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Assessing the rate of drug delivery to the central nervous system (CNS) in vitro has been used for decades to predict whether CNS drug candidates are likely to attain their pharmacological targets, located within the brain parenchyma, at an effective dose. The predictive value of in vitro blood-brain barrier (BBB) models is therefore frequently assessed by comparing in vitro BBB permeability, usually quoted as the endothelial permeability coefficient (P e ) or apparent permeability (P app ), to their rate of BBB permeation measured in vivo, the latter being commonly assessed in rodents. In collaboration with AstraZeneca (DMPK department, Södertälje, Sweden), the in vitro BBB permeability (P app and P e ) of 27 marketed CNS drugs has been determined using a bovine in vitro BBB model and compared to their in vivo permeability (P vivo ), obtained by rat in-situ brain perfusion. The latter was taken from published data from Summerfield et al. (2007). This comparison confirmed previous reports, showing a strong in vitro/in vivo correlation for hydrophilic compounds, characterized by low brain tissue binding and a weak correlation for lipophilic compounds, characterized by high brain tissue binding. This observation can be explained by the influence of brain tissue binding on the uptake of drugs into the CNS in vivo and the absence of possible brain tissue binding in vitro. The use of glial cells (GC) in the in vitro BBB model to mimic brain tissue binding and the introduction of a new calculation method for in vitro BBB permeability (P vitro ) resulted in a strong correlation between the in vitro and in vivo rate of BBB permeation for the whole set of compounds. These findings might facilitate further in vitro to in vivo extrapolation for CNS drug candidates., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Positron emission tomography microdosing of radiolabeled drugs allows for noninvasive studies of organ exposure in vivo. The aim of the present study was to examine and compare the brain exposure of 12 commercially available CNS drugs and one non-CNS drug., Methods: The drugs were radiolabeled with (11)C (t 1/2 = 20.4 minutes) and examined using a high resolution research tomograph. In cynomolgus monkeys, each drug was examined twice. In rhesus monkeys, a first positron emission tomography microdosing measurement was repeated after preadministration with unlabeled drug to examine potential dose-dependent effects on brain exposure. Partition coefficients between brain and plasma (KP) were calculated by dividing the AUC0-90 min for brain with that for plasma or by a compartmental analysis (VT). Unbound KP (KP u,u) was obtained by correction for the free fraction in brain and plasma., Results: After intravenous injection, the maximum radioactivity concentration (C max, %ID) in brain ranged from 0.01% to 6.2%. For 10 of the 12 CNS drugs, C max, %ID was >2%, indicating a preferential distribution to brain. A lower C max, %ID was observed for morphine, sulpiride, and verapamil. K P ranged from 0.002 (sulpiride) to 68 (sertraline) and 7 of 13 drugs had KP u,u close to unity. For morphine, sulpiride, and verapamil, K P u,u was <0.3, indicating impaired diffusion and/or active efflux. Brain exposure at microdosing agreed with pharmacological dosing conditions for the investigated drugs., Conclusions: This study represents the largest positron emission tomography study on brain exposure of commercially available CNS drugs in nonhuman primates and may guide interpretation of positron emission tomography microdosing data for novel drug candidates., (© The Author 2015. Published by Oxford University Press on behalf of CINP.)

Historically, the focus has been to use in vitro BBB models to optimize rate of drug delivery to the CNS, whereas total in vivo brain/plasma ratios have been used for optimizing extent. However, these two parameters do not necessarily show good correlations with receptor occupancy data or other pharmacological readouts. In line with the free drug hypothesis, the use of unbound brain concentrations (Cu,br) has been shown to provide the best correlations with pharmacological data. However, typically the determination of this parameter requires microdialysis, a technique not ideally suited for screening in early drug development. Alternative, and less resource-demanding methodologies to determine Cu,br employ either equilibrium dialysis of brain homogenates or incubations of brain slices in buffer to determine fraction unbound brain (fu,br), which is subsequently multiplied by the total brain concentration to yield Cu,br. To determine Cu,br/Cu,pl ratios this way, still requires both in vitro and in vivo experiments that are quite time consuming. The main objective of this study was to explore the possibility to directly generate Cu,br/Cu,pl ratios in a single in vitro model of the BBB, using a co-culture of brain capillary endothelial and glial cells in an attempt to mimick the in vivo situation, thereby greatly simplifying existing experimental procedures. Comparison to microdialysis brain concentration profiles demonstrates the possibility to estimate brain exposure over time in the BBB model. A stronger correlation was found between in vitro Cu,br/Cu,pl ratios and in vivo Cu,br/Cu,pl obtained using fu,br from brain slice than with fu,br from brain homogenate for a set of 30 drugs. Overall, Cu,br/Cu,pl ratios were successfully predicted in vitro for 88% of the 92 studied compounds. This result supports the possibility to use this methodology for identifying compounds with a desirable in vivo response in the CNS early on in the drug discovery process.

