Epilepsy, one of the most common neurologic disorders, is a major public health issue. Despite more than 20 approved antiepileptic drugs (AEDs), about 30% of patients are refractory to treatment. An important characteristic of pharmacoresistant epilepsy is that most patients with refractory epilepsy are resistant to several, if not all, AEDs, even though these drugs act by different mechanisms. This argues against epilepsy-induced alterations in specific drug targets as a major cause of pharmacoresistant epilepsy, but rather points to nonspecific and possibly adaptive mechanisms, such as decreased drug uptake into the brain by intrinsic or acquired over-expression of multidrug transporters in the blood-brain barrier (BBB). There is accumulating evidence demonstrating that multidrug transporters such as P-glycoprotein (PGP) and members of the multidrug resistance-associated protein (MRP) family are over-expressed in capillary endothelial cells and astrocytes in epileptogenic brain tissue surgically resected from patients with medically intractable epilepsy. PGP and MRPs in the BBB are thought to act as an active defense mechanism, restricting the penetration of lipophilic substances into the brain. A large variety of compounds, including many lipophilic drugs, are substrates for either PGP or MRPs or both. It is thus not astonishing that several AEDs, which have been made lipophilic to penetrate into the brain, seem to be substrates for multidrug transporters in the BBB. Over-expression of such transporters in epileptogenic tissue is thus likely to reduce the amount of drug that reaches the epileptic neurons, which would be a likely explanation for pharmacoresistance. PGP and MRPs can be blocked by specific inhibitors, which raises the option to use such inhibitors as adjunctive treatment for medically refractory epilepsy. However, although over-expression of multidrug transporters is a novel and reasonable hypothesis to explain multidrug resistance in epilepsy, further studies are needed to establish this concept. Furthermore, there are certainly other mechanisms of pharmacoresistance that need to be identified.

The central nucleus of the amygdala, bed nucleus of the stria terminalis, and central gray are important components of the neural circuitry responsible for autonomic and behavioral responses to threatening or stressful stimuli. Neurons of the amygdala and bed nucleus of the stria terminalis that project to the midbrain central gray were tested for the presence of peptide immunoreactivity. To accomplish this aim, a combined immunohistochemical and retrograde tracing technique was used. Maximal retrograde labeling was observed in the amygdala and bed nucleus of the stria terminalis after injections of retrograde tracer into the caudal ventrolateral midbrain central gray. The majority of the retrogradely labeled neurons in the amygdala were located in the medial central nucleus, although many neurons were also observed in the lateral subdivision of the central nucleus. Most of the retrogradely labeled neurons in the BST were located in the ventral and posterior lateral subdivisions, although cells were also observed in most other subdivisions. Retrogradely labeled neurotensin, corticotropin releasing factor (CRF), and somatostatin neurons were mainly observed in the lateral central nucleus and the dorsal lateral BST. Retrogradely labeled substance P-immunoreactive cells were found in the medial central nucleus and the posterior and ventral lateral BST. Enkephalin-immunoreactive retrogradely labeled cells were not observed in the amygdala or bed nucleus of the stria terminalis. A few cells in the hypothalamus (paraventricular and lateral hypothalamic nuclei) that project to the central gray also contained CRF and neurotensin immunoreactivity. The results suggest the amygdala and the bed nucleus of the stria terminalis are a major forebrain source of CRF, neurotensin, somatostatin, and substance P terminals in the midbrain central gray.

Multidrug resistance (MDR) is the protection of a tumor cell population against numerous drugs differing in chemical structure and mechanisms of influence on the cells. MDR is one of the major causes of failures of chemotherapy of human malignancies. Recent studies show that the molecular mechanisms of MDR are numerous. Cellular drug resistance is mediated by different mechanisms operating at different steps of the cytotoxic action of the drug from a decrease of drug accumulation in the cell to the abrogation of apoptosis induced by the chemical substance. Often several different mechanisms are switched on in the cells, but usually one major mechanism is operating. The most investigated mechanisms with known clinical significance are: a) activation of transmembrane proteins effluxing different chemical substances from the cells (P-glycoprotein is the most known efflux pump); b) activation of the enzymes of the glutathione detoxification system; c) alterations of the genes and the proteins involved into the control of apoptosis (especially p53 and Bcl-2).

BACKGROUND

Dietary carbohydrates have been associated with dyslipidemia, a lipid profile known to increase cardiovascular disease risk. Added sugars (caloric sweeteners used as ingredients in processed or prepared foods) are an increasing and potentially modifiable component in the US diet. No known studies have examined the association between the consumption of added sugars and lipid measures.

OBJECTIVE

To assess the association between consumption of added sugars and blood lipid levels in US adults.

METHODS

Cross-sectional study among US adults (n = 6113) from the National Health and Nutrition Examination Survey (NHANES) 1999-2006. Respondents were grouped by intake of added sugars using limits specified in dietary recommendations (< 5% [reference group], 5%-<10%, 10%-<17.5%, 17.5%-<25%, and>> or = 25% of total calories). Linear regression was used to estimate adjusted mean lipid levels. Logistic regression was used to determine adjusted odds ratios of dyslipidemia. Interactions between added sugars and sex were evaluated.

