Debio 0507 primarily forms diaminocyclohexane-Pt-d(GpG) and -d(ApG) DNA adducts in HCT116 cells.

PURPOSE: To characterize the cellular action mechanism of Debio 0507, we compared the major DNA adducts formed by Debio 0507- and oxaliplatin-treated HCT116 human colon carcinoma cells by a combination of inductively coupled plasma mass spectrometry (ICP-MS) and ultraperformance liquid chromatography mass spectrometry (UPLC-MS/MS). METHODS: HCT116 cells were treated with IC(50) doses of Debio 0507 or oxaliplatin for 3 days. Total cellular Pt-DNA adducts were determined by ICP-MS. The DNA was digested, and the major Pt-DNA adducts formed by both drugs were characterized by UPLC/MS/MS essentially as described previously for cisplatin (Baskerville-Abraham et al. in Chem Res Toxicol 22:905-912, 2009). RESULTS: The Pt level/deoxynucleotide was 7.4/10(4) for DNA from Debio 0507-treated cells and 5.5/10(4) for oxaliplatin-treated cells following a 3-day treatment at the IC(50) for each drug. UPLC-MS/MS in the positive ion mode confirmed the major Pt-DNA adducts formed by both drugs were dach-Pt-d(GpG) (904.2 m/z â†’ 610 m/z and 904.2 m/z â†’ 459 m/z) and dach-Pt-d(ApG) (888.2 m/z â†’ 594 m/z and 888.2 m/z â†’ 459 m/z). CONCLUSIONS: These data show that the major DNA adducts formed by Debio 0507 are the dach-Pt-d(GpG) and dach-Pt-d(ApG) adducts and at equitoxic doses Debio 0507 and oxaliplatin form similar levels of dach-Pt-d(GpG) and dach-Pt-d(ApG) adducts. This suggests that the action mechanisms of Debio 0507 and oxaliplatin are similar at a cellular level.

Aligned silver nanorod arrays as substrates for surface-enhanced infrared absorption spectroscopy.

Preferentially aligned silver nanorod arrays prepared by oblique angle vapor deposition were evaluated as substrates for surface-enhanced infrared absorption (SEIRA) spectroscopy. These nanorod arrays have an irregular surface lattice and are composed of tilted, cylindrically shaped nanorods that have an average length of 868 nm +/- 95 nm and an average diameter of 99 nm +/- 29 nm. The overall enhancement factor for chemisorbed organic films of para-nitrobenzoic acid (PNBA) deposited onto the Ag nanorod arrays analyzed by external reflection SEIRA was calculated to be 31 +/- 9 compared to infrared reflection-absorption spectroscopy (IRRAS) obtained from a 500 nm Ag film substrate. This enhancement is attributed to the unique optical properties of the nanorod arrays as well as the increased surface area provided by the nanorod substrate. SEIRA reflection-absorbance intensity was observed with both p- and s-polarized incident radiation with angles of incidence ranging from 25 degrees to 80 degrees . The largest intensity was achieved with p-polarization and incident angles larger than 75 degrees . Polarization-dependent ultraviolet/visible/near-infrared (UV/Vis/NIR) spectra of the nanorod arrays demonstrate that the red-shifted surface plasmon peaks of the elongated nanorods may be partially responsible for the observed SEIRA response. The SEIRA detection limit for the Ag nanorod arrays was estimated to be 0.08 ng/cm(2). Surface-enhanced Raman scattering (SERS) and SEIRA analysis of chemisorbed PNBA utilizing the same nanorod substrate is demonstrated.

Citrus aurantium and synephrine alkaloids in the treatment of overweight and obesity: an update.