Background: Modelling the blood-CNS barriers of the brain and spinal cord in vitro continues to provide a considerable challenge for research studying the passage of large and small molecules in and out of the central nervous system, both within the context of basic biology and for pharmaceutical drug discovery. Although there has been considerable success over the previous two decades in establishing useful in vitro primary endothelial cell cultures from the blood-CNS barriers, no model fully mimics the high electrical resistance, low paracellular permeability and selective influx/efflux characteristics of the in vivo situation. Furthermore, such primary-derived cultures are typically labour-intensive and generate low yields of cells, limiting scope for experimental work. We thus aimed to establish protocols for the high yield isolation and culture of endothelial cells from both rat brain and spinal cord. Our aim was to optimise in vitro conditions for inducing phenotypic characteristics in these cells that were reminiscent of the in vivo situation, such that they developed into tight endothelial barriers suitable for performing investigative biology and permeability studies., Methods: Brain and spinal cord tissue was taken from the same rats and used to specifically isolate endothelial cells to reconstitute as in vitro blood-CNS barrier models. Isolated endothelial cells were cultured to expand the cellular yield and then passaged onto cell culture inserts for further investigation. Cell culture conditions were optimised using commercially available reagents and the resulting barrier-forming endothelial monolayers were characterised by functional permeability experiments and in vitro phenotyping by immunocytochemistry and western blotting., Results: Using a combination of modified handling techniques and cell culture conditions, we have established and optimised a protocol for the in vitro culture of brain and, for the first time in rat, spinal cord endothelial cells. High yields of both CNS endothelial cell types can be obtained, and these can be passaged onto large numbers of cell culture inserts for in vitro permeability studies. The passaged brain and spinal cord endothelial cells are pure and express endothelial markers, tight junction proteins and intracellular transport machinery. Further, both models exhibit tight, functional barrier characteristics that are discriminating against large and small molecules in permeability assays and show functional expression of the pharmaceutically important P-gp efflux transporter., Conclusions: Our techniques allow the provision of high yields of robust sister cultures of endothelial cells that accurately model the blood-CNS barriers in vitro. These models are ideally suited for use in studying the biology of the blood-brain barrier and blood-spinal cord barrier in vitro and for pre-clinical drug discovery.

Myeloperoxidase (MPO) is a prime candidate for promoting oxidative stress during inflammation. This abundant enzyme of neutrophils uses hydrogen peroxide to oxidize chloride to highly reactive and toxic chlorine bleach. We have identified 2-thioxanthines as potent mechanism-based inactivators of MPO. Mass spectrometry and x-ray crystal structures revealed that these inhibitors become covalently attached to the heme prosthetic groups of the enzyme. We propose a mechanism whereby 2-thioxanthines are oxidized, and their incipient free radicals react with the heme groups of the enzyme before they can exit the active site. 2-Thioxanthines inhibited MPO in plasma and decreased protein chlorination in a mouse model of peritonitis. They slowed but did not prevent neutrophils from killing bacteria and were poor inhibitors of thyroid peroxidase. Our study shows that MPO is susceptible to the free radicals it generates, and this Achilles' heel of the enzyme can be exploited to block oxidative stress during inflammation.

Moving to Opportunity (MTO) offered public housing residents the opportunity to move to low-poverty neighborhoods. Several years later, boys in the experimental group fared no better on measures of risk behavior than their control group counterparts, whereas girls in the experimental group engaged in lower-risk behavior than control group girls. The authors explore these differences by analyzing data from in-depth interviews conducted with 86 teens in Baltimore and Chicago. They find that daily routines, fitting in with neighborhood norms, neighborhood navigation strategies, interactions with peers, friendship making, and distance from father figures may contribute to how girls who moved via MTO benefited more than boys.