METHODS

Adjusted mean high-density lipoprotein cholesterol (HDL-C), geometric mean triglycerides, and mean low-density lipoprotein cholesterol (LDL-C) levels and adjusted odds ratios of dyslipidemia, including low HDL-C levels (< 40 mg/dL for men; < 50 mg/dL for women), high triglyceride levels >> or = 150 mg/dL), high LDL-C levels >> or = 130 mg/dL), or high ratio of triglycerides to HDL-C >> 3.8). Results were weighted to be representative of the US population.

RESULTS

A mean of 15.8% of consumed calories was from added sugars. Among participants consuming less than 5%, 5% to less than 17.5%, 17.5% to less than 25%, and 25% or greater of total energy as added sugars, adjusted mean HDL-C levels were, respectively, 58.7, 57.5, 53.7, 51.0, and 47.7 mg/dL (P < .001 for linear trend), geometric mean triglyceride levels were 105, 102, 111, 113, and 114 mg/dL (P < .001 for linear trend), and LDL-C levels modified by sex were 116, 115, 118, 121, and 123 mg/dL among women (P = .047 for linear trend). There were no significant trends in LDL-C levels among men. Among higher consumers >> or = 10% added sugars) the odds of low HDL-C levels were 50% to more than 300% greater compared with the reference group (< 5% added sugars).

CONCLUSIONS

In this study, there was a statistically significant correlation between dietary added sugars and blood lipid levels among US adults.

BACKGROUND

Posttraumatic stress disorder (PTSD) is associated with medical and psychological morbidity. The prevalence of PTSD in urban primary care has not been well described.

OBJECTIVE

To measure the prevalence of PTSD in primary care patients overall and among those with selected conditions (chronic pain, depression, anxiety, heavy drinking, substance dependence (SD), irritable bowel syndrome (IBS), and immigrant status).

METHODS

Cross-sectional study.

METHODS

English-speaking patients aged 18-65 years old, awaiting primary care appointments in an urban academic medical center, were eligible for enrollment to determine PTSD prevalence (N = 509). Additional eligible participants (n = 98) with IBS or SD were subsequently enrolled.

METHODS

PTSD (past year) and trauma exposure were measured with Composite International Diagnostic Interview. We calculated the prevalence of PTSD associated with depression, anxiety, heavy drinking, SD, IBS, and chronic pain. Only the analyses on heavy drinking, SD, and IBS used all 607 participants.

RESULTS

Among the 509 adults in primary care, 23% (95% CI, 19-26%) had PTSD, of whom 11% had it noted in the medical record. The prevalence of PTSD, adjusted for age, gender, race, and marital and socioeconomic statuses, was higher in participants with, compared to those without, the following conditions: chronic pain (23 vs 12%, p = .003), major depression (35 vs 11%, p < .0001), anxiety disorders (42 vs 14%, p < .0001), and IBS (34 vs 18%, p = .01) and lower in immigrants (13 vs 21%, p = .05).

CONCLUSIONS

The prevalence of PTSD in the urban primary care setting, and particularly among certain high-risk conditions, compels a critical examination of optimal approaches for screening, intervention, and referral to PTSD treatment.

OBJECTIVE

Tourette syndrome (TS) is characterized by high rates of psychiatric comorbidity; however, few studies have fully characterized these comorbidities. Furthermore, most studies have included relatively few participants (<200), and none has examined the ages of highest risk for each TS-associated comorbidity or their etiologic relationship to TS.

OBJECTIVE

To characterize the lifetime prevalence, clinical associations, ages of highest risk, and etiology of psychiatric comorbidity among individuals with TS.

METHODS

Cross-sectional structured diagnostic interviews conducted between April 1, 1992, and December 31, 2008, of participants with TS (n = 1374) and TS-unaffected family members (n = 1142).

METHODS

Lifetime prevalence of comorbid DSM-IV-TR disorders, their heritabilities, ages of maximal risk, and associations with symptom severity, age at onset, and parental psychiatric history.

RESULTS

The lifetime prevalence of any psychiatric comorbidity among individuals with TS was 85.7%; 57.7% of the population had 2 or more psychiatric disorders. The mean (SD) number of lifetime comorbid diagnoses was 2.1 (1.6); the mean number was 0.9 (1.3) when obsessive-compulsive disorder (OCD) and attention-deficit/hyperactivity disorder (ADHD) were excluded, and 72.1% of the individuals met the criteria for OCD or ADHD. Other disorders, including mood, anxiety, and disruptive behavior, each occurred in approximately 30% of the participants. The age of greatest risk for the onset of most comorbid psychiatric disorders was between 4 and 10 years, with the exception of eating and substance use disorders, which began in adolescence (interquartile range, 15-19 years for both). Tourette syndrome was associated with increased risk of anxiety (odds ratio [OR], 1.4; 95% CI, 1.0-1.9; P = .04) and decreased risk of substance use disorders (OR, 0.6; 95% CI, 0.3-0.9; P = .02) independent from comorbid OCD and ADHD; however, high rates of mood disorders among participants with TS (29.8%) may be accounted for by comorbid OCD (OR, 3.7; 95% CI, 2.9-4.8; P < .001). Parental history of ADHD was associated with a higher burden of non-OCD, non-ADHD comorbid psychiatric disorders (OR, 1.86; 95% CI, 1.32-2.61; P < .001). Genetic correlations between TS and mood (RhoG, 0.47), anxiety (RhoG, 0.35), and disruptive behavior disorders (RhoG, 0.48), may be accounted for by ADHD and, for mood disorders, by OCD.

