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Amino acids, metabolized by host cells as well as commensal gut bacteria, have signaling effects on host metabolism. Oral supplementation of the essential amino acid histidine has been shown to exert metabolic benefits. To investigate whether dietary histidine aids glycemic control, we performed a case-controlled parallel clinical intervention study in participants with type 2 diabetes (T2D) and healthy controls. Participants received oral histidine for seven weeks. After 2 weeks of histidine supplementation, the microbiome was depleted by antibiotics to determine the microbial contribution to histidine metabolism. We assessed glycemic control, immunophenotyping of peripheral blood mononucelar cells (PBMC), DNA methylation of PBMCs and fecal gut microbiota composition. Histidine improves several markers of glycemic control, including postprandial glucose levels with a concordant increase in the proportion of MAIT cells after two weeks of histidine supplementation. The increase in MAIT cells was associated with changes in gut microbial pathways such as riboflavin biosynthesis and epigenetic changes in the amino acid transporter SLC7A5. Associations between the microbiome and MAIT cells were replicated in the MetaCardis cohort. We propose a conceptual framework for how oral histidine may affect MAIT cells via altered gut microbiota composition and SLC7A5 expression in MAIT cells directly and thereby influencing glycemic control. Future studies should focus on the role of flavin biosynthesis intermediates and SLC7A5 modulation in MAIT cells to modulate glycemic control.

The human absorption, distribution, metabolism, and excretion (hADME) study is the cornerstone of the clinical pharmacology package for small molecule drugs, providing comprehensive information on the rates and routes of disposition and elimination of drug-related material in humans through the use of 14 C-labeled drug. Significant changes have already been made in the design of the hADME study for many companies, but opportunity exists to continue to re-think both the design and timing of the hADME study in light of the potential offered by newer technologies, that enable flexibility in particular to reducing the magnitude of the radioactive dose used. This paper provides considerations on the variety of current strategies that exist across a number of pharmaceutical companies and on some of the ongoing debates around a potential move to the so called "human first/human only" approach, already adopted by at least one company. The paper also provides a framework for continuing the discussion in the application of further shifts in the paradigm., (© 2022 GSK & Roche, et al. Clinical Pharmacology & Therapeutics published by Wiley Periodicals LLC on behalf of American Society for Clinical Pharmacology and Therapeutics.)

Background and Objective: Omeprazole is a proton pump inhibitor that is used in acid suppression therapy in infants. Infants cannot swallow the oral tablets or capsules. Since, infants require a non-standard dose of omeprazole, the granules or tablets are often crushed or suspended in water or sodium bicarbonate, which may destroy the enteric coating. In this study we explore the efficacy and pharmacokinetics of rectally administered omeprazole in infants with gastroesophageal reflux disease (GERD) due to esophageal atresia (EA) or congenital diaphragmatic hernia (CDH) and compare these with orally administered omeprazole., Methods: Infants (6-12 weeks postnatal and bodyweight > 3 kg) with EA or CDH and GERD were randomized to receive a single dose of 1 mg/kg omeprazole rectally or orally. The primary outcome was the percentage of infants for whom omeprazole was effective according to predefined criteria for 24-h intraesophageal pH. Secondary outcomes were the percentages of time that gastric pH was < 3 or < 4, as well as the pharmacokinetic parameters., Results: Seventeen infants, 4 with EA and 13 with CDH, were included. The proportion of infants for whom omeprazole was effective was 56% (5 of 9 infants) after rectal administration and 50% (4 of 8 infants) after oral administration. The total reflux time in minutes and percentages and the number of reflux episodes of pH < 4 decreased statistically significantly after both rectal and oral omeprazole administration. Rectal and oral administration of omeprazole resulted in similar serum exposure., Conclusions: A single rectal omeprazole dose (1 mg/kg) results in consistent increases in intraesophageal and gastric pH in infants with EA- or CDH-related GERD, similar to an oral dose. Considering the challenges with existing oral formulations, rectal omeprazole presents as an innovative, promising alternative for infants with pathological GERD., Clinical Trial Register: ClinicalTrials.gov Identifier: NCT00226044.

