. Body condition, the proxy for energetic state, was inversely correlated with CORT level but discrepancies between model predictions and temporal patterns of CORT production arose because there was no evidence of a temporal decrease in body condition or increase in vocal effort. Moreover, androgen and CORT levels were not positively correlated with vocal effort. Additional ecological and physiological measures may be needed to support predictions of the EHV model.

Statistical considerations in bioequivalence of two area under the concentration–time curves obtained from serial sampling data

In this paper, we study the bioequivalence (BE) inference problem motivated by pharmacokinetic data that were collected using the serial sampling technique. In serial sampling designs, subjects are independently assigned to one of the two drugs; each subject can be sampled only once, and data are collected at K distinct timepoints from multiple subjects. We consider design and hypothesis testing for the parameter of interest: the area under the concentration–time curve (AUC). Decision rules in demonstrating BE were established using an equivalence test for either the ratio or logarithmic difference of two AUCs. The proposed t-test can deal with cases where two AUCs have unequal variances. To control for the type I error rate, the involved degrees-of-freedom were adjusted using Satterthwaites approximation. A power formula was derived to allow the determination of necessary sample sizes. Simulation results show that, when the two AUCs have unequal variances, the type I error rate is better controlled by the proposed method compared with a method that only handles equal variances. We also propose an unequal subject allocation method that improves the power relative to that of the equal and symmetric allocation. The methods are illustrated using practical examples.

Dorsal horn neuron response patterns to graded heat stimuli in the rat.

Sensory input from various receptors in the periphery first becomes integrated in the spinal cord dorsal horn. The response of the spinal cord dorsal horn neurons to mechanical stimuli are classified as low threshold, high threshold, and wide dynamic range neurons. However, the response pattern of deep dorsal horn cells to heat has not been well described. In this study, the response of the spinal cord dorsal horn neurons to graded heat stimuli were characterized in 147 neurons in rats by extracellular single cell recording. After a differentiable cell was identified, the Peltier heat stimulator was applied to the receptive field and the base temperature was set at 30 degrees C. The heat stimulus was delivered for 10 s from 37-51 degrees C in 2 degrees C increments, with an inter-stimulus interval of 30 s. Out of the 147 neurons, five statistically distinguishable response patterns were identified by latent class cluster analysis. It is concluded that variation of temperature may account for the observed results and indicate functionally different subsets of heat-responsive cells in the deep dorsal horn.

Truncated multiple inverse sampling in post-stratification

Multiple inverse sampling (MIS) is a sequential sampling procedure in a stratified population such that random samples are taken population-wide and continuously until a specified minimum number of observations are obtained in each stratum. The MIS procedure can be used in small sample post-stratification to solve the empty post-strata problem. However, the MIS procedure does not have control over the total sample size, in particular, when some of the stratum weights are small. A remedy for this drawback of MIS is to truncate the sampling procedure when the total sample size reaches a specified maximum number. The efficiency of the truncated MIS procedure is investigated and it is found that the truncated MIS is superior to the classical fixed sample size sampling procedure in post-stratification.