CONCLUSIONS

This study is, to our knowledge, the most comprehensive of its kind. It confirms the belief that psychiatric comorbidities are common among individuals with TS, demonstrates that most comorbidities begin early in life, and indicates that certain comorbidities may be mediated by the presence of comorbid OCD or ADHD. In addition, genetic analyses suggest that some comorbidities may be more biologically related to OCD and/or ADHD rather than to TS.

Opiates are the primary treatment for pain management in cancer patients reporting moderate to severe pain, and are being increasingly used for non-cancer chronic pain. However, prolonged administration of opiates is associated with significant problems including the development of antinociceptive tolerance, wherein higher doses of the drug are required over time to elicit the same amount of analgesia. High doses of opiates result in serious side effects such as constipation, nausea, vomiting, dizziness, somnolence, and impairment of mental alertness. In addition, sustained exposure to morphine has been shown to result in paradoxical pain in regions unaffected by the initial pain complaint, and which may also result in dose escalation, i.e. 'analgesic tolerance'. A concept that has been gaining considerable experimental validation is that prolonged use of opioids elicits paradoxical, abnormal pain. This enhanced pain state requires additional opioids to maintain a constant level of antinociception, and consequently may be interpreted as antinociceptive tolerance. Many substances have been shown to block or reverse antinociceptive tolerance. A non-inclusive list of examples of substances reported to block or reverse opioid antinociceptive tolerance include: substance P receptor (NK-1) antagonists, calcitonin gene-related peptide (CGRP) receptor antagonists, nitric oxide (NO) synthase inhibitors, calcium channel blockers, cyclooxygenase (COX) inhibitors, protein kinase C inhibitors, competitive and non-competitive antagonists of the NMDA (N-methyl-D-aspartate) receptor, AMPA (alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid) antagonists, anti-dynorphin antiserum, and cholecystokinin (CCK) receptor antagonists. Without exception, these substances are also antagonists of pain-enhancing agents. Prolonged opiate administration indeed induces upregulation of substance P (SP) and calcitonin gene-related peptide (CGRP) within sensory fibers in vivo, and this is accompanied by an enhanced release of excitatory neurotransmitters and neuropeptides from primary afferent fibers upon stimulation. The enhanced evoked release of neuropeptides is correlated with the onset of abnormal pain states and opioid antinociceptive tolerance. Importantly, the descending pain modulatory pathway from the brainstem rostral ventromedial medulla (RVM) via the dorsolateral funiculus (DLF) is critical for maintaining the changes observed in the spinal cord, abnormal pain states and antinociceptive tolerance, because animals with lesion of the DLF did not show enhanced evoked neuropeptide release, or develop abnormal pain or antinociceptive tolerance upon sustained exposure to opiates. Microinjection of either lidocaine or a CCK antagonist into the RVM blocked both thermal and touch hypersensitivity as well as antinociceptive tolerance. Thus, prolonged opioid exposure enhances a descending pain facilitatory pathway from the RVM that is mediated at least in part by CCK activity and is essential for the maintenance of antinociceptive tolerance.

BACKGROUND

Hepatitis C virus (HCV) infection is both prevalent and undertreated.

OBJECTIVE

To identify barriers to HCV treatment in primary care practice.

METHODS

Cross-sectional study.

METHODS

A cohort of 208 HCV-infected patients under the care of a primary care physician (PCP) between December 2001 and April 2004 at a single academically affiliated community health center.

METHODS

Data were collected from the electronic medical record (EMR), the hospital clinical data repository, and interviews with PCPs.

RESULTS

Our cohort consisted of 208 viremic patients with HCV infection. The mean age was 47.6 (+/-9.7) years, 56% were male, and 79% were white. Fifty-seven patients (27.4% of the cohort) had undergone HCV treatment. Independent predictors of not being treated included: unmarried status (adjusted odds ratio [aOR] for treatment 0.36, P=.02), female gender (aOR 0.31, P=.01), current alcohol abuse (aOR 0.08, P=.0008), and a higher ratio of no-shows to total visits (aOR 0.005 per change of 1.0 in the ratio of no-shows to total visits, P=.002). The major PCP-identified reasons not to treat included: substance abuse (22.5%), patient preference (16%), psychiatric comorbidity (15%), and a delay in specialist input (12%). For 13% of the untreated patients, no reason was identified.

CONCLUSIONS

HCV treatment was infrequent in our cohort of outpatients. Barriers to treatment included patient factors (patient preference, alcohol use, missed appointments), provider factors (reluctance to treat past substance abusers), and system factors (referral-associated delays). Multimodal interventions may be required to increase HCV treatment rates.

