Neural network vehicle models for high-performance automated driving.

Automated vehicles navigate through their environment by first planning and subsequently following a safe trajectory. To prove safer than human beings, they must ultimately perform these tasks as well or better than human drivers across a broad range of conditions and in critical situations. We show that a feedforward-feedback control structure incorporating a simple physics-based model can be used to track a path up to the friction limits of the vehicle with performance comparable with a champion amateur race car driver. The key is having the appropriate model. Although physics-based models are useful in their transparency and intuition, they require explicit characterization around a single operating point and fail to make use of the wealth of vehicle data generated by autonomous vehicles. To circumvent these limitations, we propose a neural network structure using a sequence of past states and inputs motivated by the physical model. The neural network achieved better performance than the physical model when implemented in the same feedforward-feedback control architecture on an experimental vehicle. More notably, when trained on a combination of data from dry roads and snow, the model was able to make appropriate predictions for the road surface on which the vehicle was traveling without the need for explicit road friction estimation. These findings suggest that the network structure merits further investigation as the basis for model-based control of automated vehicles over their full operating range.

Mind over motor mapping: Driver response to changing vehicle dynamics.

Improvements in vehicle safety require understanding of the neural systems that support the complex, dynamic task of real-world driving. We used functional near infrared spectroscopy (fNIRS) and pupilometry to quantify cortical and physiological responses during a realistic, simulated driving task in which vehicle dynamics were manipulated. Our results elucidate compensatory changes in driver behavior in response to changes in vehicle handling. We also describe associated neural and physiological responses under different levels of mental workload. The increased cortical activation we observed during the late phase of the experiment may indicate motor learning in prefrontal-parietal networks. Finally, relationships among cortical activation, steering control, and individual personality traits suggest that individual brain states and traits may be useful in predicting a driver's response to changes in vehicle dynamics. Results such as these will be useful for informing the design of automated safety systems that facilitate safe and supportive driver-car communication.

Neural, physiological, and behavioral correlates of visuomotor cognitive load.

Visuomotor ability is quite crucial for everyday functioning, particularly in driving and sports. While there is accumulating evidence regarding neural correlates of visuomotor transformation, less is known about the brain regions that accommodate visuomotor mapping under different cognitive demands. We concurrently measured cortical activity and pupillary response, using functional near infrared spectroscopy (fNIRS) and eye-tracking glasses, to examine the neural systems linked to pupil dilation under varying cognitive demands. Twenty-three healthy adults performed two sessions of a navigation task, in which the cognitive load was manipulated by either reversing the visuomotor mapping or increasing the speed of the moving object. We identified a region in the right superior parietal lobule that responded to both types of visuomotor load and its activity was associated with larger pupillary response and better performance in the task. Our multimodal analyses suggest that activity in this region arises from the need for increased attentional effort and alertness for visuomotor control and is an ideal candidate for objective measurement of visuomotor cognitive load. Our data extend previous findings connecting changes in pupil diameter to neural activity under varying cognitive demand and have important implications for examining brain-behavior associations in real-world tasks such as driving and sports.

Motor learning affects car-to-driver handover in automated vehicles.

Vehicles in the foreseeable future will be required to transition between autonomous driving (without human involvement) and full human control. During this transition period, the human, who has not been actively engaged in the driving process, must resume the motor control necessary to steer the car. The in-car study presented here demonstrates that when human drivers are presented with a steering behavior that is different from the last time they were in control, specifically the ratio of hand wheel angle to road wheel angle (emulating a change in vehicle speed), they undergo a significant period of adaptation before they return to their previous steering behavior. However, drivers do not require an adaptation period to return to previous driving behavior after changes in steering torque. These findings have implications for the design of vehicles that transition from automated to manual driving and for understanding of human motor control in real-world tasks.

Telemedicine for genetic and neurologic evaluation in the neonatal intensive care unit.