Obesity is a major health problem facing the developed and developing world. Efforts by individuals, health professionals, educators, and policy makers to combat the escalating trend of growing obesity prevalence have been multifaceted and mixed in outcome. Various dietary supplements have been marketed to reduce obesity. These products have been suggested to accomplish this by decreasing energy intake and energy absorption, and/or increasing metabolic rate. Ephedra, one such supplement, was banned from sale in the US market because of concerns about adverse events. Another substance, Citrus aurantium, which contains several compounds including synephrine alkaloids, has been suggested as a safe alternative. This review examines the evidence for safety and efficacy of C. aurantium and synephrine alkaloids as examined in animal studies, clinical weight loss trials, acute physiologic studies and case reports. Although at least three reviews of C. aurantium have been published, our review expands upon these by: (i) distinguishing and evaluating the efficacy of C. aurantium and related compounds; (ii) including results from previously unreviewed research; (iii) incorporating recent case reports that serve to highlight, in an anecdotal way, potential adverse events related to the use of C. aurantium and related compounds; and (iv) offering recommendations to guide the design of future trials to evaluate the safety and efficacy of C. aurantium. While some evidence is promising, we conclude that larger and more rigorous clinical trials are necessary to draw adequate conclusions regarding the safety and efficacy of C. aurantium and synephrine alkaloids for promoting weight loss.

Performance deficits of mGluR8 knockout mice in learning tasks: the effects of null mutation and the background genotype.

mGluR8 is a G-protein coupled metabotropic glutamate receptor expressed in the mammalian brain. Members of the mGluR family have been shown to be modulators of neural plasticity and learning and memory. Here we analyze the consequences of a null mutation at the mGluR8 gene locus generated using homologous recombination in embryonic stem cells by comparing the learning performance of the mutants with that of wild type controls in the Morris water maze (MWM) and the context and cue dependent fear conditioning (CFC). Our results revealed robust performance deficits associated with the genetic background, the ICR outbred strain, in both mGluR8 null mutant and the wild type control mice. Mice of this strain origin suffered from impaired vision as compared to CD1 or C57BL/6 mice, a significant impediment in MWM, a visuo-spatial learning task. The CFC task, being less dependent on visual cues, allowed us to reveal subtle performance deficits in the mGluR8 mutants: novelty induced hyperactivity and temporally delayed and blunted responding to shocks and temporally delayed responding to contextual stimuli were detected. The role of mGluR8 as a presynaptic autoreceptor and its contribution to cognitive processes are hypothesized and the utility of gene targeting as compared to pharmacological methods is discussed.

Outcome of venous stasis ulceration when complicated by arterial occlusive disease.

OBJECTIVE: to report the outcome of patients with venous stasis ulceration (VSU) and severe arterial occlusive disease (AOD). DESIGN: retrospective study. METHODS: using the International Classification of Diseases (ICD-9), codes for VSU and AOD were cross-matched to identify patients from 1989 to 1999 at two tertiary hospitals. Entry into the study required the presence of a VSU and an ipsilateral procedure to improve AOD or major amputation during the same hospitalisation. RESULTS: fourteen patients (15 extremities) with a mean age of 80 years (range: 47-93) were identified as having VSU and AOD. Mean duration of VSU up to the time of revascularisation or amputation was 6.4 years (range: 4 months-21 years). The mean number of VSUs per extremity was 2.1 and mean wound area was 71 cm(2). Mean ankle-brachial index was 0.46 (range: 0.10-0.78). Nine extremities (60%) had a bypass procedure, 3 (20%) had an interventional procedure, 1 (0.6%) had a lumbar sympathectomy, and 2 (13%) had an amputation. Over a mean follow-up of 2.8 years, 3 extremities (23%) healed of which 2 recurred. On last review, 11 patients with 12 afflicted extremities had expired. Nine of the remaining 10 extremities were not healed at the time of death. Eight of nine bypass grafts remained patent in follow-up or at death and subsequent limb salvage was 100%. CONCLUSIONS: combined VSU and AOD represents a rare condition predominantly found in elderly patients with multiple comorbidities. Few patients had complete healing despite an arterial inflow procedure and mortality was high over the short term.

Meal replacements in weight intervention.