A highly specific two-site immunoradiometric assay for insulin was used to measure the plasma insulin response to 75 g glucose administered orally to 49 patients with non-insulin-dependent diabetes (NIDDM). The plasma insulin concentration 30 min after glucose ingestion was lower in the diabetic patients than in matched controls for both non-obese (11-83 pmol/l vs 136-297 pmol/l, p less than 0.01) and obese subjects (23-119 pmol/l vs 137-378 pmol/l, p less than 0.01). By means of another two-site immunoradiometric assay, the basal intact proinsulin level was found to be higher in the NIDDM patients than in the controls for both non-obese (7.1 [SEM 1.2] pmol/l vs 2.4 [0.4] pmol/l, p less than 0.01) and obese subjects (14.4 [2.2] pmol/l vs 5.9 [1.9] pmol/l, p less than 0.01). The basal level of 32-33 split proinsulin was also raised in NIDDM. Previous failure to show clear separation between normal and NIDDM insulin responses was probably due to the high concentrations of proinsulin-like molecules in the plasma of NIDDM patients. These substances cross-react as insulin in most, if not all, insulin radioimmunoassays but have very little biological insulin-like activity. It is therefore now possible and necessary to designate most NIDDM patients as insulin deficient.

1. We describe the actions of GR43175, a 5-hydroxytryptamine1 (5-HT1)-like receptor agonist, on neurogenically-mediated plasma protein extravasation within an important pain-sensitive intracranial tissue, the dura mater. 2. GR43175 markedly attenuated extravasation of 125I-albumin from blood vessels within ipsilateral dura mater when administered to rats (100 micrograms kg-1) fifteen minutes before unilateral electrical trigeminal stimulation (1.2 mA, 5 Hz, 5 ms, 5 min); the ratio (stimulated/unstimulated sides) decreased from 1.81 to 1.23, P less than 0.005). 3. GR43175 (100 micrograms kg-1, i.v., rats; 30 micrograms kg-1, guinea-pigs) decreased the leakage of radiolabelled albumin from 163% to 119% (P less than 0.005, guinea-pig) or from 174 to 118% (P less than 0.05, rat) above vehicle-treated controls when injected ten minutes before systemic capsaicin treatment (0.5 or 1 mumol kg-1, i.v.). 4. GR43175 (30-300 micrograms kg-1) did not block plasma protein extravasation within extracranial tissues of rats and guinea-pigs innervated by the trigeminal nerve (conjunctiva, eyelid and lip). 5. The protein leakage which followed the i.v. administration of 5-HT (1 mumol kg-1) or neuropeptides which mediate neurogenic plasma extravasation, substance P (0.3 nmol kg-1 or 1 nmol kg-1) and neurokinin A (1 nmol kg-1), was not blocked by GR43175 (100, 300 micrograms kg-1) despite the presence of leakage in amounts equivalent to that following neurogenic stimulation. 6. GR43175 (100 micrograms kg-1) decreased bradykinin (10 mumol kg-1)-induced extravasation from 142 to 115% above vehicle-treated animals (P less than 0.05). 7. These results demonstrate an important action of GR43175 on neurogenic mechanisms in dural blood vessels. Since the ergot alkaloids possess a similar profile of drug activity, it is suggested that drugs useful in the treatment of acute vascular headaches may share a similar mechanism of action.

1. Substance P (SP) induces histamine release from isolated rat peritoneal mast cells at concentrations of 0.1-10 muM.2. Inhibitors of glycolysis and oxidative phosphorylation prevent the release of histamine induced by SP.3. Cells heated to 47 degrees C for 20 min release histamine when treated with an agent causing cell lysis but fail to release in response to SP.4. SP does not release histamine by interacting with cell-bound IgE.5. Histamine release by SP is rapid, with more than 90% of the response occurring within 1 min of the addition of the peptide to mast cells at 37 degrees C.6. Substance P, unlike antigen-antibody or compound 48/80, does not show enhanced release of histamine when calcium (0.1-1 mM) is present in the extracellular medium but calcium increases the response to SP when the ion is added after the peptide. Extracellular calcium (0.1-1 mM), magnesium (1-10 mM) and cobalt (0.01-0.1 mM) all inhibit SP-induced histamine release when added before the peptide. Pre-treatment of the cells with EDTA (10 mM) and washing in calcium-free medium inhibits the histamine release induced by SP.7. Histamine release induced by SP was optimum at an extracellular pH of 7.2.8. A number of peptides structurally related to SP were examined for histamine-releasing activity. At the concentrations tested, the N-terminal dipeptides Lys-Pro and Arg-Pro, tuftsin, physalaemin, eledoisin, SP(3-11), SP(4-11) and [p-Glu(6), p-amino Phe(7)]-SP(6-11) were all found to be inactive. The relative activities of the other peptides were: [Formula: see text]9. Rat basophilic leukaemia cells (RBL-2H3) fail to respond to SP at concentrations which activate rat mast cells. Release of 5-hydroxytryptamine by immunological activation of RBL cells is not changed by the presence of SP.10. The mechanism of action of SP on mast cells and the nature of the SP receptor on mast cells is discussed in relation to SP receptors in other cell types.

