Terahertz in vivo imaging of human skin: Toward detection of abnormal skin pathologies.

Terahertz (THz) imaging has long held promise for skin cancer detection but has been hampered by the lack of practical technological implementation. In this article, we introduce a technique for discriminating several skin pathologies using a coherent THz confocal system based on a THz quantum cascade laser. High resolution in vivo THz images (with diffraction limited to the order of 100 µm) of several different lesion types were acquired and compared against one another using the amplitude and phase values. Our system successfully separated pathologies using a combination of phase and amplitude information and their respective surface textures. The large scan field (50 × 40 mm) of the system allows macroscopic visualization of several skin lesions in a single frame. Utilizing THz imaging for dermatological assessment of skin lesions offers substantial additional diagnostic value for clinicians. THz images contain information complementary to the information contained in the conventional digital images.

Improved predictive ability of climate-human-behaviour interactions with modifications to the COMFA outdoor energy budget model.

The purpose of this paper is to implement current and novel research techniques in human energy budget estimations to give more accurate and efficient application of models by a variety of users. Using the COMFA model, the conditioning level of an individual is incorporated into overall energy budget predictions, giving more realistic estimations of the metabolism experienced at various fitness levels. Through the use of VO(2) reserve estimates, errors are found when an elite athlete is modelled as an unconditioned or a conditioned individual, giving budgets underpredicted significantly by -173 and -123 W m(-2), respectively. Such underprediction can result in critical errors regarding heat stress, particularly in highly motivated individuals; thus this revision is critical for athletic individuals. A further improvement in the COMFA model involves improved adaptation of clothing insulation (I (cl)), as well clothing non-uniformity, with changing air temperature (T (a)) and metabolic activity (M (act)). Equivalent T (a) values (for I (cl) estimation) are calculated in order to lower the I (cl) value with increasing M (act) at equal T (a). Furthermore, threshold T (a) values are calculated to predict the point at which an individual will change from a uniform I (cl) to a segmented I (cl) (full ensemble to shorts and a T-shirt). Lastly, improved relative velocity (v (r)) estimates were found with a refined equation accounting for the degree angle of wind to body movement. Differences between the original and improved v (r) equations increased with higher wind and activity speeds, and as the wind to body angle moved away from 90°. Under moderate microclimate conditions, and wind from behind a person, the convective heat loss and skin temperature estimates were 47 W m(-2) and 1.7°C higher when using the improved v (r) equation. These model revisions improve the applicability and usability of the COMFA energy budget model for subjects performing physical activity in outdoor environments. Application is possible for other similar energy budget models, and within various urban and rural environments.

Methods for quantifying breast cancer-related lymphedema in patients undergoing a contralateral prophylactic mastectomy.

Patients treated for breast cancer are at risk of developing breast cancer-related lymphedema (BCRL). A significant proportion of patients treated for breast cancer are opting to undergo a contralateral prophylactic mastectomy (CPM). Currently, it remains unclear as to whether the relative volume change (RVC) equation may be used as an alternative to the weight adjusted change (WAC) equation to quantify BCRL in patients who undergo CPM. In order to simplify BCRL screening, our cohort of patients who underwent a CPM (n=310) was matched by BMI to a subset of patients who underwent unilateral breast surgery (n=310). Arm volume measurements were obtained via an optoelectronic perometer preoperatively, postoperatively, and in the follow-up setting every 6-12 months. The correlation of ipsilateral RVC and WAC values for those who underwent bilateral surgery was calculated (r=0.60). Contralateral WAC values for patients in both cohorts were compared, and there was no significant difference between the two distributions in variance (p=0.446). The RVC equation shows potential to be used to quantify ipsilateral postoperative arm volume changes for patients who undergo a CPM. However, a larger trial in which RVC and WAC values are prospectively assessed is needed.

Comparison of perometry-based volumetric arm measurements and bioimpedance spectroscopy for early identification of lymphedema in a prospectively-screened cohort of breast cancer patients.

