Reverse-translational biomarker validation of Abnormal Repetitive Behaviors in mice: an illustration of the 4P's modeling approach

The NIMH's new strategic plan, with its emphasis on the "4P's" (Prediction, Pre-emption, Personalization, and Populations) and biomarker-based medicine requires a radical shift in animal modeling methodology. In particular 4P's models will be non-determinant (i.e. disease severity will depend on secondary environmental and genetic factors); and validated by reverse-translation of animal homologues to human biomarkers. A powerful consequence of the biomarker approach is that different closely related disorders have a unique fingerprint of biomarkers. Animals can be validated as a highly specific model of a single disorder by matching this 'fingerprint'; or as a model of a symptom seen in multiple disorders by matching common biomarkers. Here we illustrate this approach with two Abnormal Repetitive Behaviors (ARBs) in mice: stereotypies and barbering (hair pulling). We developed animal versions of the neuropsychological biomarkers that distinguish human ARBs, and tested the fingerprint of the different mouse ARBs. As predicted, the two mouse ARBs were associated with different biomarkers. Both barbering and stereotypy could be discounted as models of OCD (even though they are widely used as such), due to the absence of limbic biomarkers which are characteristic of OCD and hence are necessary for a valid model. Conversely barbering matched the fingerprint of trichotillomania (i.e. selective deficits in set-shifting), suggesting it may be a highly specific model of this disorder. In contrast stereotypies were correlated only with a biomarker (deficits in response shifting) correlated with stereotypies in multiple disorders, suggesting that animal stereotypies model stereotypies in multiple disorders.

Comparison of objectively measured motor behavior with ratings of the motor behavior domain of the Bern Psychopathology Scale (BPS) in schizophrenia

Motor symptoms in schizophrenia occur frequently and are relevant to diagnosis and antipsychotic therapy. To date motor symptoms are difficult to assess and their pathobiology is a widely unresolved issue. The Bern Psychopathology Scale for the assessment of system-specific psychotic symptoms (BPS) was designed to identify homogenous patient groups by focusing on three domains: language, affectivity and motor behavior. The present study aimed to validate the motor behavior domain of the BPS using wrist actigraphy. In total, 106 patients were rated with the BPS and underwent 24 h continuous actigraphy recording. The ratings of the global severity of the motor behavior domain (GSM) as well as the quantitative and the subjective items of the motor behavior domain of the BPS were significantly associated with actigraphic variables. In contrast, the qualitative items of the motor domain failed to show an association with actigraphy. Likewise, scores of the language and the affectivity domains were not related to actigraphic measures. In conclusion, we provided substantial external validity for global, quantitative and subjective ratings of the BPS motor behavior domain. Thus, the BPS is suitable to assess the dimension of quantitative motor behavior in the schizophrenia spectrum.

Cyclic nucleotide specific phosphodiesterases of Leishmania major

Background Leishmania represent a complex of important human pathogens that belong to the systematic order of the kinetoplastida. They are transmitted between their human and mammalian hosts by different bloodsucking sandfly vectors. In their hosts, the Leishmania undergo several differentiation steps, and their coordination and optimization crucially depend on numerous interactions between the parasites and the physiological environment presented by the fly and human hosts. Little is still known about the signalling networks involved in these functions. In an attempt to better understand the role of cyclic nucleotide signalling in Leishmania differentiation and host-parasite interaction, we here present an initial study on the cyclic nucleotide-specific phosphodiesterases of Leishmania major. Results This paper presents the identification of three class I cyclic-nucleotide-specific phosphodiesterases (PDEs) from L. major, PDEs whose catalytic domains exhibit considerable sequence conservation with, among other, all eleven human PDE families. In contrast to other protozoa such as Dictyostelium, or fungi such as Saccharomyces cerevisiae, Candida ssp or Neurospora, no genes for class II PDEs were found in the Leishmania genomes. LmjPDEA contains a class I catalytic domain at the C-terminus of the polypeptide, with no other discernible functional domains elsewhere. LmjPDEB1 and LmjPDEB2 are coded for by closely related, tandemly linked genes on chromosome 15. Both PDEs contain two GAF domains in their N-terminal region, and their almost identical catalytic domains are located at the C-terminus of the polypeptide. LmjPDEA, LmjPDEB1 and LmjPDEB2 were further characterized by functional complementation in a PDE-deficient S. cerevisiae strain. All three enzymes conferred complementation, demonstrating that all three can hydrolyze cAMP. Recombinant LmjPDEB1 and LmjPDEB2 were shown to be cAMP-specific, with Km values in the low micromolar range. Several PDE inhibitors were found to be active against these PDEs in vitro, and to inhibit cell proliferation. Conclusion The genome of L. major contains only PDE genes that are predicted to code for class I PDEs, and none for class II PDEs. This is more similar to what is found in higher eukaryotes than it is to the situation in Dictyostelium or the fungi that concomitantly express class I and class II PDEs. Functional complementation demonstrated that LmjPDEA, LmjPDEB1 and LmjPDEB2 are capable of hydrolyzing cAMP. In vitro studies with recombinant LmjPDEB1 and LmjPDEB2 confirmed this, and they demonstrated that both are completely cAMP-specific. Both enzymes are inhibited by several commercially available PDE inhibitors. The observation that these inhibitors also interfere with cell growth in culture indicates that inhibition of the PDEs is fatal for the cell, suggesting an important role of cAMP signalling for the maintenance of cellular integrity and proliferation.