Several lines of evidence indicate that tachykinin neuropeptides [substance P (SP), substance K (SK), and neuromedin K (NK)] play a role in regulating the inflammatory and immune responses. To test this hypothesis in a human inflammatory disease, quantitative receptor autoradiography was used to examine possible abnormalities in tachykinin binding sites in surgical specimens from patients with inflammatory bowel disease. Surgical specimens of colon were obtained from patients with ulcerative colitis (n = 4) and Crohn disease (n = 4). Normal tissue was obtained from uninvolved areas of extensive resections for carcinoma (n = 6). In all cases, specimens were obtained less than 5 min after removal to minimize influences associated with degradation artifacts and were processed for quantitative receptor autoradiography by using 125I-labeled Bolton-Hunter conjugates of NK, SK, and SP. In the normal colon a low concentration of SP receptor binding sites is expressed by submucosal arterioles and venules and a moderate concentration is expressed by the external circular muscle, whereas SK receptor binding sites are expressed in low concentrations by the external circular and longitudinal muscle. In contrast, specific NK binding sites were not observed in any area of the human colon. In colon tissue obtained from ulcerative colitis and Crohn disease patients, however, very high concentrations of SP receptor binding sites are expressed by arterioles and venules located in the submucosa, muscularis mucosa, external circular muscle, external longitudinal muscle, and serosa. In addition, very high concentrations of SP receptor binding sites are expressed within the germinal center of lymph nodules, whereas the concentrations of SP and SK binding sites expressed by the external muscle layers are not altered significantly. These results demonstrate that receptor binding sites for SP, but not SK or NK, are ectopically expressed in high concentrations (1000-2000 times normal) by cells involved in mediating inflammatory and immune responses. These data suggest that SP may be involved in the pathophysiology of inflammatory bowel disease and might provide some insight into the interaction between the nervous system and the regulation of inflammation and the immune response in human inflammatory disease.

Using postmortem immunohistochemical analysis, we have identified degeneration of several different neuronal cell groups in the brainstem of patients dying with idiopathic Parkinson's disease. We report the first chemically identified loss of presumed serotonin neurons in the median raphe nucleus of the pons and of substance P-containing preganglionic neurons in the dorsal motor vagal nucleus. This evidence is concordant with other evidence that the primary neuropathological process is not confined either to a single pathway or to neurons containing a particular transmitter. Rather it appears that Parkinson's disease affects several classes of neurons in localized areas of the brainstem.

OBJECTIVE

To determine antioxidant vitamin concentrations, lipid peroxidation, and an index of nitric oxide production in patients in the intensive care unit (ICU) with septic shock and relate the findings to the presence of secondary organ failure.

METHODS

A prospective, observational study.

METHODS

A nine-bed ICU in a University teaching hospital.

METHODS

Sixteen consecutive patients with septic shock, defined as: a) clinical evidence of acute infection; b) hypo- or hyperthermia (< 35.6 degrees C or>> 38.3 degrees C); c) tachypnea >> 20 breaths/min or being mechanically ventilated); d) tachycardia >> 90 beats/min); e) shock (systolic pressure < 90 mm Hg) or receiving inotropes. Fourteen patients also had secondary organ dysfunction.

METHODS

None.

RESULTS

Antioxidant vitamin concentrations were significantly lower in the patients than the reference range obtained from a comparable group of healthy controls. The mean plasma retinol (vitamin A) concentration was 26.5 +/- 19.3 micrograms/dL compared with 73.5 +/- 18.3 micrograms/dL in healthy subjects (p < .01). Additionally, 13 (81%) patients had retinol values below the lower limit of our reference range (< 37.0 micrograms/dL). Tocopherol (vitamin E) plasma concentrations were below the reference range in all patients (< 9.0 mg/L), with a mean value of 3.6 +/- 2.0 mg/L compared with 11.5 +/- 1.3 mg/L in healthy subjects (p < .001). Plasma beta carotene and lycopene concentrations were undetectable (< 15 micrograms/L) in eight (50%) patients, and below our reference range (< 101 micrograms/L and < 154 micrograms/L, respectively) in the remaining patients. In the five patients with three or more dysfunctional secondary organs, plasma thiobarbituric acid-reactive substances were significantly increased (p < .05), suggesting increased lipid peroxidation. Concentrations of thiobarbituric acid-reactive substances correlated negatively with both plasma retinol and plasma tocopherol (r2 = .42, p < .01 and r2 = .48, p < .005, respectively). In the five patients from whom we were able to collect urine, nitrite excretion was increased approximately 400-fold (p < .001).

CONCLUSIONS

These data indicate decreased antioxidant status in the face of enhanced free radical activity, and suggest potential therapeutic strategies involving antioxidant repletion.

OBJECTIVE

The methionine choline-deficient (MCD) diet leads to steatohepatitis in rodents. The aim of the present study was to investigate species, strain and sex differences in this nutritional model of non-alcoholic steatohepatitis (NASH).