Background: Improvement of balance control is an important rehabilitation goal for patients with motor and sensory impairments. To quantify balance control during walking, various stability outcome measures have described differences between healthy controls and patient groups with balance problems. To be useful for the evaluation of interventions or monitoring of individual patients, stability outcome measures need to be reliable., Research Question: What is the test-retest reliability of six stability outcome measures during gait?, Methods: Patients with balance problems (n = 45) and healthy controls (n = 20) performed two times a two-minute walk test (2MWT). The intraclass correlation coefficient (ICC) and Bland-Altman analysis (coefficient of repeatability; CR) were used to evaluate the test-retest reliability of six stability outcome measures: dynamic stability margin (DSM), margin of stability (MoS), distance between the extrapolated centre of mass (XCoM) and centre of pressure (CoP) in anterior-posterior (XCoM-CoP AP ) and medial-lateral (XCoM-CoP ML ) direction, and inclination angle between centre of mass (CoM) and CoP in anterior-posterior (CoM-CoP AP-angle ) and medial-lateral (CoM-CoP ML-angle ) direction. A two way mixed ANOVA was performed to reveal measurement- and group-effects., Results: The ICCs of all stability outcome measures ranged between 0.51 and 0.97. Significant differences between the measurements were found for the DSM (p = 0.017), XCoM-CoP AP (p = 0.008) and CoM-CoP AP-angle (p = 0.001). Significant differences between controls and patients were found for all stability outcome measures (p < 0.01) except for the MoS (p = 0.32). For the XCoM-CoP distances and CoM-CoP angles, the CRs were smaller than the difference between patients and controls., Significance: Based on the ICCs, the reliability of all stability outcome measures was moderate to excellent. Since the XCoM-CoP ML and CoM-CoP ML-angle showed no differences between the measurements and smaller CRs than the differences between patients and controls, the XCoM-CoP ML and CoM-CoP ML-angle seem the most promising stability outcome measures to evaluate interventions and monitor individual patients., Competing Interests: Declaration of Competing Interest All authors declare that they do not have any conflict of interest. The authors state that there was no financial or personal relationship with other people or organizations that influenced the outcome of this study., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Stroke survivors often show reduced walking velocity and gait asymmetry. These gait abnormalities are associated with reduced propulsion of the paretic leg. This review aimed to provide an overview of the potential effectiveness of post-stroke rehabilitation interventions to improve paretic propulsion, ankle kinetics and walking velocity., Methods: A systematic search was performed in Pubmed, Web of Science, Embase, and Pedro. Studies were eligible if they reported changes in propulsion measures (impulse, peak value and symmetry ratios) or ankle kinetics (moment and power) following intervention in stroke survivors (group size ≥10). Study selection, data extraction and quality assessment were performed independently by two authors., Findings: A total of 28 studies were included, of which 25 studies applied exercise interventions, two studies focused on surgical interventions, and one on non-invasive brain stimulation. The number of high-quality trials was limited (N = 6; score Downs and Black scale ≥19). Propulsion measures were the primary outcome in eight studies. In general, mixed results were reported with 14 interventions yielding improvements in propulsion and ankle kinetics. In contrast, gains in walking velocity were observed in the vast majority of studies (N = 20 out of 23)., Interpretation: Interventions that yielded gains in propulsion appeared to have in common that they challenged and/or enabled the utilization of latent propulsive capacity of the paretic leg during walking. Walking speed generally increased, regardless of the observed change in propulsion, suggesting the use of compensatory mechanisms. Findings should, however, be interpreted with some caution, as the evidence base for this emerging focus of rehabilitation is limited., Competing Interests: Declaration of competing interest None of the involved authors have any conflicts of interest to report., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Many research groups have studied fall impact mechanics to understand how fall severity can be reduced to prevent hip fractures. Yet, direct impact force measurements with force plates are restricted to a very limited repertoire of experimental falls. The purpose of this study was to develop a generic model for estimating hip impact forces (i.e. fall severity) in in vivo sideways falls without the use of force plates. Twelve experienced judokas performed sideways Martial Arts (MA) and Block ('natural') falls on a force plate, both with and without a mat on top. Data were analyzed to determine the hip impact force and to derive 11 selected (subject-specific and kinematic) variables. Falls from kneeling height were used to perform a stepwise regression procedure to assess the effects of these input variables and build the model. The final model includes four input variables, involving one subject-specific measure and three kinematic variables: maximum upper body deceleration, body mass, shoulder angle at the instant of 'maximum impact' and maximum hip deceleration. The results showed that estimated and measured hip impact forces were linearly related (explained variances ranging from 46 to 63%). Hip impact forces of MA falls onto the mat from a standing position (3650±916N) estimated by the final model were comparable with measured values (3698±689N), even though these data were not used for training the model. In conclusion, a generic linear regression model was developed that enables the assessment of fall severity through kinematic measures of sideways falls, without using force plates., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The aim of this study was to explore the feasibility of the Mastication Observation and Evaluation (MOE) instrument, dynamic ultrasound and 3D kinematic measurements to describe mastication in children with spastic cerebral palsy and typically developing children. Masticatory movements during five trials of eating a biscuit were assessed in 8 children with cerebral palsy, spastic type (mean age 9.08years) and 14 typically developing children (mean age 9.01years). Differences between trials were tested (t-test) and the mastication of individual children with cerebral palsy was analyzed. MOE scores ranged from 17 to 31 (median 24) for the children with cerebral palsy and from 28 to 32 (median 31) for the typically developing children. There was an increased chewing cycle duration, a smaller left-right and up-down tongue displacement and larger anterior mandible movements for the trials (n=40) of cerebral palsy children (p<0.000 for all comparisons) compared to the trials of typically developing children (n=70). The MOE captures differences in mastication between individual children with cerebral palsy. The MOE items 'jaw movement' and 'fluency and coordination' showed the most similarity with the objective measurements. Objective measurements of dynamic ultrasound and 3D kinematics complemented data from the MOE instrument., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Background: Patients with differentiated thyroid carcinoma (DTC) have a favorable prognosis after treatment with thyroidectomy, radioiodine, and TSH suppression. However, treatment is associated with long-term cardiovascular toxicity. The aim of this study was to evaluate whether there is an increased risk of atrial fibrillation (AF) in DTC patients and whether AF occurrence is related to DTC treatment., Patients and Methods: Incident AF was compared between 518 DTC patients and 1563 matched controls. A cumulative incidence curve was plotted, and competing risk regression analyses with adjustment for all-cause mortality were performed. Within the DTC cohort, associations between time-varying DTC treatment variables and incident AF were analyzed., Results: For both cohorts, the mean age was 48.6 years (75% of subjects were women). The AF incidence rate was 6.2/1000 person-years for DTC patients and 2.7/1000 person-years for controls. DTC patients had a 2.25-fold (95% confidence interval [CI], 1.40-3.63) and 2.47-fold (95% CI, 1.55-3.95) increased AF risk in crude and fully adjusted analyses, respectively. Within the DTC cohort, the TSH level (which was suppressed in 85.7% of patients) was not associated with AF, whereas a higher cumulative radioiodine dose slightly increased AF risk: subdistribution hazard ratio, 1.04 (95% CI, 1.01-1.08) per 50 mCi (1.85 GBq) increase, after adjustment., Conclusion: Patients with DTC have an increased AF risk, independent from established AF risk factors. We could not demonstrate a relation between TSH and AF, whereas a higher cumulative radioiodine dose was associated with a slightly increased AF risk. Electrocardiogram screening for AF may be warranted during follow-up of DTC patients to allow early diagnosis and treatment of AF and to prevent its complications.