Native EEG and treatment effects in neuroleptic-naïve schizophrenic patients: time and frequency domain approaches

Time domain analysis of electroencephalography (EEG) can identify subsecond periods of quasi-stable brain states. These so-called microstates assumingly correspond to basic units of cognition and emotion. On the other hand, Global Field Synchronization (GFS) is a frequency domain measure to estimate functional synchronization of brain processes on a global level for each EEG frequency band [Koenig, T., Lehmann, D., Saito, N., Kuginuki, T., Kinoshita, T., Koukkou, M., 2001. Decreased functional connectivity of EEG theta-frequency activity in first-episode, neuroleptic-naive patients with schizophrenia: preliminary results. Schizophr Res. 50, 55-60.]. Using these time and frequency domain analyzes, several previous studies reported shortened microstate duration in specific microstate classes and decreased GFS in theta band in drug naïve schizophrenia compared to controls. The purpose of this study was to investigate changes of these EEG parameters after drug treatment in drug naïve schizophrenia. EEG analysis was performed in 21 drug-naive patients and 21 healthy controls. 14 patients were reevaluated 2-8 weeks (mean 4.3) after the initiation of drug administration. The results extended findings of treatment effect on brain functions in schizophrenia, and imply that shortened duration of specific microstate classes seems a state marker especially in patients with later neuroleptic responsive, while lower theta GFS seems a state-related phenomenon and that higher gamma GFS is a trait like phenomenon.

Modulation of human CYP19A1 activity by mutant NADPH P450 oxidoreductase

Mutations in NADPH P450 oxidoreductase (POR) cause a broad spectrum of human disease with abnormalities in steroidogenesis. We have studied the impact of P450 reductase mutations on the activity of CYP19A1. POR supported CYP19A1 activity with a calculated Km of 126 nm for androstenedione and a Vmax of 1.7 pmol/min. Mutations R457H and V492E located in the FAD domain of POR that disrupt electron transfer caused a complete loss of CYP19A1 activity. The A287P mutation of POR decreased the activities of CYP17A1 by 60-80% but had normal CYP19A1 activity. Molecular modeling and protein docking studies suggested that A287P is involved in the interaction of POR:CYP17A1 but not in the POR:CYP19A1 interaction. Mutations C569Y and V608F in the NADPH binding domain of POR had 49 and 28% of activity of CYP19A1 compared with normal reductase and were more sensitive to the amount of NADPH available for supporting CYP19A1 activity. Substitution of NADH for NADPH had a higher impact on C569Y and V608F mutants of POR. Similar effects were obtained at low/high (5.5/8.5) pH, but using octanol to limit the flux of electrons from POR to CYP19A1 inhibited activity supported by all variants. High molar ratios of KCl also reduced the CYP19A1 supporting activities of C569Y and V608F mutants of POR to a greater extent compared to normal POR and A287P mutant. Because POR supports many P450s involved in steroidogenesis, bone formation, and drug metabolism, variations in the effects of POR mutations on specific enzyme activities may explain the broad clinical spectrum of POR deficiency.