OBJECTIVE: Evaluate whether telemedicine can be used to perform dysmorphology and neurologic examinations in the neonatal intensive care unit (NICU) by determining the examination accuracy, limitations and optimized procedures. STUDY DESIGN: Prospective evaluation of NICU patients referred for subspecialty consultation for dysmorphic features (n=10) or encephalopathy (n=10). A physician at bedside (bedside clinician) performed an in-person examination that was viewed in real time by a remote physician (remote consultant). Standardized examinations were recorded and compared. Subsequently, a qualitative approach established technique adjustments and optimization procedures necessary to improve visualization. RESULT: Telemedicine examinations identified 81 of 87 (93%) dysmorphology examination abnormalities and 37 of 39 (92%) neurologic examination abnormalities. Optimization of remote consultant visualization required an active bedside clinician assisting in camera and patient adjustments. CONCLUSION: Telemedicine can be used to perform accurately many components of the dysmorphology or neurologic examinations in NICU patients, but physicians must be mindful of specific limitations.

Generation and evaluation of a monoclonal antibody, designated MAdL, as a new specific marker for adenocarcinomas of the lung.

BACKGROUND: Different therapy regimens in non-small-cell lung cancer (NSCLC) are of rising clinical importance, and therefore a clear-cut subdifferentiation is mandatory. The common immunohistochemical markers available today are well applicable for subdifferentiation, but a fraction of indistinct cases still remains, demanding upgrades of the panel by new markers. METHODS: We report here the generation and evaluation of a new monoclonal antibody carrying the MAdL designation, which was raised against primary isolated human alveolar epithelial cells type 2. RESULTS: Upon screening, one clone (MAdL) was identified as a marker for alveolar epithelial cell type II, alveolar macrophages and adenocarcinomas of the lung. In a large-scale study, this antibody, with an optimised staining procedure for formalin-fixed tissues, was then evaluated together with the established markers thyroid transcription factor-1, surfactant protein-A, pro-surfactant protein-B and napsin A in a series of 362 lung cancer specimens. The MAdL displays a high specificity (>99%) for adenocarcinomas of the lung, together with a sensitivity of 76.5%, and is capable of delivering independent additional diagnostic information to the established markers. CONCLUSION: We conclude that MAdL is a new specific marker for adenocarcinomas of the lung, which helps to clarify subdifferentiation in a considerable portion of NSCLCs.

Chromophore-assisted light inactivation of pKi-67 leads to inhibition of ribosomal RNA synthesis.

OBJECTIVES: Expression of the nuclear Ki-67 protein (pKi-67) is strongly associated with cell proliferation. For this reason, antibodies against this protein are widely used as prognostic tools for the assessment of cell proliferation in biopsies from cancer patients. Despite this broad application in histopathology, functional evidence for the physiological role of pKi-67 is still missing. Recently, we proposed a function of pKi-67 in the early steps of ribosomal RNA (rRNA) synthesis. Here, we have examined the involvement of pKi-67 in this process by photochemical inhibition using chromophore-assisted light inactivation (CALI). MATERIALS AND METHODS: Anti-pKi-67 antibodies were labelled with the fluorochrome fluorescein 5(6)-isothiocyanate and were irradiated after binding to their target protein. RESULTS: Performing CALI in vitro on cell lysates led to specific cross-linking of pKi-67. Moreover, the upstream binding factor (UBF) necessary for rRNA transcription was also partly subjected to cross-link formation, indicating a close spatial proximity of UBF and pKi-67. CALI in living cells, using micro-injected antibody, caused a striking relocalization of UBF from foci within the nucleoli to spots located at the nucleolar rim or within the nucleoplasm. pKi-67-CALI resulted in dramatic inhibition of RNA polymerase I-dependent nucleolar rRNA synthesis, whereas RNA polymerase II-dependent nucleoplasmic RNA synthesis remained almost unaltered. CONCLUSIONS: Our data presented here argue for a crucial role of pKi-67 in RNA polymerase I-dependent nucleolar rRNA synthesis.

Rare cancer cell analyzer for whole blood applications: microcytometer cell counting and sorting subcircuits.