Despite adequate glycemic control, pregnancy outcome of women with type 1 diabetes (T1D) is still unfavorable as compared to healthy women. In a rat-model of T1D under normoglycemic conditions, adverse pregnancy outcome was also observed, which was associated with aberrant immunological adaptations to pregnancy. Because similar processes may occur in women with T1D we studied the systemic immune response in non-pregnant and pregnant women with and without T1D. The systemic immune response was assessed by using flow cytometry to evaluate the number and activational status of subpopulations of lymphocytes, Natural Killer cells and monocytes in peripheral blood of non-pregnant and pregnant women with and without T1D. An increased white blood cell count, an increased Th1/Th2 ratio, increased Natural Killer cell expression of CD335 and enhanced activation of intermediate and non-classical monocytes was observed in pregnant women with T1D vs. healthy pregnant women. Also, the pregnancy outcome (i.e. incidence of preterm delivery and macrosomia) of women with T1D was unfavorable as compared to healthy women. This study showed that in T1D, the immunological adaptations to pregnancy are disturbed. In addition to hyperglycemia, these different immunological adaptations may be responsible for the greater frequency of complications in pregnant women with T1D.

Femoral fractures are a major health issue. Most experimental and finite element (FE) fracture studies use polymethylmethacrylate cups on the greater trochanter (GT) to simulate fall impact loads. However, in vivo fall studies showed that the femur is loaded distally from the GT. Our objective was to incorporate in vivo fall data in FE models to determine the effects of loading position and direction, and size of simulated impact site on the fracture load and fracture type for a healthy and an osteoporotic femur. Twelve sets of loading position and angles were applied through 'near point loads' on the models. Additional simulations were performed with 'cup loads' on the GT, similar to the literature. The results showed no significant difference between fracture loads from simulations with near point loads distally from the GT and those with cup loads on the GT. However, simulated fracture types differed, as near point loads distally from the GT generally resulted in various neck fractures, whilst cup load simulations predicted superior neck and trochanteric fractures only. This study showed that incorporating in vivo fall assessments in FE models by loading the models distally from the GT results in prediction of realistic fracture loads and fracture types., (Copyright © 2015 IPEM. Published by Elsevier Ltd. All rights reserved.)

Objective. To assess the incidence of adverse pregnancy outcome in native and nonnative Dutch women with pregestational type 2 diabetes (T2D) in a multicenter study in The Netherlands. Methods. Maternal characteristics and pregnancy outcome were retrospectively reviewed and the influence of ethnicity on outcome was evaluated using independent t-test, Mann-Whitney U-test, and chi-square test. Results. 272 pregnant women (80 native and 192 non-native Dutch) with pregestational T2D were included. Overall outcome was unfavourable, with a perinatal mortality of 4.8%, major congenital malformations of 6.3%, preeclampsia of 11%, preterm birth of 19%, birth weight >90th percentile of 32%, and a Caesarean section rate of 42%. In nonnative Dutch women, the glycemic control was slightly poorer and the gestational age at booking somewhat later as compared to native Dutch women. However, there were no differences in incidence of preeclampsia/HELLP, preterm birth, perinatal mortality, macrosomia, and congenital malformations between those two groups. Conclusions. A high incidence of adverse pregnancy outcomes was found in women with pregestational T2D, although the outcome was comparable between native and non-native Dutch women. This suggests that easy access to and adequate participation in the local health care systems contribute to these comparable outcomes, offsetting potential disadvantages in the non-native group.

To investigate whether double-leg stance could reveal balance deficits in subjects with functional ankle instability (FAI) and whether such an assessment of static balance would be correlated with measures of dynamic instability, 16 individuals with FAI and 16 healthy controls participated in this study. Static postural control was tested using double-leg stance (either with the eyes open (EO) or closed (EC)) on a dual-plate force platform. Dynamic balance was evaluated using the Multiple Hop Test (MHT) and a weight-shifting task. FAI subjects were significantly less stable in the anteroposterior direction during double-leg stance (as assessed by velocity of centre of pressure, VCP), both for the EO and EC condition. In the mediolateral direction the VCP values were also higher in FAI, but significance was only found for the EC condition (p=.02). FAI subjects made significantly more balance errors compared to healthy controls (p<.001) on both the affected and less affected leg during MHT. There were no significant differences between FAI and healthy subjects during the weight-shifting task. No relationship was found between double-leg stance and MHT measures (all correlations (rs) less than .30). This study suggests that static postural control during double-leg stance is impaired in FAI subjects. Although dynamic balance during MHT is also affected, no significant relationship was found between static and dynamic measurements, which indicate that they are most probably related to different aspects of postural control., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Introduction: Despite tight glycemic control, pregnancy complication rate in type 1 diabetes patients is higher than in normal pregnancy. Other etiological factors may be responsible for the development of adverse pregnancy outcome. Acceptance of the semi-allogeneic fetus is accompanied by adaptations in the maternal immune-response. Maladaptations of the immune-response has been shown to contribute to pregnancy complications. We hypothesized that type 1 diabetes, as an autoimmune disease, may be associated with maladaptations of the immune-response to pregnancy, possibly resulting in pregnancy complications., Methods: We studied pregnancy outcome and pregnancy-induced immunological adaptations in a normoglycemic rat-model of type 1 diabetes, i.e. biobreeding diabetes-prone rats (BBDP; 5 non-pregnant rats, 7 pregnant day 10 rats and 6 pregnant day 18 rats) , versus non-diabetic control rats (i.e. congenic non-diabetic biobreeding diabetes-resistant (BBDR; 6 non-pregnant rats, 6 pregnant day 10 rats and 6 pregnant day 18 rats) and Wistar-rats (6 non-pregnant, 6 pregnant day 10 rats and 5 pregnant day 18 rats))., Results: We observed reduced litter size, lower fetal weight of viable fetuses and increased numbers of resorptions versus control rats. These complications are accompanied by various differences in the immune-response between BBDP and control rats in both pregnant and non-pregnant animals. The immune-response in non-pregnant BBDP-rats was characterized by decreased percentages of lymphocytes, increased percentages of effector T-cells, regulatory T-cells and natural killer cells, an increased Th1/Th2-ratio and activated monocytes versus Wistar and BBDR-rats. Furthermore, pregnancy-induced adaptations in BBDP-rats coincided with an increased Th1/Th2-ratio, a decreased mean fluorescence intensity CD161a/NKR-P1b ratio and no further activation of monocytes versus non-diabetic control rats., Conclusion: This study suggests that even in the face of strict normoglycemia, pregnancy complications still occur in type 1 diabetic pregnancies. This adverse pregnancy outcome may be related to the aberrant immunological adaptations to pregnancy in diabetic rats.