Breast cancer-related lymphedema (BCRL) affects more than one in five women treated for breast cancer, and women remain at lifelong risk. Screening for BCRL is recommended by several national and international organizations for women at risk of BCRL, and multiple methods of objective screening measurement exist. The goal of this study was to compare the use of perometry and bioimpedance spectroscopy (BIS) for early identification of BCRL in a cohort of 138 prospectivelyscreened patients. At each screening visit, a patient's relative volume change (RVC) from perometer measurements and change in L-Dex from baseline (ΔL-Dex) using BIS was calculated. There was a negligible correlation between RVC and ΔL-Dex (r=0.195). Multiple thresholds of BCRL were examined: RVC ≥5% and ≥10% as well as and ΔL-Dex ≥6.5 and ≥10. While some patients developed an elevated RVC and ΔL-Dex, many demonstrated elevations in only one threshold category. Moreover, the majority of patients with RVC ≥5%, ΔL-Dex ≥6.5, or ΔL-Dex ≥10 regressed to non-elevated measurements without intervention. These findings suggest a role for combining multiple screening methods for early identification of BCRL; furthermore, BCRL diagnosis must incorporate patient symptoms and clinical evaluation with objective measurements obtained from techniques such as perometry and bioimpedance spectroscopy.

Reducing the potential for phlebotomy tourniquets to act as a reservoir for meticillin-resistant Staphylococcus aureus.

The contamination rate of phlebotomy tourniquets with meticillin-resistant Staphylococcus aureus (MRSA) was assessed, and it was determined whether this could be reduced by changes in practice or by the use of a physical barrier. Initially, the tourniquets of both preregistration house officers and phlebotomists were investigated, but as phlebotomists reported significantly more venepunctures daily, the trial continued solely with phlebotomists. Each day, the phlebotomists were supplied with a fresh sterile tourniquet, and after use, the tourniquets were swabbed and cultured. The rate of contamination with MRSA was 32 of 131 (25%) tourniquets. An audit of hand hygiene practice was undertaken and revealed that phlebotomists were performing hand decontamination inadequately between patients and wore wristwatches while working. Education comprising standard infection control methods to encourage good practice was given. After this, a polythene strip was used as a barrier by half of the phlebotomists during all venepunctures. Tourniquets were cultured and replaced daily as before. During this stage of the trial, the rates of contamination were 1 of 46 tourniquets (using a polythene strip) and 1 of 42 tourniquets (without using a polythene strip). In conclusion, phlebotomy tourniquets may be potential vectors for transferring bacteria, including MRSA. Contamination rates, and hence potential risk, can be reduced if hand decontamination is performed. This suggests that contamination of tourniquets is via phlebotomists' hands, not directly from patients' skin. Hand hygiene should be regarded as the most important method by which the spread of organisms can be reduced.

Effects of dobutamine in patients with acute myocardial infarction.

This study was designed to evaluate the effects of dobutamine, a new cardioselective beta adrenergic agonist, on cardiac performance and myocardial injury in patients with evolving myocardial infarction. Results in 16 patients given dobutamine (1 to 40 microng/kg per min for 24 hours) were compared with those in two groups of control patients: one of 16 patients matched for predicted infarct size, and the other of 16 patients matched for early ventricular dysrhythmia, analyzed by computer. Infarct size was predicted from plasma creatine kinase (CK) values during the first 7 hours after the initial elevation, before infusion of dobutamine. Overall observed infarct size was estimated from hourly CK values for 48 hours (including those before and after administration of dobutamine). In all patients technetium-99m (stannous) pyrophosphate scans were positive for myocardial infarction. Dobutamine increased cardiac output (assessed by thermodilution) from 4.9 +/- 0.37 (mean +/- standard error) to 6.0 +/- 0.38 liters/min (P less than 0.05) and decreased pulmonary arterial occlusive pressure from 21.5 +/- 2.7 to 16.7 +/- 1.6 mmHg (P less than 0.01) without significantly altering heart rate or systemic arterial blood pressure. The ratio of observed to predicted infarct size, the frequency of independently detected reinfarction or extension of infarction and the frequency of premature ventricular complexes were similar in control and treated patients. Thus administration of dobutamine in doses sufficient to improve ventricular performance after myocardial infarction does not exacerbate myocardial injury or ventricular dysrhythmia.