We demonstrate sorting of rare cancer cells from blood using a thin ribbon monolayer of cells within a credit-card sized, microfluidic laboratory-on-a-card ("lab card") structure. This enables higher cell throughput per minute thereby speeding up cell interrogation. In this approach, multiple cells are viewed and sorted, not individually, but as a whole cell row or section of the ribbon at a time. Gated selection of only the cell rows containing a tagged rare cell provides enrichment of the rare cell relative to background blood cells. We also designed the cell injector for laminar flow antibody labeling within 20s. The approach combines rapid laminar flow cell labeling with monolayer cell sorting thereby enabling rare cell target detection at sensitivity levels 1000 to 10,000 times that of existing flow cytometers. Using this method, total cell labeling and data acquisition time on card may be reduced to a few minutes compared to 30-60 min for standard flow methods.

Rare cancer cell analyzer for whole blood applications: automated nucleic acid purification in a microfluidic disposable card.

One current challenge facing point-of-care cancer detection is that existing methods make it difficult, time consuming and too costly to (1) collect relevant cell types directly from a patient sample, such as blood and (2) rapidly assay those cell types to determine the presence or absence of a particular type of cancer. We present a proof of principle method for an integrated, sample-to-result, point-of-care detection device that employs microfluidics technology, accepted assays, and a silica membrane for total RNA purification on a disposable, credit card sized laboratory-on-card ('lab card") device in which results are obtained in minutes. Both yield and quality of on-card purified total RNA, as determined by both LightCycler and standard reverse transcriptase amplification of G6PDH and BCR-ABL transcripts, were found to be better than or equal to accepted standard purification methods.

Microtubule transport defects in neurological and ciliary disease.

Microtubules are primarily responsible for facilitating long-distance transport of both proteins and organelles. Given the critical role of this process in cellular function, it is not surprising that perturbation of microtubule-based transport can lead to diverse phenotypes in humans, including cancer and neurodegenerative disorders such as Alzheimer or Huntington disease. Recent investigations have also indicated that defects in specialized microtubule-based transport systems, such as mutations affecting the transport of protein particles along the length of cilia (intraflagellar transport) can cause retinal dystrophy, polycystic kidney disease or more complex syndromic phenotypes, such as Bardet-Biedl syndrome. In this review, we discuss recent findings implicating defects in microtubule-associated transport and motor proteins in a variety of diseases, particularly the role of defective microtubular transport in neurological and ciliary disease. These defects frequently display phenotypic consequences that manifest as human disease yet do not cause organismal lethality.

Expression of the proliferation marker Ki-67 during early mouse development.

In somatic tissues, the mouse Ki-67 protein (pKi-67) is expressed in proliferating cells only. Depending on the stage of the cell cycle, pKi-67 is associated with different nuclear domains: with euchromatin as part of the perichromosomal layer, with centromeric heterochromatin, and with the nucleolus. In gametes, sex-specific expression is evident. Mature MII oocytes contain pKi-67, whereas pKi-67 is not detectable in mature sperm. We investigated the re-establishment of the cell cycle-dependent distribution of pKi-67 during early mouse development. After fertilization, male and female pronuclei exhibited very little or no pKi-67, while polar bodies were pKi-67 positive. Towards the end of the first cell cycle, prophase chromosomes of male and female pronuclei simultaneously got decorated with pKi-67. In 2-cell embryos, the distribution pattern changed, presumably depending on the progress of development of the embryo, from a distribution all over the nucleus to a preferential location in the nucleolus precursor bodies (NPBs). From the 4-cell stage onwards, pKi-67 showed the regular nuclear relocations known from somatic tissues: during mitosis the protein was found covering the chromosome arms as a constituent of the perichromosomal layer, in early G1 it was distributed in the whole nucleus, and for the rest of the cell cycle it was associated with NPBs or with the nucleolus.

Are we underestimating the full potential of early thrombolytic treatment in patients with acute myocardial infarction?