Introduction: The role of nutrition in modulating postexercise overnight recovery remains to be elucidated. We assessed the effect of protein ingestion immediately before sleep on digestion and absorption kinetics and protein metabolism during overnight recovery from a single bout of resistance-type exercise., Methods: Sixteen healthy young males performed a single bout of resistance-type exercise in the evening (2000 h) after a full day of dietary standardization. All subjects were provided with appropriate recovery nutrition (20 g of protein, 60 g of CHO) immediately after exercise (2100 h). Thereafter, 30 min before sleep (2330 h), subjects ingested a beverage with (PRO) or without (PLA) 40 g of specifically produced intrinsically [1-C]phenylalanine-labeled casein protein. Continuous intravenous infusions with [ring-H5]phenylalanine and [ring-H2]tyrosine were applied with blood and muscle samples collected to assess protein digestion and absorption kinetics, whole-body protein balance and mixed muscle protein synthesis rates throughout the night (7.5 h)., Results: During sleep, casein protein was effectively digested and absorbed resulting in a rapid rise in circulating amino acid levels, which were sustained throughout the remainder of the night. Protein ingestion before sleep increased whole-body protein synthesis rates (311 ± 8 vs 246 ± 9 μmol·kg per 7.5 h) and improved net protein balance (61 ± 5 vs -11 ± 6 μmol·kg per 7.5 h) in the PRO vs the PLA experiment (P < 0.01). Mixed muscle protein synthesis rates were ∼22% higher in the PRO vs the PLA experiment, which reached borderline significance (0.059%·h ± 0.005%·h vs 0.048%·h ± 0.004%·h, P = 0.05)., Conclusions: This is the first study to show that protein ingested immediately before sleep is effectively digested and absorbed, thereby stimulating muscle protein synthesis and improving whole-body protein balance during postexercise overnight recovery.

Sideways falls onto the hip are a major cause of femoral fractures in the elderly. Martial arts (MA) fall techniques decrease hip impact forces in sideways falls. The femoral fracture risk, however, also depends on the femoral loading configuration (direction and point of application of the force). The purpose of this study was to determine the effect of fall techniques, landing surface and fall height on the impact force and the loading configuration in sideways falls. Twelve experienced judokas performed sideways MA and Block ('natural') falls on a force plate, both with and without a judo mat on top. Kinematic and force data were analysed to determine the hip impact force and the loading configuration. In falls from a kneeling position, the MA technique reduced the impact force by 27%, but did not change the loading configuration. The use of the mat did not change the loading configuration. Falling from a standing changed the force direction. In all conditions, the point of application was distal and posterior to the greater trochanter, but it was less distal and more posterior in falls from standing than from kneeling position. The present decrease in hip impact force with an unchanged loading configuration indicates the potential protective effect of the MA technique on the femoral fracture risk. The change in loading configuration with an increased fall height warrant further studies to examine the effect of MA techniques on fall severity under more natural fall circumstances., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Whey protein ingestion has been shown to effectively stimulate postprandial muscle protein accretion in older adults. However, the impact of the amount of whey protein ingested on protein digestion and absorption kinetics, whole body protein balance, and postprandial muscle protein accretion remains to be established. We aimed to fill this gap by including 33 healthy, older men (73 ± 2 yr) who were randomly assigned to ingest 10, 20, or 35 g of intrinsically l-[1-¹³C]phenylalanine-labeled whey protein (n = 11/treatment). Ingestion of labeled whey protein was combined with continuous intravenous l-[ring-²H₅]phenylalanine and l-[ring-²H₂]tyrosine infusion to assess the metabolic fate of whey protein-derived amino acids. Dietary protein digestion and absorption rapidly increased following ingestion of 10, 20, and 35 g whey protein, with the lowest and highest (peak) values observed following 10 and 35 g, respectively (P < 0.05). Whole body net protein balance was positive in all groups (19 ± 1, 37 ± 2, and 58 ± 2 μmol/kg), with the lowest and highest values observed following ingestion of 10 and 35 g, respectively (P < 0.05). Postprandial muscle protein accretion, assessed by l-[1-¹³C]phenylalanine incorporation in muscle protein, was higher following ingestion of 35 g when compared with 10 (P < 0.01) or 20 (P < 0.05) g. We conclude that ingestion of 35 g whey protein results in greater amino acid absorption and subsequent stimulation of de novo muscle protein synthesis compared with the ingestion of 10 or 20 g whey protein in healthy, older men.