Differences in the low-energy collision-activated dissociation of carboxylate anions from structurally similar prostaglandins: E2, F 2α, D 2, and DHKF 2α.

Some intriguing discoveries were made concerning the collision-activated dissociation behavior of the derivatized carboxylate anions of PGE2 and PGF2α. The carboxylate anion [MPFB](-) formed from electron-capture negative chemical ionization of the pentafluorobenzyl ester-trimethylsilyl derivative of PGF2α showed little fragmentation under typical collision gas pressures and energies (<2.0 mtorr N2 and <20 eV). In contrast, the daughter spectra of the carboxylate anion of the methoxime-pentafluorobenzyl ester-trimethylsilyl derivative of PGE2 produced many intense fragments under the same conditions.

The effect of specimen processing delay on borate urine preservation.

AIM: To investigate the effect on urine culture results and their clinical interpretation of delaying the processing of urine samples in which boric acid had been used as a preservative. METHODS: 792 mid-stream specimens of urine from patients attending their general practitioner were received in borate containing plastic jars. The specimens were cultured upon receipt, stored at room temperature, and then recultured the following morning. RESULTS: After overnight delayed culture, the results were altered in 16% of samples and the clinical interpretation of these findings differed in 8% of specimens. In 28 samples (3.5%) the bacterium isolated on initial culture was not the same as that obtained by culture after overnight storage. CONCLUSIONS: Boric acid urine preservation used for overnight delayed processing of samples is associated with a significant alteration in culture results and the attendant clinical interpretation of such specimens. Rapid transportation/processing of urine specimens must remain the optimum procedure.

Randomized study of interleukin 2 (IL-2) alone vs IL-2 plus lymphokine-activated killer cells for treatment of melanoma and renal cell cancer.

The purpose of this study was to evaluate the efficacy and safety of a continuous-infusion interleukin 2 (IL-2) regimen for patients with metastatic melanoma and renal cell cancer. To investigate the contribution of adoptively transferred lymphokine-activated killer cells, patients were randomized to receive either IL-2 alone or IL-2 plus lymphokine-activated killer cells. Twenty-three patients with renal cell carcinoma and 20 with melanoma were entered into the protocol. There were no objective responses noted in the 38 assessable patients (20 with renal cell carcinoma, 18 with melanoma). Most patients demonstrated progressive disease following one 31-day cycle of weekly continuous-infusion IL-2. Grade I and II toxic reactions, including fever, rash, anorexia, and weight gain, were common and treated symptomatically. Significant in vivo stimulation of lymphokine-activated killer and natural killer cell activity was noted in most patients. This continuous-infusion IL-2 regimen with or without lymphokine-activated killer cells was ineffective in the treatment of melanoma and renal cell carcinoma.

Peptide fragments derived from the beta-chain of hemoglobin (hemorphins) are centrally active in vivo.