OBJECTIVE: To evaluate the effectiveness of meal replacements (MRs) in weight loss interventions in premenopausal women. RESEARCH METHODS AND PROCEDURES: Overweight premenopausal women (n = 113; body mass index: 25 to 35 kg/m(2); 30 to 50 years old) were randomized into three interventions: group A, a dietitian-led intervention; group B, a dietitian-led intervention incorporating MRs; and group C, a clinical office-based intervention incorporating MRs. In year 1, groups A and B attended 26 group sessions, whereas group C received the same educational materials during 26 10-minute office visits with a physician-nurse team. In year 2, participants attended monthly group seminars and drop-in visits with a dietitian. RESULTS: For the 74 subjects completing year 1, weight loss in the office-based group C was as effective as the traditional dietitian-led group A (4.3 +/- 6.5% vs. 4.1 +/- 6.4%), while group B maintained a significantly greater weight loss (9.1 +/- 8.9%; p < 0.02; mean +/- SD). For the 43 subjects completing year 2, group B showed significant differences in the percentage of weight loss (-8.5 +/- 7.0%) compared with group A (-1.5 +/- 5.0%) and group C (-3.0 +/- 7.0%; p < 0.001). DISCUSSION: Study results showed that a traditional weight loss intervention incorporating MRs was effective as a weight loss tool in the medical office practice and in the dietitian-led group setting.

Weight control in the physician's office.

BACKGROUND: Lifestyle changes involving diet, behavior, and physical activity are the cornerstone of successful weight control. Incorporating meal replacements (1-2 per day) into traditional lifestyle interventions may offer an additional strategy for overweight patients in the primary care setting. METHODS: One hundred thirteen overweight premenopausal women (mean +/- SD age, 40.4 +/- 5.5 years; weight, 82 +/- 10 kg; and body mass index, 30 +/- 3 kg/m(2)) participated in a 1-year weight-reduction study consisting of 26 sessions. The women were randomly assigned to 3 different traditional lifestyle-based groups: (1) dietitian-led group intervention (1 hour per session), (2) dietitian-led group intervention incorporating meal replacements (1 hour per session), or (3) primary care office intervention incorporating meal replacements with individual physician and nurse visits (10-15 minutes per visit). RESULTS: For the 74 subjects (65%) completing 1 year, the primary care office intervention using meal replacements was as effective as the traditional dietitian-led group intervention not using meal replacements (mean +/- SD weight loss, 4.3% +/- 6.5% vs 4.1% +/- 6.4%, respectively). Comparison of the dietitian-led groups showed that women using meal replacements maintained a significantly greater weight loss (9.1% +/- 8.9% vs 4.1% +/- 6.4%) (P =.03). Analysis across groups showed that weight loss of 5% to 10% was associated with significant (P =.01) reduction in percentage of body fat, body mass index, waist circumference, resting energy expenditure, insulin level, total cholesterol level, and low-density lipoprotein cholesterol level. Weight loss of 10% or greater was associated with additional significant (P =.05) improvements in blood pressure and triglyceride level. CONCLUSIONS: A traditional lifestyle intervention using meal replacements can be effective for weight control and reduction in risk of chronic disease in the physician's office setting as well as in the dietitian-led group setting.

Injury severity and sensitivity to treatment after controlled cortical impact in rats.

We sought to determine sensitivity of the cortical impact injury model of traumatic brain injury (TBI) to severity of injury and to treatment. We examined the pattern of motor and cognitive deficits and recovery following TBI over a range of injury severities, and examined the efficacy of surface-induced moderate hypothermia at three disparate injury levels. In experiment I, Sprague-Dawley rats were injured at one of eight injury severity levels from 0 mm (sham) to 2.5 mm depth of penetration. On postinjury day 1, balance beam, rotorod performance, and posture reflexes were evaluated. Motor outcome was increasingly impaired with increasing injury levels, with the pattern of deficits showing a step-like function. Cognitive deficits, assessed using water maze on day 7, were more severe for the 2.5-mm group than for the 1.6-mm injury group, while the 1.0-mm group did not differ from the sham controls. In experiments II-IV, hypothermia, 30 degrees C for 3-h duration or normothermia was applied to three injury levels: 1.0 mm, the least cortical deformation; 2.5 mm, the most deformation; and 1.6 mm, representing a level in-between. Neurologic outcome was assessed relative to shams on postinjury days 1, 3, and 5. The 1.0-mm group exhibited small deficits that recovered completely by day 3; the 1.6-mm group recovered to the level of shams by day 5, and the 2.5-mm group did not show significant recovery during the testing period. Hypothermia effectively attenuated behavioral deficits for the 1.6-mm group, but had no effect on the other two groups. These three observations--that increasing injury severity is associated with increasing motor and cognitive deficits, that injury severity is related to recovery time, and that hypothermia treatment is selectively effective--have each been reported in the human TBI population; thus, moderate cortical impact injury in rats may be a model with clinical predictability for evaluating neuroprotective therapies.