Gait data need to be reliable to be valuable for clinical decision-making. To reduce the impact of marker placement errors, the Optimized Lower Limb Gait Analysis (OLGA) model was developed. The purpose of this study was to assess the sensitivity of the kinematic gait data to a standard marker displacement of the OLGA model compared with the standard Vicon Clinical Manager (VCM) model and to determine whether OLGA reduces the errors due to the most critical marker displacements. Healthy adults performed six gait sessions. The first session was a standard gait session. For the following sessions, 10mm marker displacements were applied. Kinematic data were collected for both models. The root mean squares of the differences (RMS) were calculated for the kinematics of the displacement sessions with respect to the first session. The results showed that the RMS values were generally larger than the stride-to-stride variation except for the pelvic kinematics. For the ankle, knee and hip kinematics, OLGA significantly reduced the averaged RMS values for most planes. The shank, knee and thigh anterior-posterior marker displacements resulted in RMS values exceeding 10°. OLGA reduced the errors due to the knee and thigh marker displacements, but not the errors due to the ankle marker displacements. In conclusion, OLGA reduces the effect of erroneous marker placement, but does not fully compensate all effects, indicating that accurate marker placement remains of crucial importance for adequate 3D-gait analysis and subsequent clinical decision-making., (Copyright © 2011 Elsevier B.V. All rights reserved.)

We hypothesized that imagery training would improve the fast onset of neuromuscular activation and thereby fast knee extensor isometric torque development. Forty young healthy participants, not involved in strength training, were assigned to one of four groups: physical training, imagery training, placebo training or control. The three training groups had three 15 min sessions per week for 4 weeks, with a 90 ° knee angle but were tested also at 120 °. At 90 ° knee angle, maximal torque increased (-8%) similarly in all three training groups. The torque-time integral (contractile impulse) over the first 40 ms after torque onset (TTI40) increased (P < 0.05) after physical training (by -100%), but only at 90 °. This increase was significantly different from the delta values (change pre to post) in the control and placebo groups, whereas delta values in the imagery group were similar to those in the placebo group. The increases in TTI40 following physical training were related (r (2) = 0.81, P < 0.05) to significant increases of knee extensor rectified surface EMG at torque onset (EMG40). In conclusion, only physical training led to a knee angle specific increase of contractile impulse that was significantly different from placebo and controls and that was related to improved onset of neuromuscular activation.

Introduction: Both nonclassical and intermediate monocytes have been implicated in different inflammatory conditions. We hypothesized that these monocytes would increase during pregnancy, a condition associated with generalized activation of inflammatory responses and that they would increase even more during preeclampsia, in which inflammatory responses are further stimulated. In the present study we investigated changes in monocyte subsets during healthy pregnancy and preeclampsia in humans and rats., Methods: Blood monocyte subsets of nonpregnant, preeclamptic and healthy pregnant women were identified with CD14 and CD16. In nonpregnant and pregnant rats, blood monocytes were identified with CD172a and CD43, as well as in rats infused with adenosine triphosphate (ATP), a pro-inflammatory stimulus known to induce preeclampsia-like symptoms. Total and CD206-positive macrophages were quantified in placentas of these animals., Results: Lower percentages of classical monocytes were found in pregnant women (91%-[83-98%]) compared to nonpregnant women (94%-[90-98%]) and even less in preeclamptic patients (90%-[61-92%]). In contrast, the percentage of combined nonclassical/intermediate monocytes was higher in pregnant women (8.5%-[2.3-16.6%] vs. 5.6%-[1.9-9.5%]) and even higher in preeclamptic patients (9.9%-[7.8-38.7%]), which was caused by a selective increase of intermediate monocytes. In rats, we also found lower percentages of classical monocytes and higher percentages of nonclassical monocytes in pregnant versus nonpregnant rats. ATP infusion increased the percentage of nonclassical monocytes in pregnant rats even further but not in nonpregnant rats. These nonclassical monocytes showed a more activated phenotype in pregnant ATP-infused rats only. Mesometrial triangles of ATP-infused rats had less CD206-positive macrophages as compared to those of saline-infused rats., Conclusion: The higher percentage of nonclassical/intermediate monocytes found in pregnancy and preeclampsia confirms their association with inflammatory responses. The observation that ATP stimulated numbers/activation of nonclassical monocytes in pregnant rats only, suggests that nonclassical monocytes are specifically altered in pregnancy and may play a role in the pathophysiology of preeclampsia.

We examined the effect of an acute bout of resistance exercise on fractional muscle protein synthesis rates in human type I and type II muscle fibres. After a standardised breakfast (31 ± 1 kJ kg(-1) body weight, consisting of 52 Energy% (En%) carbohydrate, 34 En% protein and 14 En% fat), 9 untrained men completed a lower-limb resistance exercise bout (8 sets of 10 repetitions leg press and leg extension at 70% 1RM). A primed, continuous infusion of L: -[ring-(13)C(6)]phenylalanine was combined with muscle biopsies collected from both legs immediately after exercise and after 6 h of post-exercise recovery. Single muscle fibres were dissected from freeze-dried biopsies and stained for ATPase activity with pre-incubation at a pH of 4.3. Type I and II fibres were separated under a light microscope and analysed for protein-bound L: -[ring-(13)C(6)]phenylalanine labelling. Baseline (post-exercise) L: -[ring-(13)C(6)]phenylalanine muscle tissue labelling, expressed as (∂(13)C/(12)C), averaged -32.09 ± 0.28, -32.53 ± 0.10 and -32.02 ± 0.16 in the type I and II muscle fibres and mixed muscle, respectively (P = 0.14). During post-exercise recovery, muscle protein synthesis rates were marginally (8 ± 2%) higher in the type I than type II muscle fibres, at 0.100 ± 0.005 versus 0.094 ± 0.005%/h, respectively (P < 0.05), whereby rates of mixed muscle protein were 0.091 ± 0.005%/h. Muscle protein synthesis rates following resistance-type exercise are only marginally higher in type I compared with type II muscle fibres.