NXY-059, a polar compound with limited transport across the blood-brain barrier, has demonstrated neuroprotection in several animal models of acute ischemic stroke but failed to confirm clinical benefit in the second phase III trial (SAINT-II). To improve the understanding of the mechanisms responsible for its neuroprotective action in preclinical models a series of experiments was carried out in an in vitro blood-brain barrier (BBB) model. A clinically attainable concentration of 250 mumol/L of NXY-059 administered at the onset or up to 4 h after oxygen glucose deprivation (OGD) produced a significant reduction in the increased BBB permeability caused by OGD. Furthermore, OGD produced a huge influx of tissue plasminogen activator across the BBB, which was substantially reduced by NXY-059. This study suggests that the neuroprotective effects of NXY-059 preclinically, may at least in part be attributed to its ability to restore functionality of the brain endothelium.

There is a growing interest to use in vitro BBB cell assays in early safety assessment of compounds. By modifying a well-validated co-culture model of brain capillary endothelial and glial cells, developed by Dehouck et al. [Dehouck, M.P., Meresse, S., Delorme, P., Fruchart, J.C., Cecchelli, R., 1990. An easier, reproducible, and mass-production method to study the blood-brain barrier in vitro. Journal of Neurochemistry 54 (5), 1798-1801], it has been possible to develop a new in vitro BBB system suitable for high throughput screening (HTS). In addition, this new procedure substantially reduces the use of experimental animals and considerably facilitates the process of obtaining a functional in vitro BBB model. The model is ready to use after only 4 days of culture and then shows the typical expression and localization of tight junction proteins. The function of the P-glycoprotein and the transcriptional expression of other efflux transporters such as MRP 1, 4 and 5 have been demonstrated. In addition, the model produces a good in vitro/in vivo correlation for 10 compounds (R2=0.81). Furthermore, studies were undertaken within the European ACuteTox consortium with the objective to assess BBB toxicity and make risk assessments of potentially toxic compounds according to their predicted ability to reach the CNS compartment. These investigations demonstrated that the results produced in the HTS BBB model were similar to the standard co-culture model.

The market for neuropharmaceuticals is potentially one of the largest sectors of the global pharmaceutical market owing to the increase in average life expectancy and the fact that many neurological disorders have been largely refractory to pharmacotherapy. The brain is a delicate organ that can efficiently protect itself from harmful compounds and precisely regulate its microenvironment. Unfortunately, the same mechanisms can also prove to be formidable hurdles in drug development. An improved understanding of the regulatory interfaces that exist between blood and brain may provide novel and more effective strategies to treat neurological disorders.

The penetration of the free radical trapping neuroprotectant NXY-059 into the brain has been examined in rats subjected to permanent middle cerebral artery occlusion (pMCAO). NXY-059 (125 mg/kg bolus followed by 125 mg/kg/h) was infused for 4 h 45 min starting 15 min after right pMCAO or sham operation. At 5 h, there was a similar plasma total NXY-059 concentration (micromol/L) in both groups [sham: 623 +/- 44 (6); pMCAO: 605 +/- 43 (5)] and a similar drug concentration (nmol/g) in the right cortex [sham: 6.92 +/- 1.05 (6); pMCAO: 6.14 +/- 2.18 (6)]. A subsequent experiment in normal rats, infusing NXY-059 at both a similar and higher concentration (252 mg/kg bolus and 252 mg/kg/h), demonstrated that the concentration of NXY-059 in cortex increased linearly with respect to the plasma concentration. These data demonstrate that NXY-059 does penetrate brain tissue in control animals and ischemic tissue of animals subjected to pMCAO.

A complication of percutaneous endoscopic gastrostomy (PEG) is perforation of a hollow viscus. This is typically detected by finding of pneumoperitoneum (PP) on radiographs. However, PP can occasionally be a benign finding. A review of the literature shows many causes for a benign PP, and it has been noted to occur frequently after PEG placement. In the absence of signs or symptoms of peritoneal inflammation, PP usually requires no further investigation or treatment.