METHODS

Male and female Wistar, Long-Evans and Sprague-Dawley rats, and C57/BL6 mice (n = 6 per group) were fed a MCD diet for 4 weeks. Control groups received an identical diet supplemented with choline bitartrate (0.2% w/w) and methionine (0.3% w/w). Liver pathology (steatosis and inflammation) and ultrastructure, liver lipid profile (total lipids, triglycerides, lipid peroxidation products), liver : body mass ratios and serum alanine aminotransferase (ALT) levels were compared between these groups.

RESULTS

The MCD diet-fed male rats developed greater steatosis (P < 0.001), had higher liver lipid content (P < 0.05) and had higher serum ALT levels (P < 0.005) than did female rats. Wistar rats (both sexes) had higher liver lipid levels (P < 0.05), serum ALT levels (P < 0.05), and liver mass : body mass ratios (P < 0.025) than did Long-Evans and Sprague-Dawley rats. In female groups, Wistar rats showed greater fatty change than did the other two strains (P < 0.05). All rats fed the MCD diet developed hepatic steatosis, but necrosis and inflammation were minor features and fibrosis was absent. Compared with Wistar rats, male C57/BL6 mice showed a marked increase in inflammatory foci (P < 0.001), end products of lipid peroxidation (free thiobarbituric acid reactive substances) (P < 0.005), and mitochondrial injury, while showing less steatosis (P < 0.005), lower hepatic triglyceride levels, (P < 0.005) and lower early lipid peroxidation products (conjugated dienes and lipid hydroperoxides; P < 0.005 and P < 0.01, respectively).

CONCLUSIONS

The Wistar strain and the male sex are associated with the greatest degree of steatosis in rats subjected to the MCD diet. Of the groups studied, male C57/BL6 mice develop the most inflammation and necrosis, lipid peroxidation, and ultrastructural injury, and best approximate the histological features of NASH.

BACKGROUND

Contingency management (CM) and significant other involvement (SO) were evaluated as strategies to enhance treatment retention, medication compliance, and outcome for naltrexone treatment of opioid dependence.

METHODS

One hundred twenty-seven recently detoxified opioid-dependent individuals were randomly assigned to 1 of 3 conditions delivered for 12 weeks: (1) standard naltrexone treatment, given 3 times a week; (2) naltrexone treatment plus contingency management (CM), with delivery of vouchers contingent on naltrexone compliance and drug-free urine specimens; or (3) naltrexone treatment, CM, plus significant other involvement (SO), where a family member was invited to participate in up to 6 family counseling sessions. Principal outcomes were retention in treatment, compliance with naltrexone therapy, and number of drug-free urine specimens.

RESULTS

First, CM was associated with significant improvements in treatment retention (7.4 vs 5.6 weeks; P =.05) and in reduction in opioid use (19 vs 14 opioid-free urine specimens; P =.04) compared with standard naltrexone treatment. Second, assignment to SO did not significantly improve retention, compliance, or substance abuse outcomes compared with CM. Significant effects for the SO condition over CM on retention, compliance, and drug use outcomes were seen only for the subgroup who attended at least 1 family counseling session. The SO condition was associated with significant (P =.02) improvements in family functioning.

CONCLUSIONS

Behavioral therapies, such as CM, can be targeted to address weaknesses of specific pharmacotherapies, such as noncompliance, and thus can play a substantial role in broadening the utility of available pharmacotherapies.

Substance P belongs to a group of neurokinins (NKs), small peptides that are broadly distributed in the central nervous system (CNS) and peripheral nervous system (PNS). The biological effects of substance P in the CNS, namely regulation of affective behavior and emesis in the brain and nociception in the spinal cord, are mediated by its binding to the NK1 receptor. The substance P-NK1 (SP-NK1) receptor system is the most extensively studied NK pathway, and in contrast to receptors for other neurotransmitters, such as glutamate, which have high expression throughout the CNS, only a minority of neurons (5% to 7%) in certain CNS areas express the NK1 receptor. The NK1 receptor is distributed in the plasma membrane of cell bodies and dendrites of unstimulated neurons, but upon substance P binding, the NK1 receptor undergoes rapid internalization, followed by rapid recycling to the plasma membrane. Release of substance P is induced by stressful stimuli, and the magnitude of its release is proportional to the intensity and frequency of stimulation. More potent and more frequent stimuli allow diffusion of substance P farther from the site of release, allowing activation of an approximately 3- to 5-times greater number of NK1 receptor-expressing neurons. Recent studies employing pharmacologic or genetic inactivation of NK1 receptors demonstrate the important role of the SP-NK1 receptor system in the regulation of affective behavior and suggest that inhibition of this pathway may be a useful approach to treatment of depression and associated anxiety.