Although the practice of fall techniques has been introduced in fall prevention programs, it is not clear whether people can apply acquired techniques during a real-life fall. It would be helpful to know the time it takes to initiate and to successfully execute such techniques, as well as the effect of experience on the execution of these techniques. In this study we investigated the neuromuscular control of voluntary fall techniques in five seasoned judokas and nine non-judokas. After they had started falling from a kneeling position, they received an auditory cue prompting either a lateral natural fall arrest (block) or a martial arts (MA) fall. EMG data of shoulder and trunk muscles were collected. The requested technique was successfully applied in 85% of the falls. Following the cue, EMG amplitudes of the fall techniques started to diverge after 180-190 ms. EMG amplitudes were generally similar in both groups, but experience-related differences could be demonstrated in the pectoralis and trapezius. In conclusion, voluntary motor control is possible within the duration of a fall, even in inexperienced fallers. Differences in EMG activity might suggest that experienced fallers changed their reaction to possible falls from a preparation for arm abduction into a preparation for trunk rotation.

Hip fractures are among the most serious consequences of falls in the elderly. Martial arts (MA) fall techniques may reduce hip fracture risk, as they are known to reduce hip impact forces by approximately 30% in experienced fallers. The purpose of this study was to investigate whether hip impact forces and velocities in MA falls would be smaller than in a 'natural' fall arrest strategy (Block) in young adults (without any prior experience) after a 30-min training session in sideways MA fall techniques. Ten subjects fell sideways from kneeling height. In order to identify experience-related differences, additional EMG data of both fall types were collected in inexperienced (n=10) and experienced fallers (n=5). Compared to Block falls, MA falls had significantly smaller hip impact forces (-17%) and velocities (-7%). EMG results revealed experience-related differences in the execution of the MA fall, indicative of less pronounced trunk rotation in the inexperienced fallers. This may explain their smaller reduction of impact forces compared to experienced fallers. In conclusion, the finding that a substantial reduction in impact forces can be achieved after a short training in MA techniques is very promising with respect to their use in interventions to prevent fall injuries.

Fall techniques that reduce fall severity may decrease the risk of hip fractures. A fundamental variable for fall severity is impact force, but impact velocity is also used. The purpose of the study was to determine whether impact velocity is valid to determine differences in fall severity between different techniques. Five young adults with martial arts (MA) experience performed sideways falls from kneeling height using three techniques: Block with arm (Block) and MA techniques with and without use of the arm to break the fall. In addition, one subject also performed MA falls from standing height. Linear regression analysis showed a moderate relation between hip impact velocity and force, which was depended on technique. In falls with comparable impact velocities, forces in MA falls were lower than forces in Block falls. Hence, differences in impact force could not be predicted by velocity. In conclusion, hip impact velocity may be useful to make an approximate prediction of impact force within fall techniques. However, to determine differences between techniques it was not always a valid predictor. When direct impact force measurements are not possible, methods combining impact velocity with energy estimates before and after impact might be more valid.

Falls to the side and those with impact on the hip are risky for hip fractures in the elderly. A previous study has indicated that martial arts (MA) fall techniques can reduce hip impact force, but the underlying mechanism is unknown. Furthermore, the high impact forces at the hand used to break the fall have raised concerns because of the risk for wrist fractures. The purpose of the study was to get insight into the role of hand impact, impact velocity, and trunk orientation in the reduction of hip impact force in MA techniques. Six experienced judokas performed sideways falls from kneeling height using three fall techniques: block with arm technique (control), MA technique with use of the arm to break the fall (MA-a), and MA technique without use of the arm (MA-na). The results showed that the MA-a and MA-na technique reduced the impact force by 27.5% and 30%, respectively. Impact velocity was significantly reduced in the MA falls. Trunk orientation was significantly less vertical in the MA-a falls. No significant differences were found between the MA techniques. It was concluded that the reduction in hip impact force was associated with a lower impact velocity and less vertical trunk orientation. Rolling after impact, which is characteristic for MA falls, is likely to contribute to the reduction of impact forces, as well. Using the arm to break the fall was not essential for the MA technique to reduce hip impact force. These findings provided support for the incorporation of MA fall techniques in fall prevention programs for elderly.

The objective of the present study was to investigate whether reduction of central acetylcholine (ACh) accumulation by adenosine receptor agonists could serve as a generic treatment against organophosphate (OP) poisoning. The OPs studied were tabun ( O-ethyl- N-dimethylphosphoramidocyanidate), sarin (isopropylmethylphosphonofluoridate), VX ( O-ethyl- S-2-diisopropylaminoethylmethylphosphonothiolate) and parathion ( O, O-diethyl- O-(4-nitrophenyl)phosphorothioate). The efficacy of the adenosine A(1) receptor agonist N(6)-cyclopentyladenosine (CPA) against an OP intoxication was examined on the basis of the occurrence of clinical symptoms that are directly associated with such intoxication. CPA (1-2 mg/kg) effectively attenuated the cholinergic symptoms and prevented mortality in lethally tabun- or sarin-intoxicated rats. In contrast, CPA (2 mg/kg) proved to be ineffective against VX or parathion intoxication. Intracerebral microdialysis studies revealed that survival of sarin-poisoned and CPA-treated animals coincided with a minor elevation of extracellular ACh concentrations in the brain relative to the baseline value, whereas an 11-fold increase in transmitter levels was observed in animals not treated with CPA. In VX-intoxicated rats, however, the ACh amounts increased 18-fold, irrespective of treatment with CPA. The striatal acetylcholinesterase (AChE) activity following a lethal sarin intoxication was completely abolished in the vehicle-treated animals, whereas 10% and 60% AChE activity remained in animals treated with 2 mg/kg CPA 1 min after or 2 min prior to the poisoning, respectively. In VX-intoxicated animals the AChE activity in the brain was strongly reduced (striatum 10%, hippocampus 1%) regardless of the CPA treatment. These results demonstrate that CPA is highly effective against tabun or sarin poisoning, but fails to protect against VX or parathion. Survival and attenuation of clinical signs in tabun- or sarin-poisoned animals are associated with a reduction of ACh accumulation and with protection of AChE activity in the brain.