Use of the Boersma curve in order to describe the beneficial effect of thrombolytic treatment at different treatment delays seems questionable, because the curve may underestimate the favourable prognostic effects of early thrombolysis in patients with acute myocardial infarction

Telemedicine used for remote prehospital diagnosing in patients suspected of acute myocardial infarction.

INTRODUCTION: In patients with acute myocardial infarction (AMI), considerable time elapses from symptom onset until initiation of thrombolytic therapy or primary percutaneous coronary intervention. Prehospital diagnosing can reduce time delays, and remote diagnosing using telemedicine may be appropriate in areas and countries where ambulances are not staffed with physicians. OBJECTIVES: To evaluate whether it was technically feasible for physicians at a remote university hospital to diagnose ST-segment-elevation-AMI (AMI(STelev)) in patients suspected of AMI, who were transported by ambulances to a local hospital. To determine associated prehospital delays and in-hospital treatment delays. METHODS: Patients carried in telemetry equipped ambulances had 12-lead electrocardiograms (ECGs) acquired as soon as possible. En route to the local hospital the ECGs were transmitted to a remote university hospital, by use of the GSM-system. The physician on call at the university hospital interviewed the patients, who were provided with cellular phone headsets, and alerted the local hospital if signs of AMI(STelev), bundle-branch-block-AMI or malignant arrhythmia were detected. Patients transported by traditional ambulances were included in a prospective control group. RESULTS: In 214 (86%) of 250 patients prehospital diagnosing was successful. Geographically related transmission problems were the primary reason for failure. Ninety-eight per cent of transmitted electrocardiograms and obtained history takings were technically acceptable for diagnostic purposes. Door-to-needle times were shorter amongst patients with AMI(STelev) who were subjected to prehospital diagnosing (n = 13) as compared with patients transported by traditional ambulances (n = 14) (38 vs. 81 min) (P = 0.004). CONCLUSIONS: It was technically feasible to use telemedicine for remote prehospital diagnosing of patients suspected of AMI. Patients subjected to prehospital diagnosing had shorter door-to-needle times compared with a prospective control group.

The expression of Ki-67, MCM3, and p27 defines distinct subsets of proliferating, resting, and differentiated cells.

The mini-chromosome maintenance proteins (MCM), which are involved in the control of DNA replication, and the cyclin-dependent kinase inhibitors, such as p27/KIP1, represent two groups of proteins that are currently under investigation as diagnostic tumour markers. The expression of p27 and MCM3 was compared with the expression of the Ki-67 protein, an approved marker for proliferating cells, extensively used in histopathology and cancer research. The expression pattern of all three proteins was assessed on germinal centres and oral mucosa, which display a well-defined spatio-temporal organization. The expression of the p27 protein was closely related to differentiated cells, whereas MCM3 and Ki-67 were predominantly localized to the regions of proliferating cells. However, it is important to note that considerable numbers of cells that were growth-arrested, as confirmed by the absence of the Ki-67 protein, stained positive for the MCM3 protein. These results were verified in vitro using growth-arrested Swiss 3T3. The MCM3 protein is therefore expressed in cells that have ceased to proliferate, but are not terminally differentiated, according to the absence of p27 protein expression. In conclusion, a combined analysis of Ki-67, MCM3, and p27 protein expression may provide a more detailed insight into the cell proliferation and differentiation processes that determine individual tumour growth.

Reduced T-cell receptor CD3zeta-chain protein and sustained CD3epsilon expression at the site of mycobacterial infection.