The effect of DNA structure on the catalytic efficiency and fidelity of human DNA polymerase beta on templates with platinum-DNA adducts.

DNA adducts formed by platinum-based anticancer drugs interfere with DNA replication. The carrier ligand of the platinum compound is likely to affect the conformation of the Pt-DNA adducts. In addition, the conformation of the adduct can also change upon binding of damaged DNA to the active site of DNA polymerase. From the crystal structures of pol beta ternary complexes it is evident that undamaged gapped and primed single-stranded (non-gapped) DNA templates exist in very different conformations when bound to pol beta. Therefore, one might expect that the constraints imposed on the damaged templates by binding to the polymerase active site should also affect the conformation of the Pt-DNA adducts and their ability to inhibit DNA replication. In support of this hypothesis we have found that the efficiency, carrier ligand specificity, site of discrimination (3'-G versus 5'-G of the Pt-GG adducts), and fidelity of translesion synthesis past Pt-DNA adducts by pol beta are strongly affected by the structure of the DNA template. Previous studies have suggested that the conformation of Pt-DNA adducts may be affected by the sequence context of the adduct. In support of this hypothesis, our data show that sequence context affects the efficiency, fidelity, and pattern of misincorporation by pol beta.

Nutrition and cancer education: ten years of progress.

The Nutrition Education and Research Program at the University of Nevada School of Medicine was awarded two separate NIH/NCI R25 cancer education grants over a ten-year period. With this support, a four-year longitudinal nutrition curriculum was implemented, including the required 20-hour freshman Medical Nutrition Course, junior and senior nutrition electives, and a senior assignment in nutrition and cancer during the rural rotation with faculty preceptors. Funding has also supported nutrition integration into the basic science courses, patient care courses, and specialty clerkships. A unique nutrition fellowship for medical students who specialize in nutrition during their four years of training and graduate with special Qualifications in Nutrition (SQIN) has also been instituted. The curriculum reflects a longitudinal, interdisciplinary, but flexible, integration of nutrition into an already crowded medical school education.

Oxaliplatin-induced damage of cellular DNA.

Damage to cellular DNA is believed to determine the antiproliferative properties of platinum (Pt) drugs. This study characterized DNA damage by oxaliplatin, a diaminocyclohexane Pt drug with clinical antitumor activity. Compared with cisplatin, oxaliplatin formed significantly fewer Pt-DNA adducts (e.g., 0.86+/-0.04 versus 1.36+/- 0.01 adducts/10(6) base pairs/10 microM drug/1 h, respectively, in CEM cells, P<.01). Oxaliplatin was found to induce potentially lethal bifunctional lesions, such as interstrand DNA cross-links (ISC) and DNA-protein cross-links (DPC) in CEM cells. As with total adducts, however, oxaliplatin produced fewer (P<.05) bifunctional lesions than did cisplatin: 0.7+/-0.2 and 1.8+/-0.3 ISC and 0.8+/-0.1 and 1.5+/-0.3 DPC/10(6) base pairs/10 microM drug, respectively, after a 4-h treatment. Extended postincubation (up to 12 h) did not compensate the lower DPC and ISC levels by oxaliplatin. ISC and DPC determinations in isolated CEM nuclei unequivocally verified that oxaliplatin is inherently less able than cisplatin to form these lesions. Reactivation of drug-treated plasmids, observed in four cell lines, suggests that oxaliplatin adducts are repaired with similar kinetics as cisplatin adducts. Oxaliplatin, however, was more efficient than cisplatin per equal number of DNA adducts in inhibiting DNA chain elongation ( approximately 7-fold in CEM cells). Despite lower DNA reactivity, oxaliplatin exhibited similar or greater cytotoxicity in several other human tumor cell lines (50% growth inhibition in CEM cells at 1.1/1.2 microM, respectively). The results demonstrate that oxaliplatin-induced DNA lesions, including ISC and DPC, are likely to contribute to the drug's biological properties. However, oxaliplatin requires fewer DNA lesions than does cisplatin to achieve cell growth inhibition.