A novel tetrapeptide (hemorphin-4) and pentapeptide (hemorphin-5), derived from the beta-chain of hemoglobin, were synthesized by solid-phase methodology, purified and the amino acid sequences confirmed. The central (ICV) effects of hemorphin-4 and -5 were studied in two models of phasic and tonic nociception, the mouse warm water tail-flick assay and hindpaw formalin assay, respectively. Additionally, two physiological endpoints, central modulation of bladder motility and central effects on intestinal propulsion, were studied in rats and mice, respectively. In the tail-flick assay, both peptides (40-100 nmoles) produced a dose-related naloxone-reversible antinociceptive effect when tested 10 min after peptide administration, with the tetrapeptide being slightly more potent than the pentapeptide. No effect was noted for either peptide using the tonic nociception assay, except at a dose of 150 nmoles for hemorphin-5. Inhibition of gastrointestinal propulsion was also not affected by either peptide. However, both peptides (10-40 nmoles) inhibited micturition contractions in a dose-related and naloxone-reversible fashion, with the tetrapeptide being twice as potent as the pentapeptide. These findings provide evidence that hemorphin-4 and -5 exert naloxone-reversible opioid actions in vivo and, therefore, may be physiologically important blood-borne peptides.

Neurotensin may function as a regulatory peptide in small cell lung cancer.

Neurotensin (NT) has been postulated to act as a modulatory agent in the central nervous system. Besides its presence in mammalian brain, NT is produced by small cell carcinoma of the lung (SCLC) and cell lines derived from these tumors. Receptors have also been characterized in some SCLC cell lines leading to the suggestion that NT could regulate the growth of SCLC in an autocrine fashion similar to bombesin/GRP. Previously, we had reported that a 10 nM dose of NT and NT(8-13), but not NT(1-8), elevated cytosolic Ca2+, indicating that SCLC NT receptors may use Ca2+ as a second messenger. Using intact SCLC cells we report that time-course incubations with NT lead to the formation of the amino-terminal fragment NT(1-8) and small amounts of the C-terminal fragment NT(9-13). These fragments are formed by metalloendopeptidase 3.4.24.15 cleaving enzyme at the Arg8-Arg9 bond of NT. Significant levels of soluble 3.4.24.15 (10-17 nmoles/mg Pr-/min) are present in SCLC cell lines. Using the in vitro clonogenic assay we tested the effect of 0.5, 5.0 and 10.0 nM doses of NT, NT(1-8) and NT(8-13) on SCLC clonal growth. NT and the C-terminal fragment NT(8-13) stimulated colony formation whereas the N-terminal fragment did not. In summary, NT may function as a regulatory peptide in SCLC through the formation of peptide fragments.

Ethanol treatment alters beta-endorphin metabolism by purified synaptosomal plasma membranes.

Ethanol administration has been shown to affect beta-endorphin (beta-E) levels in most brain areas. Chronic ethanol treatment has also lead to changes in the levels of Met- and Leu-enkephalin which may be due to recent finding that enkephalin A activity is significantly altered. To determine if proteolytic enzymes responsible for beta-E metabolism at the pSPM are also altered, we studied the effect of chronic ethanol (7% v/v; 8 days) administration on in vitro central beta-E metabolism in male C57/BL mice. Purified SPM was time-course incubated with beta-E (20 microM) for 30-120 min and subjected to HPLC analyses for determination of beta-endorphin and related fragments. Chronic ethanol significantly reduced the half-life for beta-E at the pSPM (T1/2 = 50/min) versus controls (T1/2 = 100.4 min). Chronic ethanol also caused significant accumulation of the behaviorally active alpha- and gamma-type endorphins formed at the pSPM. These results suggest that chronic ethanol treatment leads to an increase in the activity of peptidases responsible for beta-E metabolism at pSPM leading to an increased formation of both alpha- and gamma-type endorphins which may affect alcohol related behaviors.

Peptide drug modifications to enhance bioavailability and blood-brain barrier permeability.

Peptides have the potential to be potent pharmaceutical agents for the treatment of many central nervous system derived maladies. Unfortunately peptides are generally water-soluble compounds that will not enter the central nervous system, via passive diffusion, due to the existence of the blood-brain barrier. Peptides can also undergo metabolic deactivation by peptidases, thus further reducing their therapeutic benefits. In targeting peptides to the central nervous system consideration must be focused both on increasing bioavailability and enhancing brain uptake. To date multiple strategies have been examined with this focus. However, each strategy comes with its own complications and considerations. In this review we assess the strengths and weaknesses of many of the methods currently being examined to enhance peptide entry into the central nervous system.