1. Physical activity is known to increase type I collagen synthesis measured as the concentration of biomarkers in plasma. By the use of microdialysis catheters with a very high molecular mass cut-off value (3000 kDa) we aimed to determine local type I collagen synthesis and degradation in the peritendinous region by measuring interstitial concentrations of a collagen propeptide (PICP; 100 kDa) and a collagen degradation product (ICTP; 9 kDa) as well as an inflammatory mediator (PGE2). 2. Seven trained human runners were studied before and after (2 and 72 h) 3 h of running (36 km). Two microdialysis catheters were placed in the peritendinous space ventral to the Achilles' tendon under ultrasound guidance and perfused with a Ringer-acetate solution containing 3H-labelled human type IV collagen and [15-3H(N)]PGE2 for in vivo recovery determination. Relative recovery was 37-59 % (range of the s.e.m. values) for both radioactively labelled substances. 3. PICP concentration decreased in both interstitial peritendinous tissue and arterial blood immediately after exercise, but rose 3-fold from basal 72 h after exercise in the peritendinous tissue (55 +/- 10 microg l-1, mean +/- s.e.m. (rest) to 165 +/- 40 microg l-1 (72 h), P < 0.05) and by 25 % in circulating blood (160 +/- 10 microg l-1 (rest) to 200 +/- 12 microg l-1 (72 h), P < 0.05). ICTP concentration did not change in blood, but decreased transiently in tendon-related tissue during early recovery after exercise only. PGE2 concentration increased in blood during running, and returned to baseline in the recovery period, whereas interstitial PGE2 concentration was elevated in the early recovery phase. 4. The findings of the present study indicate that acute exercise induces increased formation of type I collagen in peritendinous tissue as determined with microdialysis and using dialysate fibre with a very high molecular mass cut-off. This suggests an adaptation to acute physical loading also in non-bone-related collagen in humans.

Although microglial activation may be an initial beneficial response to a variety of insults, prolonged activation can release toxic substances and lead to cell death. Microglial activation secondary to hypoxia-ischemia and/or infection in immature cerebral white matter is important in the pathogenesis of periventricular leukomalacia (PVL), the major pathological substrate of cerebral palsy in the premature infant. We hypothesize that a transient overexpression in activated microglial density occurs normally in the cerebral white matter of the human fetus during the peak window of vulnerability for PVL. Such an increase could render this region susceptible to insults that cause prolonged microglial activation, as conceptualized in PVL. To examine the developmental profile of microglia in the human fetus and infant brain, immunocytochemistry with microglial specific markers were used in 23 control (non-PVL) cases ranging from 20 to 183 postconceptional (PC) weeks. Tomato lectin, used to identify microglial morphology, revealed that the cerebral white matter of the human fetus and infant is densely populated with intermediate and amoeboid microglia; the latter is indicative of an activated state. Quantitative analysis with CD68 showed increased density of activated microglia in the cerebral white matter of the fetus (<37 PC weeks) relative to the neonate/infant >> or =37 PC weeks) and to the overlying cortex of either age group (P = 0.01). The primary finding of a transient, developmental-dependent overabundance of CD68-activated microglia in the cerebral white matter of the fetus suggests a potential "priming" of this area for diverse brain insults characterized by activation of microglia, particularly PVL. J.

Angiotensin I converting enzyme (ACE) and neutral endopeptidase ("enkephalinase"; NEP), were purified to homogeneity from human kidney. NEP cleaved substance P (SP) at Gln6-Phe7,-Phe8, and Gly9-Leu10 and neurotensin (NT) at Pro10-Tyr11 and Tyr11-Ile12. NEP hydrolyzed 0.1 mM SP, NT and their C-terminal fragments at the following rates (mumol/min/mg): SPPPPPPurified ACE rapidly inactivated SP as measured in bioassay. HPLC analysis showed that ACE cleaved SP at Phe8-Gly9 and Gly9-Leu10 to release C-terminal tri- and dipeptide (ratio = 4:1). The hydrolysis was Cl- dependent and inhibited by captopril. ACE released mainly C-terminal tripeptide from SP methyl ester, but only dipeptide from SP free acid. Modification of arginine residues in ACE with cyclohexanedione or butanedione similarly inhibited hydrolysis of SP, bradykinin and Bz-Gly-Phe-Arg (80-93%) indicating an active site arginine is required for hydrolysis of SP. ACE hydrolyzed NT at Tyr11-Ile12 to release Ile12-Leu13. SP, NT and their derivatives (0.1 mM) were cleaved by ACE at the following rates (mumol/min/mg): SPP methyl ester = 0.7, SP free acid = 8.5, SPPPPPeptide substrates were used as inhibitors of ACE (substrate = FA-Phe-Gly-Gly) and NEP (substrate = Leu5-enkephalin).(ABSTRACT TRUNCATED AT 250 WORDS)

Saliva contains a large number of proteins that participate in the protection of oral tissue. We found, for the first time, small vesicles (30-130 nm in diameter) in human whole saliva. Vesicles from saliva were identified by electron microscopy after isolation by gel-filtration on Sepharose CL-4B. They resemble exosomes, which are vesicles with an endosome-derived limiting membrane that are secreted by a diverse range of cell types. We performed a biochemical characterization of these vesicles by amino acid sequence analysis and Western blot analysis. We found that they contain dipeptidyl peptidase IV (DPP IV), galectin-3 and immunoglobulin A, which have potential to influence immune response. The DPP IV in the vesicles was metabolically active in cleaving substance P and glucose-dependent insulinotropic polypeptide to release N-terminal dipeptides. Our results demonstrate that human whole saliva contains exosome-like vesicles; they might participate in the catabolism of bioactive peptides and play a regulatory role in local immune defense in the oral cavity.