Modeling of electroosmotic and electrophoretic mobilization in capillary and microchip isoelectric focusing

Our dynamic capillary electrophoresis model which uses material specific input data for estimation of electroosmosis was applied to investigate fundamental aspects of isoelectric focusing (IEF) in capillaries or microchannels made from bare fused-silica (FS), FS coated with a sulfonated polymer, polymethylmethacrylate (PMMA) and poly(dimethylsiloxane) (PDMS). Input data were generated via determination of the electroosmotic flow (EOF) using buffers with varying pH and ionic strength. Two models are distinguished, one that neglects changes of ionic strength and one that includes the dependence between electroosmotic mobility and ionic strength. For each configuration, the models provide insight into the magnitude and dynamics of electroosmosis. The contribution of each electrophoretic zone to the net EOF is thereby visualized and the amount of EOF required for the detection of the zone structures at a particular location along the capillary, including at its end for MS detection, is predicted. For bare FS, PDMS and PMMA, simulations reveal that EOF is decreasing with time and that the entire IEF process is characterized by the asymptotic formation of a stationary steady-state zone configuration in which electrophoretic transport and electroosmotic zone displacement are opposite and of equal magnitude. The location of immobilization of the boundary between anolyte and most acidic carrier ampholyte is dependent on EOF, i.e. capillary material and anolyte. Overall time intervals for reaching this state in microchannels produced by PDMS and PMMA are predicted to be similar and about twice as long compared to uncoated FS. Additional mobilization for the detection of the entire pH gradient at the capillary end is required. Using concomitant electrophoretic mobilization with an acid as coanion in the catholyte is shown to provide sufficient additional cathodic transport for that purpose. FS capillaries dynamically double coated with polybrene and poly(vinylsulfonate) are predicted to provide sufficient electroosmotic pumping for detection of the entire IEF gradient at the cathodic column end.

Hypnotic and opioid anesthetic drug interactions on the CNS, focus on response surface modeling

This chapter will present the conceptual and applied approaches to capture the interaction of anesthetic hypnotic drugs with opioid drugs, as used in the clinical anesthetic state. The graphic and mathematical approaches used to capture hypnotic/opiate anesthetic drug interactions will be presented. This chapter is not a review article about interaction modeling, but focuses on specific drug interactions within a quite narrow field, anesthesia.

A novel FIP1L1-PDGFRA mutant destabilizing the inactive conformation of the kinase domain in chronic eosinophilic leukemia/hypereosinophilic syndrome

BACKGROUND: The Fip1-like-1-platelet-derived growth factor receptor alpha (FIP1L1-PDGFRA) gene fusion is a common cause of chronic eosinophilic leukemia (CEL)/hypereosinophilic syndrome (HES), and patients suffering from this particular subgroup of CEL/HES respond to low-dose imatinib therapy. However, some patients may develop imatinib resistance because of an acquired T674I mutation, which is believed to prevent drug binding through steric hindrance. METHODS: In an imatinib resistant FIP1L1-PDGFRA positive patient, we analyzed the molecular structure of the fusion gene and analyzed the effect of several kinase inhibitors on FIP1L1-PDGFRA-mediated proliferative responses in vitro. RESULTS: Sequencing of the FIP1L1-PDGFRA fusion gene revealed the occurrence of a S601P mutation, which is located within the nucleotide binding loop. In agreement with the clinical observations, imatinib did not inhibit the proliferation of S601P mutant FIP1L1-PDGFRA-transduced Ba/F3 cells. Moreover, sorafenib, which has been described to inhibit T674I mutant FIP1L1-PDGFRA, failed to block S601P mutant FIP1L1-PDGFRA. Structural modeling revealed that the newly identified S601P mutated form of PDGFRA destabilizes the inactive conformation of the kinase domain that is necessary to bind imatinib as well as sorafenib. CONCLUSIONS: We identified a novel mutation in FIP1L1-PDGFRA resulting in both imatinib and sorafenib resistance. The identification of novel drug-resistant FIP1L1-PDGFRA variants may help to develop the next generation of target-directed compounds for CEL/HES and other leukemias.

Near-infrared digital photography to estimate snow correlation length for microwave emission modeling

The study is based on experimental work conducted in alpine snow. We made microwave radiometric and near-infrared reflectance measurements of snow slabs under different experimental conditions. We used an empirical relation to link near-infrared reflectance of snow to the specific surface area (SSA), and converted the SSA into the correlation length. From the measurements of snow radiances at 21 and 35 GHz , we derived the microwave scattering coefficient by inverting two coupled radiative transfer models (the sandwich and six-flux model). The correlation lengths found are in the same range as those determined in the literature using cold laboratory work. The technique shows great potential in the determination of the snow correlation length under field conditions.