The efficiency and fidelity of translesion synthesis past cisplatin and oxaliplatin GpG adducts by human DNA polymerase beta.

DNA polymerase beta (pol beta) is the only mammalian DNA polymerase identified to date that can catalyze extensive bypass of platinum-DNA adducts in vitro. Previous studies suggest that DNA synthesis by pol beta is distributive on primed single-stranded DNA and processive on gapped DNA. The data presented in this paper provide an analysis of translesion synthesis past cisplatin- and oxaliplatin-DNA adducts by pol beta functioning in both distributive and processive modes using primer extension and steady-state kinetic experiments. Translesion synthesis past Pt-DNA adducts was greater with gapped DNA templates than with single-stranded DNA templates. In the processive mode pol beta did not discriminate between cisplatin and oxaliplatin adducts, while in the distributive mode it displayed about 2-fold increased ability for translesion synthesis past oxaliplatin compared with cisplatin adducts. The differentiation between cisplatin and oxaliplatin adducts resulted from a K(m)-mediated increase in the efficiency of dCTP incorporation across from the 3'-G of oxaliplatin-GG adducts. Rates of misincorporation across platinated guanines determined by the steady-state kinetic assay were higher in reactions with primed single-stranded templates than with gapped DNA and a slight increase in the misincorporation of dTTP across from the 3'-G was found for oxaliplatin compared with cisplatin adducts.

Efficient translesion replication past oxaliplatin and cisplatin GpG adducts by human DNA polymerase eta.

Platinum anticancer agents form bulky DNA adducts which are thought to exert their cytotoxic effect by blocking DNA replication. Translesion synthesis, one of the pathways of postreplication repair, is thought to account for some resistance to DNA damage and much of the mutagenicity of bulky DNA adducts in dividing cells. Oxaliplatin has been shown to be effective in cisplatin-resistant cell lines and less mutagenic than cisplatin in the Ames assay. We have shown that the eukaryotic DNA polymerases yeast pol zeta, human pol beta, and human pol gamma bypass oxaliplatin-GG adducts more efficiently than cisplatin-GG adducts. Human pol eta, a product of the XPV gene, has been shown to catalyze efficient translesion synthesis past cis, syn-cyclobutane pyrimidine dimers. In the present study we compared translesion synthesis past different Pt-GG adducts by human pol eta. Our data show that, similar to other eukaryotic DNA polymerases, pol eta bypasses oxaliplatin-GG adducts more efficiently than cisplatin-GG adducts. However, pol eta-catalyzed translesion replication past Pt-DNA adducts was more efficient and less accurate than that seen for previously tested polymerases. We show that the efficiency and fidelity of translesion replication past Pt-DNA adducts appear to be determined by both the structure of the adduct and the DNA polymerase active site.

Ectopic G-protein expression in dopamine and serotonin neurons blocks cocaine sensitization in Drosophila melanogaster.

Sensitization to repeated doses of psychostimulants is thought to be an important component underlying the addictive process in humans [1] [2] [3] [4]. In all vertebrate animal models, including humans [5], and even in fruit flies, sensitization is observed after repeated exposure to volatilized crack cocaine [6]. In vertebrates, sensitization is thought to be initiated by processes occurring in brain regions that contain dopamine cell bodies [2] [7]. Here, we show that modulated cell signaling in the Drosophila dopamine and serotonin neurons plays an essential role in cocaine sensitization. Targeted expression of either a stimulatory (Galpha(s)) or inhibitory (Galpha(i)) Galpha subunit, or tetanus toxin light chain (TNT) in dopamine and serotonin neurons of living flies blocked behavioral sensitization to repeated cocaine exposures. These flies showed alterations in their initial cocaine responsiveness that correlated with compensatory adaptations of postsynaptic receptor sensitivity. Finally, repeated drug stimulation of a nerve cord preparation that is postsynaptic to the brain amine cells failed to induce sensitization, further showing the importance of presynaptic modulation in sensitization.

Child abuse: clinical findings and management.