Control of mycobacterial infection by the cellular immune system relies both on antigen-presenting cells and on T lymphocytes. The quality of an effective cellular immune response is dependent on functional signal transduction residing in the cytoplasmic tails of the T-cell receptor CD3 components. In order to investigate potential effects of mycobacteria on T-cell receptor signalling, we examined the protein expression of T-cell signal transduction molecules (CD3zeta, ZAP-70, p59fyn, p56lck). In Western blots of peripheral blood mononuclear cells of Mycobacterium tuberculosis infected patients, only the CD3zeta-chain showed a marked reduction in protein expression. To investigate the situation in situ, immunoenzymatic and immunofluorescence stainings for CD3epsilon and CD3zeta expression were performed on sections of normal lymphoid tissue, M. leprae infected and sarcoid tissue. CD3epsilon and CD3zeta expression were similar with respect to intensity, localization and the number of cells stained in normal lymphoid tissue and in sarcoid granulomas. In contrast, the granulomas of M. leprae infected tissues showed a significantly reduced expression of CD3zeta compared to CD3epsilon. Using double immunofluorescence analysis, virtually no CD3zeta expression could be detected in comparison to the CD3epsilon expression in the lesions. Apparently, mycobacteria are capable of significantly reducing CD3zeta-chain expression, which may be restored by cytokines. IL-2-enhanced zeta-chain expression and T-cell effector functions, defined by interferon-gamma release, in M. tuberculosis-specific and human leucocyte antigen-DR restricted CD4+ T cells isolated from granuloma lesions from patients with pulmonary tuberculosis. Because CD3zeta is essential for CD3 signalling and for eliciting T-cell effector functions, reduced CD3zeta protein expression could result in altered signal transduction and inefficient T-cell effector functions. Alternatively, reduced CD3zeta-chain expression may protect T cells from repetitive TCR stimulation associated with anergy or apoptosis.

Evidence for disease phenotype associated haplotypes (DR.TNF) in sarcoidosis.

BACKGROUND AND AIM OF THE WORK: In a previous study, gene polymorphisms of the MHC locus (-308 TNFalpha promotor, HLA-DR) in patients with pulmonary sarcoidosis had been investigated and significant correlations with the presentation of the disease as defined by the presence of Löfgren syndrome had been found. Since genotyping of both loci was necessary to reveal a significant difference on the genetic level between the two patient groups Löfgren and non-Löfgren, a working hypothesis was derived in which a disease course associated haplotype, rather than single specific genes, was considered to play a role in the pathogenesis of sarcoidosis. METHODS: A first step to test this hypothesis was taken by calculating virtual haplotypes from these previous data. Statistical analysis of disease phenotype association and relative risk of these virtual haplotypes was performed to assess correlations with sarcoidosis and the two disease phenotypes. RESULTS: The results not only substantiated the previous findings, but also showed that the virtual haplotype DR3.TNFalpha2 was significantly associated with Löfgren syndrome and that the virtual haplotype DR2.TNFalpha1 was noticeably although not significantly associated with the non-Löfgren patients. CONCLUSIONS: Using theoretical calculations based on actual data, haplotypes, rather than single genes interacting with each other, were found to be a highly likely explanation for the previously published observations. In addition, the results allow the conclusion to be drawn that disease course associated haplotypes in sarcoidosis are highly probable and that further investigation of polymorphisms in the MHC gene region holds the potential of defining prognostic markers.

Impaired glucocorticoid synthesis in premature infants developing chronic lung disease.

Premature infants have higher cortisol precursor concentrations than term infants; however, many sick preterm infants have surprisingly low cortisol concentrations. Those who develop chronic lung disease (CLD) have lower cortisol values than those who recover. We hypothesized that some infants have a decreased ability to synthesize cortisol, leading to physiologic disruptions including amplified inflammatory responses, thereby resulting in CLD. We measured cortisol, 11-deoxycortisol, 17-hydroxyprogesterone, 17-hydroxypregnenolone, dehydroepiandrosterone sulfate, and ACTH in 40 extremely low birth weight infants enrolled in a study of low-dose hydrocortisone therapy to prevent CLD. Thirty-four infants survived and 15 developed CLD. Hydrocortisone therapy did not suppress ACTH or any measured steroid value. Before study (<48 h of life), 17-OH progesterone was higher in CLD infants, as was the ratio of 17-OH progesterone to 11-deoxycortisol. On d 15-19 (> or =72 h after end of therapy), basal and stimulated cortisol concentrations were lower in CLD infants. In contrast, the basal ratio of 11-deoxycortisol to cortisol was higher in CLD infants, as were stimulated values of 17-OH progesterone and stimulated ratios of 17-OH progesterone to 11-deoxycortisol and 11-deoxycortisol to cortisol. Thus, infants who developed CLD had lower basal and stimulated cortisol values, but elevated cortisol precursors and precursor to product ratios, compared with infants who recovered. These data support the hypothesis that these immature infants have a decreased capacity to synthesize cortisol, which may lead to a relative adrenal insufficiency in the face of significant illness.