In vitro stability of some reduced peptide bond pseudopeptide analogues of dynorphin A.

Eight analogues of DYN A(1-11)-NH2 incorporating the nonhydrolyzable psi [CH2-NH] peptide bond surrogate were tested for their in vitro enzymatic stability in mouse brain homogenates. Results show that the Leu(5)-Arg6 and to a lesser extent the Arg(7)-Ile8 and Ile(8)-Arg9 peptide bonds are the more susceptible to enzymatic cleavage in the native peptide. (Leu5 psi[CH(2)-NH]Arg6)DYN A(1-11)-NH2 exhibits an almost complete resistance to enzymatic cleavage with a half-life greater than 500 min in brain, compared to 42 min for the standard peptide, DYN A(1-11)-NH2.

Neurotensin is metabolized by endogenous proteases in prostate cancer cell lines.

The formation and processing of neurotensin (NT) by three prostate cancer cell lines was investigated. Neurotensin (NT) immunoreactivity was detected in conditioned media and extracts of LNCaP cells. Using HPLC techniques, the immunoreactivity extracted from LNCaP cells coeluted with synthetic NT standard. Metalloendopeptidase 3.4.24.15 activity was detected in PC-3, DU-145 and LNCaP cells, whereas high levels of neutral endopeptidase 3.4.24.1 1 activity was detected only in LNCaP cells. NT was relatively stable when incubated with PC-3 or D-145 cells but was rapidly degraded by LNCaP cells to NT1-11 and NT1-10. Phosphoramidon inhibited the metabolism of NT by LNCaP cells. These data suggest that NT is present in and metabolized by LNCaP cellular enzymes.

The effect of halogenation on blood-brain barrier permeability of a novel peptide drug.

The utility of a drug depends on its ability to reach appropriate receptors at the target tissue and remain metabolically stable to produce the desired effect. To improve central nervous system entry of the opioid analgesic [D-Pen2, L-Pen5, Phe6] Enkephalin (DPLPE-Phe), our research group synthesized analogs that had chloro, bromo, fluoro, and iodo halogens on the para positions of the phenylalanine-4 residue. This study reports on investigation of the effect of halogenation on stability, lipophilicity, and in vitro blood-brain barrier permeability of a novel enkephalin analog DPLPE-Phe. The stability of each halogenated DPLPE-Phe analog as well as the amidated and nonamidated parent peptide was tested in plasma and brain. All peptides tested had a half-time disappearance >300 min except for DPLPE-Phe-NH2, which was found to have a half-life of 30 min in plasma. Octanol/saline distribution studies indicated addition of halogens to DPLPE-Phe-OH significantly increased lipophilicity except for p-[F-Phe4]DPLPE-Phe-OH. p-[Cl-Phe4]DPLPE-Phe-OH exhibited the most pronounced increase in lipophilicity. Para-bromo and para-chloro halogen additions significantly enhanced in vitro blood-brain barrier permeability, providing evidence for improved delivery to the central nervous system.

Characterization of olanzapine (LY170053) in human liver slices by liquid chromatography/tandem mass spectrometry.

Olanzapine metabolism was investigated using incubation of olanzapine with human liver slices. The intent of the investigation was to identify olanzapine metabolites and determine if the human liver slice incubations could potentially produce quantities of the olanzapine glucuronides for future studies. Along with known Phase 1 olanzapine metabolites, N-desmethyl-, 2-hydroxymethyl-, and 4'-N-oxide-, a new hydroxylated species was detected. Detection of Phase 2 metabolites included known N-10-glucuronides, a quaternary glucuronide and a novel glucuronide conjugate. This investigation showed the feasibility of using human liver slices to produce sufficient quantities of olanzapine glucuronides for further studies.

Application of packed capillary liquid chromatography/mass spectrometry with electrospray ionization to the study of the human biotransformation of the anti-emetic drug dolasetron.