PURPOSE: To provide an overview of the role of the nurse practitioner (NP) in identifying, reporting, and managing child abuse in primary care. DATA SOURCES: Selected research, national guidelines, and the author's experience. CONCLUSIONS: Child abuse is a complex phenomenon characterized by maladaptive behaviors between children and their parents. IMPLICATIONS FOR PRACTICE: The role of the NP includes identification of families at risk, recognition of clinical findings of abuse, diagnosis of abuse, education for families identified at risk, and management of children diagnosed with abuse. The ultimate goal is the safe return of the child to a loving family.

Critical pathways: a means of managing the operating room.

Critical pathways help institutions in efficient and appropriate resource use to increase the quality of health care and minimize health care costs. However, many opportunities for pathway development and implementation are unexplored. This article delineates the development process for critical pathways and discusses the outcomes realized from use of the total joint pathway at the Medical College of Georgia, Augusta, GA.

Efficient nucleotide excision repair of cisplatin, oxaliplatin, and Bis-aceto-ammine-dichloro-cyclohexylamine-platinum(IV) (JM216) platinum intrastrand DNA diadducts.

Tumors exhibit a spectrum of cellular responses to chemotherapy ranging from extreme sensitivity to resistance, either intrinsic or acquired. These variable responses are both patient and tumor specific. For platinum DNA-damaging agents, drug resistance depends on the carrier ligand of the platinum complex and is due to a combination of mechanisms including DNA repair. Nucleotide excision repair is the only known mechanism by which bulky adducts, including those generated by platinum chemotherapeutic agents, are removed from DNA in human cells. In this report, we show that the types of DNA lesions generated by three platinum drugs, cisplatin, oxaliplatin, and (Bis-aceto-ammine-dichloro-cyclohexylamine-platinum(IV) (JM216), are repaired in vitro with similar kinetics by the mammalian nucleotide excision repair pathway.

Effect of DNA polymerases and high mobility group protein 1 on the carrier ligand specificity for translesion synthesis past platinum-DNA adducts.

Translesion synthesis past Pt-DNA adducts can affect both the cytotoxicity and mutagenicity of the platinum adducts. We have shown previously that the extent of replicative bypass in vivo is influenced by the carrier ligand of platinum adducts. The specificity of replicative bypass may be determined by the DNA polymerase complexes that catalyze translesion synthesis past Pt-DNA adducts and/or by DNA damage-recognition proteins that bind to the Pt-DNA adducts and block translesion replication. In the present study, primer extension on DNA templates containing site-specifically placed cisplatin, oxaliplatin, JM216, or chlorodiethylenetriamine-Pt adducts revealed that the eukaryotic DNA polymerases beta, zeta, gamma, and human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) had a similar specificity for translesion synthesis past Pt-DNA adducts (dien >> oxaliplatin >/= cisplatin > JM216). Primer extension assays performed in the presence of high mobility group protein 1 (HMG1), which is known to recognize cisplatin-damaged DNA, revealed that inhibition of translesion synthesis by HMG1 also depended on the carrier ligand of the Pt-DNA adduct (cisplatin > oxaliplatin = JM216 >> dien). These data were consistent with the results of gel-shift experiments showing similar differences in the affinity of HMG1 for DNA modified with the different platinum adducts. Our studies show that both DNA polymerases and damage-recognition proteins can impart specificity to replicative bypass of Pt-DNA adducts. This information may serve as a model for further studies of translesion synthesis.

Requirement of circadian genes for cocaine sensitization in Drosophila.

The circadian clock consists of a feedback loop in which clock genes are rhythmically expressed, giving rise to cycling levels of RNA and proteins. Four of the five circadian genes identified to date influence responsiveness to freebase cocaine in the fruit fly, Drosophila melanogaster. Sensitization to repeated cocaine exposures, a phenomenon also seen in humans and animal models and associated with enhanced drug craving, is eliminated in flies mutant for period, clock, cycle, and doubletime, but not in flies lacking the gene timeless. Flies that do not sensitize owing to lack of these genes do not show the induction of tyrosine decarboxylase normally seen after cocaine exposure. These findings indicate unexpected roles for these genes in regulating cocaine sensitization and indicate that they function as regulators of tyrosine decarboxylase.