An amidohydrolase activity present in rat brain microsomes catalyzes the hydrolysis of N-arachidonoyl-[3H]ethanolamine ([3H]anandamide), an endogenous cannabimimetic substance, forming [3H]ethanolamine and arachidonic acid. Amidohydrolase activity is maximal at pH 6 and 8, is independent of divalent cations, has an apparent Km for [3H]anandamide of 12.7 +/- 1.8 microM, and has a Vmax of 5630 +/- 200 pmol/min/mg of protein. Phenylmethylsulfonyl fluoride, a serine protease inhibitor, and p-bromophenacyl bromide, a histidine-alkylating reagent, inhibit the activity, whereas N-ethylmaleimide and various nonselective peptidase inhibitors (EDTA, o-phenanthroline, bacitracin) have no effect. Brain amidohydrolase activity exhibits high substrate specificity for [3H]anandamide; N-gamma-linolenoyl-, N-homo-gamma-linolenoyl-, and N-11,14-eicosadienoyl- are hydrolyzed at markedly slower rates. Moreover, N-11-eicosaenoyl- and N-palmitoyl-[3H]ethanolamine are not hydrolyzed. [3H]Anandamide hydrolysis is inhibited competitively by nonradioactive anandamide and by other N-acylethanolamines with the following rank order of potency: anandamide>> N-linoleoyl- = N-cis-linolenoyl- = N-gamma-linolenoyl- = N- homo-gamma-linolenoyl->> N-11,14-eicosadienoyl->> N-oleoyl->> N- docosahexaenoyl->> N-docosatetraenoyl>> N-linoelaidoyl->> N-eicosaenoyl->> N- palmitoyl>> or = N-elaidoyl- = N-eicosanoyl-ethanolamine = no effect. Amidohydrolase activity is high in liver and brain and low in heart, kidney, intestine, stomach, lung, spleen, and skeletal muscle. Within the central nervous system, highest activity is found in globus pallidus and hippocampus, two regions rich in cannabinoid receptors, and lowest activity is found in brainstem and medulla, where cannabinoid receptors are sparse. The results, showing that brain amidohydrolase activity is selective for anandamide and enriched in areas of the central nervous system with high density of cannabinoid receptors, suggest that this activity may participate in the inactivation of anandamide at its sites of action.

A brief overview of recent developments in the substance P field is provided, in addition to a historical introduction. It is emphasized that there are multiple tachykinins and tachykinin receptors and that there are examples of coexistence of several tachykinin peptides and of several tachykinin receptors in single cells, and there is evidence for tachykininergic cotransmission. The distribution and functional significance of tachykinins in the gastrointestinal tract and in sensory neurones, and interactions with other peptides and transmitters, are reviewed. The recent production of knock-out mice for either substance P or the NK1 receptor is discussed, as well as the exciting concept of substance P receptor internalization. Finally, the development of specific substance P antagonists is summarized, and possible clinical implications discussed, and, in particular, a recent study which reports that a substance P antagonist shows clinical efficacy in depression.

Traditional Chinese medicine (TCM) has been practiced for thousands of years, but only within the last few decades has its use become more widespread outside of Asia. Concerns continue to be raised about the efficacy, legality, and safety of many popular complementary alternative medicines, including TCMs. Ingredients of some TCMs are known to include derivatives of endangered, trade-restricted species of plants and animals, and therefore contravene the Convention on International Trade in Endangered Species (CITES) legislation. Chromatographic studies have detected the presence of heavy metals and plant toxins within some TCMs, and there are numerous cases of adverse reactions. It is in the interests of both biodiversity conservation and public safety that techniques are developed to screen medicinals like TCMs. Targeting both the p-loop region of the plastid trnL gene and the mitochondrial 16S ribosomal RNA gene, over 49,000 amplicon sequence reads were generated from 15 TCM samples presented in the form of powders, tablets, capsules, bile flakes, and herbal teas. Here we show that second-generation, high-throughput sequencing (HTS) of DNA represents an effective means to genetically audit organic ingredients within complex TCMs. Comparison of DNA sequence data to reference databases revealed the presence of 68 different plant families and included genera, such as Ephedra and Asarum, that are potentially toxic. Similarly, animal families were identified that include genera that are classified as vulnerable, endangered, or critically endangered, including Asiatic black bear (Ursus thibetanus) and Saiga antelope (Saiga tatarica). Bovidae, Cervidae, and Bufonidae DNA were also detected in many of the TCM samples and were rarely declared on the product packaging. This study demonstrates that deep sequencing via HTS is an efficient and cost-effective way to audit highly processed TCM products and will assist in monitoring their legality and safety especially when plant reference databases become better established.