Packed capillary liquid chromatography/mass spectrometry (LC/MS) using electrospray ionization (ESI) was used to study the human biotransformation of the anti-emetic drug dolasetron. Urine from subjects given a single 100 mg intravenous dose, containing 14C-labeled dolasetron (50 microCi), was de-salted and concentrated for LC/MS with minimal loss of radioactivity (97% recovery). Aliquots of the de-salted material were injected directly onto a C8 packed capillary column (25 cm x 0.32 mm i.d.) and eluted with an acetonitrile-water gradient, buffered with 1% acetic acid, at a flow rate of 2 microliters min-1. Five metabolites were detected by LC ESI-MS which, yielded molecular mass information but no fragmentation. The identity of each metabolite was confirmed in a subsequent analysis using product ion scans in conjunction with collisionally induced dissociation. Precursor ion scanning was also employed and did not reveal any new biotransformation products. In addition to defining the major routes of biotransformation, the data obtained were compared with a 14C radioprofile prepared in a separate experiment. Qualitative agreement in the two chromatographic profiles enabled the major clusters of radioactivity to be assigned to specific metabolites of dolasetron. An important observation in this comparison was that the signal obtained by ESI did not provide an accurate assessment of the quantity of each metabolite. This was especially true for acidic conjugates (i.e. glucuronides, sulfates), which in the case of dolasetron can exist as zwitterions (no net charge). The results demonstrate the power of packed capillary LC ESI-MS for use in drug biotransformation studies and suggest that caution should be exercised when interpreting relative metabolite abundances from ESI data in the absence of actual reference standards.

Degradation of insulin-like growth factors in small intestine of suckling rats.

Insulin-like growth factors (IFGs), IGF-I and IGF-II, present in mammalian milk, play an important role during gastrointestinal tract development. In this study we identified and localized the activities of the common intestinal proteolytic enzymes and investigated their degradation effect on IGFs. Results indicated that the enzymatic activities of chymotrypsin, trypsin, and elastase progressed from the lowest in the duodenum, to the highest in the midjejunum, and declined in the ileum. Chymotrypsin exhibited the greatest IGFs degradation activities in neonatal intestinal lumen followed by elastase. These data furnish a potential strategic design to supplement IGFs into milk formulas.

Eradication of a resistant Pseudomonas aeruginosa strain after a cluster of infections in a hematology/oncology unit.

OBJECTIVES: This report chronicles an outbreak of a multiply resistant strain of Pseudomonas aeruginosa and the measures required to contain this outbreak. METHODS: Laboratory-based ward-liaison surveillance allowed the detection of a multiply resistant strain of P. aeruginosa infecting patients in our hematology/oncology unit. Sampling of the immediate environment was carried out. Pulsed field gel electrophoresis was used to compare the patients' organisms with those found in the environment. Extensive dismantling of the drainage system, repeated cleaning and disinfection, and a review of the departmental antibiotic policy were some of the infection control measures instigated. RESULTS: During a period of 11 months, three patients in the hematology department and two patients in the oncology department were infected with multiply resistant P. aeruginosa. There were two cases of pneumonia, one of which was fatal, and two cases of neutropenic septicaemia. Pulsed field gel electrophoresis performed on the isolates showed that the isolates from geographically separate areas could be divided into two strains that were closely related but distinct. Two genotypically identical strains were also isolated from the plumbing systems in the areas of each ward where patients had been treated. CONCLUSIONS: The potential for serious nosocomial infections with P. aeruginosa is well recognized. Eradication of the organism from the environment may require the co-ordinated efforts of clinicians, nurses, pharmacy and hospital engineers, working in collaboration with the hospital infection control team. To date, the same strains have not been isolated despite repeated surveillance over the past 18 months and therefore these measures have, in our opinion, successfully removed the potential for nosocomial infection with this resistant organism in our hospital.