The SAKK cancer-specific geriatric assessment (C-SGA): a pilot study of a brief tool for clinical decision-making in older cancer patients

BACKGROUND Recommendations from international task forces on geriatric assessment emphasize the need for research including validation of cancer-specific geriatric assessment (C-SGA) tools in oncological settings. The objective of this study was to evaluate the feasibility of the SAKK Cancer-Specific Geriatric Assessment (C-SGA) in clinical practice. METHODS A cross sectional study of cancer patients >=65 years old (N = 51) with pathologically confirmed cancer presenting for initiation of chemotherapy treatment (07/01/2009-03/31/2011) at two oncology departments in Swiss canton hospitals: Kantonsspital Graubunden (KSGR N = 25), Kantonsspital St. Gallen (KSSG N = 26). Data was collected using three instruments, the SAKK C-SGA plus physician and patient evaluation forms. The SAKK C-SGA includes six measures covering five geriatric assessment domains (comorbidity, function, psychosocial, nutrition, cognition) using a mix of medical record abstraction (MRA) and patient interview. Five individual domains and one overall SAKK C-SGA score were calculated and dichotomized as below/above literature-based cut-offs. The SAKK C-SGA was evaluated by: patient and physician estimated time to complete, ease of completing, and difficult or unanswered questions. RESULTS Time to complete the patient questionnaire was considered acceptable by almost all (>=96%) patients and physicians. Patients reported slightly shorter times to complete the questionnaire than physicians (17.33 +/- 7.34 vs. 20.59 +/- 6.53 minutes, p = 0.02). Both groups rated the patient questionnaire as easy/fairly easy to complete (91% vs. 84% respectively, p = 0.14) with few difficult or unanswered questions. The MRA took on average 8.32 +/- 4.72 minutes to complete. Physicians (100%) considered time to complete MRA acceptable, 96% rated it as easy/fairly easy to complete. Individual study site populations differed on health-related characteristics (excellent/good physician-rated general health KSGR 71% vs. KSSG 32%, p = 0.007). The overall mean C-SGA score was 2.4 +/- 1.12. Patients at KSGR had lower C-SGA scores (2.00 +/- 1.19 vs. 2.81 +/- 0.90, p = 0.009) and a smaller proportion (28% vs.65%, p = 0.008) was above the C-SGA cut-off score compared to KSSG. CONCLUSIONS These results suggest the SAKK C-SGA is a feasible practical tool for use in clinical practice. It demonstrated discriminative ability based on objective geriatric assessment measures, but additional investigations on use for clinical decision-making are warranted. The SAKK C-SGA also provides important usable domain information for intervention to optimize outcomes in older cancer patients.

Inhibition of Fas-associated death domain-containing protein (FADD) protects against myocardial ischemia/reperfusion injury in a heart failure mouse model

AIM As technological interventions treating acute myocardial infarction (MI) improve, post-ischemic heart failure increasingly threatens patient health. The aim of the current study was to test whether FADD could be a potential target of gene therapy in the treatment of heart failure. METHODS Cardiomyocyte-specific FADD knockout mice along with non-transgenic littermates (NLC) were subjected to 30 minutes myocardial ischemia followed by 7 days of reperfusion or 6 weeks of permanent myocardial ischemia via the ligation of left main descending coronary artery. Cardiac function were evaluated by echocardiography and left ventricular (LV) catheterization and cardiomyocyte death was measured by Evans blue-TTC staining, TUNEL staining, and caspase-3, -8, and -9 activities. In vitro, H9C2 cells transfected with ether scramble siRNA or FADD siRNA were stressed with chelerythrin for 30 min and cleaved caspase-3 was assessed. RESULTS FADD expression was significantly decreased in FADD knockout mice compared to NLC. Ischemia/reperfusion (I/R) upregulated FADD expression in NLC mice, but not in FADD knockout mice at the early time. FADD deletion significantly attenuated I/R-induced cardiac dysfunction, decreased myocardial necrosis, and inhibited cardiomyocyte apoptosis. Furthermore, in 6 weeks long term permanent ischemia model, FADD deletion significantly reduced the infarct size (from 41.20 ± 3.90% in NLC to 26.83 ± 4.17% in FADD deletion), attenuated myocardial remodeling, improved cardiac function and improved survival. In vitro, FADD knockdown significantly reduced chelerythrin-induced the level of cleaved caspase-3. CONCLUSION Taken together, our results suggest FADD plays a critical role in post-ischemic heart failure. Inhibition of FADD retards heart failure progression. Our data supports the further investigation of FADD as a potential target for genetic manipulation in the treatment of heart failure.