The free radical trapping nitrone compounds alpha-phenyl-N-tert-butylnitrone (PBN), 2-sulfophenyl-N-tert-butylnitrone (S-PBN) and disodium 2,4-disulfophenyl-N-tert-butyl nitrone (NXY-059) are effective neuroprotective agents in experimental models of both transient and permanent focal ischemia. A recent in vivo study suggested that NXY-059 had poor brain uptake in a transient ischemia model. We have now examined its blood-brain barrier permeability and cerebral endothelial uptake during hypoxic and ischemic conditions using an in vitro model of the blood-brain barrier. The in vitro blood-brain barrier permeability and cerebral endothelial uptake of NXY-059 and S-PBN were low during normoxic conditions. In contrast, PBN had very high blood-brain barrier penetration in vitro which confirmed earlier in vivo results. The permeability of [14C]NXY-059 increased 3.5 times after 9 h of hypoxia or 3 h of ischemia. There was, respectively, a 5-fold and more than 10-fold increase, after 6 and 9 h of ischemia. The control molecule [3H]inulin (M(r) approximately 5000) showed a similar increase in permeability under the same experimental conditions indicating a major change in the transport properties of the endothelium. There was a 60% reduction in the ATP levels of astrocytes after 3 h of ischemia and a 90% reduction after 9 h. The reduction in ATP levels in endothelial cells was somewhat lower. The uptake of NXY-059 in cerebral endothelial cells under normoxic, hypoxic or 9 h of ischemic conditions was negligible. NXY-059, S-PBN and PBN showed no effects on vesicular transport or the integrity of the blood-brain barrier in normoxic or ischemic conditions, nor did the compounds induce any change in the ATP levels of the cells. In conclusion, it is possible that the increase in blood-brain barrier permeability of [14C]NXY-059 which occurs during prolonged ischemia in vitro reflects a change which may be of importance to the neuroprotective effects of this nitrone free radical trapping agent.

Purpose: Studies were conducted to evaluate whether the use of an in vitro model of the blood-brain barrier (BBB) resulted in more accurate predictions of the in vivo transport of compounds compared to the use of a human intestinal cell line (Caco-2)., Methods: The in vitro BBB model employs bovine brain capillary endothelial cells co-cultured with primary rat astrocytes. The Caco-2 cells originate from a human colorectal carcinoma. The rat was used as experimental animal for the in vivo studies., Results: Strong correlations (r = 0.93-0.95) were found between the results generated by the in vitro model of the BBB and two different methodologies to measure the permeability across the BBB in vivo. In contrast, a poor correlation (r = 0.68) was obtained between Caco-2 cell data and in vivo BBB transport. A relatively poor correlation (r = 0.74) was also found between the two in vitro models., Conclusion: The present study illustrates the limitations of the Caco-2 model to predict BBB permeability of compounds in vivo. The results emphasize the fact that the BBB and the intestinal mucosa are two fundamentally different biologic barriers, and to be able to make accurate predictions about the in vivo CNS penetration of potential drug candidates, it is important that the in vitro model possesses the main characteristics of the in vivo BBB.

Successful drug delivery to the central nervous system (CNS) is highly dependent on DMPK as well as physicochemical properties and it is therefore important to characterise these properties and take them into account when designing chemical lead series that act at CNS targets. Since the drug discovery/development process is becoming increasingly focused on reducing the time required to enter molecules into the market, industrial DMPK scientists have emerged from their traditional supportive role in drug development to provide valuable support in the drug discovery process, using novel methods to meet the demands of combinatorial chemistry and bioscience groups.

Diseases of the foreskin, penis, and urethra are uncommon conditions that constitute a small percentage of emergency department visits. However, it is important that the emergency physician recognize and treat these conditions and understand which conditions require urologic consultation. Rapid and proper treatment in the emergency department can alleviate patient discomfort, increase patient satisfaction, and prevent future morbidity.

Objectives: To assess the prevalence of elevated blood lead levels (EBLLs) among children before and after foster care placement, and to compare the prevalence of EBLLs among children in foster care with that of their siblings and the general population., Methods: We conducted a retrospective cohort study using administrative databases from the Philadelphia Department of Human Services and the Birth Certificate Registry and the Childhood Lead Poisoning Prevention Program at the Philadelphia Department of Public Health. Logistic regression analyses were performed to control for confounding variables, including age, race, gender, and the year, seasonal timing, and source (capillary vs venous) of test., Results: From June 1992 to May 1997, there were 1824 children in foster care with available blood lead results in the Childhood Lead Poisoning Prevention Program database. Of these, 519 (28%) had initial lead screening before foster care placement and 654 (36%) after placement. There were 821 siblings and 73 608 children in the general population with available blood lead results. Before entering foster care, children were nearly twice as likely to have EBLLs as their siblings (adjusted odds ratio [OR] = 1.7; 95% confidence interval [CI] = 1.4, 2.0), those in placement (adjusted OR = 1.9; 95% CI = 1.6, 2.2), and the general population (adjusted OR = 1.7; 95% CI = 1.5, 2.0). At the highest point prevalence, 50% of children before placement had lead levels >/=20 microg/dL, and nearly 90% had levels >/=10 microg/dL. For all age categories, siblings of children in foster care placement had a higher prevalence of EBLLs than did the general population. After placement, children in foster care were nearly half as likely as the other groups to have EBLLs., Conclusions: Our findings suggest that children are at high risk for lead poisoning before entering foster care and that placement in foster care may have a beneficial effect on lead exposure. Children before foster care placement are nearly twice as likely to have EBLLs compared with children in foster care placement, the general population, and their siblings. Furthermore, siblings of children in foster care are at high risk for lead poisoning. Children receiving social services in their own homes and children suffering from abuse and neglect should be actively screened for lead poisoning. Greater efforts at preventing lead poisoning among these children must be made.