Current treatment of acute organophosphate (OP) poisoning includes a combined administration of a cholinesterase reactivator (oxime), a muscarinic receptor antagonist (atropine) and an anticonvulsant (diazepam). This treatment is not adequate since it does not prevent neuronal brain damage and incapacitation. Here, as in a recent review it is stated that other therapeutic approaches may improve protection. Former studies on the "direct effects" of oximes led to the conclusion that drug-induced inhibition of acetylcholine (ACh)-release shortly (1 min) after the acute OP-intoxication, could prevent and counteract convulsions and improve survival. In general, the accumulation of ACh in the synaptic cleft is considered to be responsible for the symptoms that ultimately lead to death. Therefore, prevention or suppression of this excessive accumulation of ACh could be a generic approach to antagonize OP-poisoning. Preliminary evidence for this concept has been put forward. Evaluation of drugs that would be able to prevent and counteract ACh accumulation, led to the conclusion that adenosine receptor agonists could be promising candidates. Pilot experiments demonstrated that intramuscular administration of the adenosine receptor agonists NECA (5'-N-ethylcarboxamido-adenosine) or CPA (N6-cyclopentyl adenosine) 1 min following a subcutaneous soman poisoning (1.5-2LD50) in rats, resulted in (1) prevention or postponement of chewing, salivation, convulsive activity, and respiratory distress (cholinergic symptoms), (2) improvement of survival rate (24 h), (3) a low level of extracellular brain ACh, as opposed to high levels of extracellular brain ACh in untreated animals. It is concluded that (1) adenosine agonists protect acutely soman-poisoned rats without the need of additional treatment with atropine, oxime or diazepam, (2) prevention of ACh accumulation in this way may be a new generic approach in the treatment of OP-poisoning.

Hexose oxidase from the red seaweed, Chondrus crispus was purified to homogeneity. The enzyme appeared to be encapsulated in particles obtained after mechanical disintegration of the fronds. Liberation of the enzyme in soluble form required either waiting for the spontaneous development of a suitable microbial flora in the suspension, or treatment with a mixture of proteases (pronase). As deduced from (SDS/)PAGE, the enzyme has a molecular mass of 87 kDa and probably consists of subunits of 36 kDa and 25 kDa. The low isoelectric point of 2.8 and the presence of 25% (by mass) sugars indicate that the enzyme is a strongly acidic glycoprotein. The absorption spectrum of isolated enzyme minus that of the substrate-reduced enzyme, and the EPR spectrum of the free radical observed in the reduced enzyme revealed the presence of a flavin. This cofactor is probably covalently bound since flavins were not released upon denaturation of the enzyme by heat or acid treatment. Taking free FAD as a reference compound, the enzyme contains 1 mol flavin/mol enzyme. EPR spectroscopy of the purified preparation showed the presence of Cu2+. However, since the amount was substoichiometric, substrate addition did not affect the signal, and the addition of chelator or Cu2+ did not affect the activity, the presence of this metal ion seems adventitious. It is concluded that the large discrepancies between the presently and the previously reported [Sullivan, J. D. & Ikawa, M. (1973) Biochim. Biophys. Acta 309, 11-22] characteristics of the enzyme probably originate from the characterization of a contaminating protein in the latter case.

Pig kidney diamine oxidase was purified to homogeneity. The reaction product of the cofactor with p-nitrophenylhydrazine (pNPH) was liberated with pronase treatment and purified. 1H NMR, uv/vis, and electrospray tandem mass spectroscopy revealed it to be a dipeptide with the sequence topaquinone-pNPH and aspartate. No heterogeneity was observed, indicating that no intramolecular cyclization of the quinone moiety occurs in the time span of the isolation and of the measurements. Similar results were obtained with the more widely applicable reagent, phenylhydrazine, and using the aromatic amine oxidase from Escherichia coli. From the amount and ease with which the dipeptide could be isolated, the procedure used here is more convenient than the existing one for the identification of protein-integrated quinone cofactors.

To investigate at which dose levels undesirable effects started, behavioural performance and several physiological parameters were measured in marmosets (Callithrix jacchus) after soman (1.75 and 3.5 micrograms/kg), sarin (3 and 6 micrograms/kg), physostigmine (10 and 20 micrograms/kg), and pyridostigmine (200 and 400 micrograms/kg). Effects on performance were investigated with a discrete-trial, two-choice visual discrimination task and a hand-eye coordination task. The former test appeared more sensitive to disruption than the hand-eye coordination task. "Motor speed" was not disrupted by any of the four compounds. However, "choice time" as well as "no attempts" increased and were clearly more disturbed by soman and physostigmine than by sarin and pyridostigmine. All effects had disappeared after 24 h. Except for a small effect of sarin on heart rate and blood pressure, none of the cholinesterase (ChE) inhibitors affected a number of physiological parameters at behavioural effective does that caused a profound ChE inhibition in blood. Take together, these results strongly suggest that both soman and physostigmine may interfere with higher CNS functions at low dose levels. These effects may go undetected because physical signs are absent.

Four lines of mutant mice were used to investigate (1) the neuronal localization of cannabinoid receptors in the cerebellar molecular layer and (2) the anatomical association of these receptors with elements of the two second messenger systems in the brain. Two of the mutant lines--Purkinje cell degeneration and nervous--are selectively deficient in Purkinje cells; the other two--weaver and reeler--are deficient in granule cells. In the heterozygous mice, [3H]CP 55,940 binding to cannabinoid receptors was discretely and densely localized to the molecular layer, as was [3H]forskolin binding to adenylate cyclase and [3H]phorbol 12,13-dibutyrate binding to protein kinase C, a component of the phosphoinositide cycle. [3H]CP 55,940 and [3H]forskolin binding was selectively reduced in weaver and reeler homozygous mice but unchanged in Purkinje cell deficient and nervous homozygotes. No decreases in [3H]phorbol 12,13-dibutyrate binding were found in any of the homozygous mutants relative to the heterozygous littermates. The results suggest that cannabinoid receptors and adenylate cyclase are localized to granule cell axons in the molecular layer, whereas protein kinase C is equally distributed in parallel fibers and Purkinje cell dendrites.