Patient-specific finite-element simulation of the human cornea: A clinical validation study on cataract surgery

The planning of refractive surgical interventions is a challenging task. Numerical modeling has been proposed as a solution to support surgical intervention and predict the visual acuity, but validation on patient specific intervention is missing. The purpose of this study was to validate the numerical predictions of the post-operative corneal topography induced by the incisions required for cataract surgery. The corneal topography of 13 patients was assessed preoperatively and postoperatively (1-day and 30-day follow-up) with a Pentacam tomography device. The preoperatively acquired geometric corneal topography – anterior, posterior and pachymetry data – was used to build patient-specific finite element models. For each patient, the effects of the cataract incisions were simulated numerically and the resulting corneal surfaces were compared to the clinical postoperative measurements at one day and at 30-days follow up. Results showed that the model was able to reproduce experimental measurements with an error on the surgically induced sphere of 0.38D one day postoperatively and 0.19D 30 days postoperatively. The standard deviation of the surgically induced cylinder was 0.54D at the first postoperative day and 0.38D 30 days postoperatively. The prediction errors in surface elevation and curvature were below the topography measurement device accuracy of ±5μm and ±0.25D after the 30-day follow-up. The results showed that finite element simulations of corneal biomechanics are able to predict post cataract surgery within topography measurement device accuracy. We can conclude that the numerical simulation can become a valuable tool to plan corneal incisions in cataract surgery and other ophthalmosurgical procedures in order to optimize patients' refractive outcome and visual function.

Relations between the attentional blink and aspects of psychometric intelligence: A fixed-links modeling approach

The attentional blink phenomenon (AB) represents impaired identification of the second of two targets presented in rapid succession within a stream of stimuli. Despite the well-known association between attentional processes and psychometric intelligence (PI), evidence for a relationship between AB and PI is highly inconsistent. Theory and empirical findings suggest AB to be multifaceted. Hence, relations between AB and PI may be blurred when AB is measured as a single process. Furthermore, different aspects of PI might be differentially related to AB. The present study explored the relationship between processes underlying AB and general PI as well as specific aspects of PI (Reasoning, Speed, Memory, and Creativity) in 201 female students. Fixed-links modeling revealed three processes underlying AB: (1) a U-shaped process positively related to Speed and negatively related to Memory but unrelated to Reasoning, Creativity, and general PI, (2) an increasing process positively related to Reasoning, Speed, Memory, and general PI but not to Creativity, and (3) a decreasing process positively related to general PI and Memory but not to other aspects of PI. Our findings demonstrate that dissociating processes underlying AB and considering specific aspects of PI is required to understand the relationship between AB and PI.

Kinetic modeling of sodium in the lunar exosphere

Our knowledge about the lunar environment is based on a large volume of ground-based, remote, and in situ observations. These observations have been conducted at different times and sampled different pieces of such a complex system as the surface-bound exosphere of the Moon. Numerical modeling is the tool that can link results of these separate observations into a single picture. Being validated against previous measurements, models can be used for predictions and interpretation of future observations results. In this paper we present a kinetic model of the sodium exosphere of the Moon as well as results of its validation against a set of ground-based and remote observations. The unique characteristic of the model is that it takes the orbital motion of the Moon and the Earth into consideration and simulates both the exosphere as well as the sodium tail self-consistently. The extended computational domain covers the part of the Earth’s orbit at new Moon, which allows us to study the effect of Earth’s gravity on the lunar sodium tail. The model is fitted to a set of ground-based and remote observations by tuning sodium source rate as well as values of sticking, and accommodation coefficients. The best agreement of the model results with the observations is reached when all sodium atoms returning from the exosphere stick to the surface and the net sodium escape rate is about 5.3 × 1022 s−1.

U7 snRNP-specific Lsm11 protein: dual binding contacts with the 100 kDa zinc finger processing factor (ZFP100) and a ZFP100-independent function in histone RNA 3' end processing

The 3' cleavage generating non-polyadenylated animal histone mRNAs depends on the base pairing between U7 snRNA and a conserved histone pre-mRNA downstream element. This interaction is enhanced by a 100 kDa zinc finger protein (ZFP100) that forms a bridge between an RNA hairpin element upstream of the processing site and the U7 small nuclear ribonucleoprotein (snRNP). The N-terminus of Lsm11, a U7-specific Sm-like protein, was shown to be crucial for histone RNA processing and to bind ZFP100. By further analysing these two functions of Lsm11, we find that Lsm11 and ZFP100 can undergo two interactions, i.e. between the Lsm11 N-terminus and the zinc finger repeats of ZFP100, and between the N-terminus of ZFP100 and the Sm domain of Lsm11, respectively. Both interactions are not specific for the two proteins in vitro, but the second interaction is sufficient for a specific recognition of the U7 snRNP by ZFP100 in cell extracts. Furthermore, clustered point mutations in three phylogenetically conserved regions of the Lsm11 N-terminus impair or abolish histone RNA processing. As these mutations have no effect on the two interactions with ZFP100, these protein regions must play other roles in histone RNA processing, e.g. by contacting the pre-mRNA or additional processing factors.