Immunohistochemical detection of cell growth fraction in formalin-fixed and paraffin-embedded murine tissue.

Monoclonal antibody MIB-1 is a reliable tool for determining proliferating cells in human tissues, but does not react with the homologous mouse antigen and is therefore useless in experimental pathology using mice as model systems. Standard method for assessment of cellular proliferation in formalin-fixed, paraffin-embedded murine tissues is immunohistochemical detection of DNA synthesis using antibodies against exogenously injected 5-bromodeoxyuridine (BrdU), which is a tedious procedure and not useful for routine investigations. We tested monoclonal antibody MIB-5 and monoclonal and polyclonal anti-MCM3 antibodies as immunohistochemical proliferation markers for paraffin-embedded nonneoplastic and neoplastic tissues of wild-type and transgenic mice, compared to anti-BrdU immunostaining. Percentage of proliferating cells was determined with continuously decreasing antibody dilutions. Percentages of MIB-5 and anti-BrdU immunostained cells correlated strongly, as well as percentage of MIB-5-decorated cells and frequency of mitotic figures. Anti-MCM3 antibodies labeled significantly higher percentages of cells than anti-BrdU or MIB-5, and showed a linear decrease with increasing antibody dilutions. We conclude that MIB-5 detects reliably the cell growth fraction in formalin fixed, paraffin-embedded murine tissues, bypassing methodological drawbacks of BrdU. Anti-MCM3 antibodies are less useful for determination of proliferating cells although they might detect the fraction of cells remaining competent for proliferation.

The fate of bacillus Calmette-Guerin after intravesical instillation.

PURPOSE: Long-term activation of immunocompetent cells of the bladder wall as well as case reports of systemic infections some months or years after intravesical bacillus Calmette-Guerin (BCG) therapy imply that mycobacteria may persist in the body. Therefore. we investigated the fate of BCG in patients after uncomplicated intravesical instillation therapy. MATERIAL AND METHODS: A total of 49 patients were included in the study, from whom various numbers of specimens were used for mycobacterial culture and molecular biological detection techniques. In 23 patients who received a total of 128 instillations urine, sputum, venous blood and bladder biopsies were screened for BCG by acid-fast staining and culture at different times before and after instillation. From 16 of the 23 patients and from an additional 26 a total of 180 bladder biopsies obtained at intervals 3 to 30 months after instillation were screened for mycobacterial 16S ribosomal DNA by a nested polymerase chain reaction protocol. RESULTS: No viable BCG was found in venous blood or in 127 of 128 sputum specimens before and 2 hours after instillation. Two of 56 bladder biopsies were culture positive. In urine BCG was detected in 96.4% of the specimens after 2 hours and in 67.9% after 24 hours after instillation. The number of positive specimens decreased and it was 27.1% on day 7 immediately before the next instillation. In 14 of 44 bladder biopsies (31.8%) mycobacterial ribosomal DNA was found within 1 week after the sixth instillation. A positive polymerase chain reaction was evident up to 24 months in between 4.2% and 37.5% of the investigated biopsies. After 30 months no ribosomal DNA was evident in the 6 samples available for testing. CONCLUSIONS: Nontraumatic intravesical instillation of BCG is not accompanied by systemic mycobacterial spread. Local persistence during the instillation course is evident since viable BCG is commonly found in the urine. Long lasting and persistent BCG DNA in the bladder wall may account for long-term immuno-activation. However, the remaining BCG may be a possible source of late systemic infections.