Due to increasing financial pressure on maintaining and improving pharmacy services, there is a need for productivity data and time distribution among different activities for pharmacy practice residents and their faculty preceptors (pharmacotherapists). This study measured the clinical productivity of 13 residents and 25 pharmacotherapists for a 14-day period. The study identified the average time (minutes) and frequency spent each day on categories of activities, which included direct patient care, chart use, rounds, professional encounter, teaching, research, and administration. Results showed that the productivity profiles for residents and pharmacotherapists were comparable. Findings were utilized to defend the educational programs for the department and as a baseline for periodic monitoring of the productivity of these programs.

Genetic recombination involves classical crossing-over and gene conversion (aberrant segregation). In fungi that produce an ascus containing four spores, a gene conversion event is manifested as 3:1 or 1:3 (or more rarely 4:0 or 0:4) segregations, in contrast to the normal mendelian 2:2 segregation. Polarity is one of the properties of gene conversion; in almost all cases the frequency of conversion exhibits a gradient across the gene monitored. The frequency of conversion is usually independent of the specific allele used as a marker, but dependent on its location. An interpretation of conversion polarity is that it is caused by the existence of specific initiation sites for meiotic recombination, located at the high end of the polarity gradient. Here we show that the polarity gradient for the HIS2 gene of Saccharomyces cerevisiae is high at the 3' end of the gene, implying that the promoter of HIS2 is not the initiation site.

With increasing competition among hospitals, primary care referral development and management programs offer an opportunity for hospitals to increase their admissions. Such programs require careful development, the commitment of the hospital staff to the strategy, an integration of hospital activities, and an understanding of medical practice management.

Eating styles of 30 obese women and 37 matched women of normal weight were observed unobtrusively in a fast-food restaurant in the search for an "obese eating style" or other differences between the two groups. The size and character of the food was carefully matched by giving each woman a coupon entitling her to a free meal of either 985 or 1,800 calories. There were only small and inconsistent differences between obese and normal-weight women. No evidence of an "obese eating style" was found.

An experimental model has been developed in the pig in order to evaluate the effect of percutaneous transluminal angioplasty on fibrous renal artery stenosis. Stenoses were induced by bilateral subtotal ligation with chromic catgut. Six weeks following ligation unilateral transluminal dilatation was performed in 9 pigs using Grüntzig balloon catheters. In only one case a serious complication occurred with rupture of the renal artery. Seven pigs with dilated stenoses were re-examined by angiography 8 to 10 weeks after dilatation. In 6 cases the stenoses were practically completely eliminated while in one cases a re-stenosis developed. Microscopy of the dilated arteries revealed several small and large ruptures of th internal elastic membrane.

The effect of changes in weather on headache was investigated in two separate studies in Uppsala. In a pilot study 60 healthy persons (all medical staff) reported their headache during a period of 2 months. Later 53 patients with migraine and 20 with tension headache registered their headache during 4 weeks. Several climate factors were recorded during the observation periods. Both studies showed a significant weekly periodicity of headache. In patients with migraine the headache frequency was lowest on Sunday-Monday and highest on Thursday and on Saturday. In patients with tension headache the highest frequency was noted on Thursday-Friday and lowest frequency on Sunday-Tuesday. The pilot study showed that personnel off work had significantly more headache when bad weather occurred. There was a tendency for headache the day before windy weather. In the 53 migraine patients there was also a correlation between some climate factors and headache frequency. In this group the deviation from the expected headache frequency (actual headache frequency minus that of observed weekly periodicity) correlated significantly to the air-pressure and air-temperature measured 1-3 days later.