We have measured the thermodynamic efficiency of oxidative phosphorylation of isolated rat-liver mitochondria during oxidation of succinate. Furthermore, we have calculated what the effect of proton leak or slip in the redox pumps should be on the efficiency of energy transduction in oxidative phosphorylation. These calculations were compared with experiments in which the efficiency was determined in the presence of induced proton leak or redox slip. The results of these experiments are in agreement with the predictions. It is concluded that it is possible to distinguish experimentally between effects of proton leak and redox slip on energy transduction.

Based on the results of the so-called redox-cycling assay it has been claimed that various common foods and beverages as well as mammalian body fluids and tissues contain substantial quantities (microM) of free PQQ [M. Paz et al. (1989) in: PQQ and Quinoproteins (J.A. Jongejan and J.A. Duine, eds.) Kluwer Academic Publishers, Dordrecht, pp. 131-143 and J. Killgore et al. (1989) Science 245, 850-852]. However, by investigating samples from such sources with a biological assay of nM sensitivity, we could not confirm these claims. Analysis of the samples with procedures that proved adequate for the detection of PQQ adducts and conjugates gave equally negative results. To account for the positive response in the redox-cycling assay, as opposed to the negative results obtained by other methods, a search was made for those substances in these samples that caused the false-positive reactions. It was found that a number of commonly occurring biochemicals like ascorbic and dehydroascorbic acid, riboflavin and to a lesser extent pyridoxal phosphate, gave a positive response in the redox-cycling assay. The amounts of these interfering substances that were determined in the samples by independent methods could well explain the response. In separate experiments it was found that the effect of PQQ added to biological samples was obscured over an appreciable range of concentrations. For these reasons it must be concluded that the redox-cycling assay is not suited for the detection of PQQ in these samples. Any claims that are based on the results of this method should be disregarded.

Analysis of glutamic acid decarboxylase (GDC) (EC 4.1.1.15) from Escherichia coli ATCC 11246 revealed the presence of six pyridoxal phosphates (PLPs) as well as six covalently bound pyrroloquinoline quinones (PQQs) per hexameric enzyme molecule. This is the second example of a pyridoxo-quinoprotein, suggesting that other atypical pyridoxoproteins (PLP-containing enzymes) have similar cofactor composition. Since the organism did not produce free PQQ and its quinoprotein glucose dehydrogenase was present in the apo form, free PQQ is not used in the assemblage of GDC. Most probably, biosynthesis of covalently bound cofactor occurs in situ via a route which is different from that of free PQQ. Thus, organisms previously believed to be unable to synthesize (free) PQQ could in fact be able to produce quinoproteins with covalently bound cofactor. Implications for the role of PQQ in eukaryotic cells are discussed.

Pig kidney 3,4-dihydroxyphenylalanine (dopa) decarboxylase (EC 4.1.1.28) was purified to homogeneity. Treatment of the enzyme with phenylhydrazine (PH) according to a procedure developed for analysis of quinoproteins gave products which were identified as the hydrazone of pyridoxal phosphate (PLP) and the C(5)-hydrazone of pyrroloquinoline quinone (PQQ). This method failed, however, in quantifying the amounts of cofactor. Direct hydrolysis of the enzyme by refluxing with hexanol and concentrated HCl led to detachment of PQQ from the protein in a quantity of 1 PQQ per enzyme molecule. In view of the reactivity of PQQ towards amines and amino acids, we postulate that it participates as a covalently bound cofactor in the catalytic cycle of the enzyme, in interplay with PLP. Since several other enzymes have been reported to show the atypical behaviour of dopa decarboxylase, it seems that the PLP-containing group of enzymes can be subdivided into pyridoxoproteins and pyridoxo-quinoproteins.

A guinea pig model was used to study correlations between cis-platinum dosage and level of hearing loss, hair cell loss in the cochlea. Other parameters measured included weight loss and serum urea, creatinine, and cis-platinum levels. The damage to the inner ear expressed as hearing loss and hair cell loss demonstrated a highly positive correlation with the dosage of cis-platinum, and only a moderately positive correlation with the cis-platinum serum levels. Weight loss correlated well with hearing loss.

Aldehyde dehydrogenase from Pseudomonas testosteroni was purified to homogeneity. The enzyme has a pH optimum of 8.2, uses a wide range of aldehydes as substrates and cationic dyes (Wurster's blue, phenazine methosulphate and thionine), but not anionic dyes (ferricyanide and 2.6-dichloroindophenol), NAD(P)+ or O2, as electron acceptors. Haem c and pyrroloquinoline quinone appeared to be absent but the common cofactors of molybdenum hydroxylases were present. Xanthine was not a substrate and allopurinol was not an inhibitor. Alcohols were inhibitors only when turnover of the enzyme occurred in aldehyde conversion. The enzyme has a relative molecular mass of 186,000, consists of two subunits of equal size (Mr 92,000), and 1 enzyme molecule contains 1 FAD, 1 molybdopterin cofactor, 4 Fe and 4 S. It is a novel type of NAD(P)+-independent aldehyde dehydrogenase since its catalytic and physicochemical properties are quite different from those reported for already known aldehyde-converting enzymes like haemoprotein aldehyde dehydrogenase (EC 1.2.99.3), quino-protein alcohol dehydrogenases (EC 1.1.99.8) and molybdenum hydroxylases.