Impact of Robotic Assistance on Precision of Vitreoretinal Surgical Procedures

Purpose

To elucidate the merits of robotic application for vitreoretinal maneuver in comparison to conventional manual performance using an in-vitro eye model constructed for the present study.

Methods

Capability to accurately approach the target on the fundus, to stabilize the manipulator tip just above the fundus, and to perceive the contact of the manipulator tip with the fundus were tested. The accuracies were compared between the robotic and manual control, as well as between ophthalmologists and engineering students.

Results

In case of manual control, ophthalmologists were superior to engineering students in all the 3 test procedures. Robotic assistance significantly improved accuracy of all the test procedures performed by engineering students. For the ophthalmologists including a specialist of vitreoretinal surgery, robotic assistance enhanced the accuracy in the stabilization of manipulator tip (from 90.9 µm to 14.9 µm, P = 0.0006) and the perception of contact with the fundus (from 20.0 mN to 7.84 mN, P = 0.046), while robotic assistance did not improve pointing accuracy.

Conclusions

It was confirmed that telerobotic assistance has a potential to significantly improve precision in vitreoretinal procedures in both experienced and inexperienced hands.     

Extracellular Sphingosine-1-Phosphate: A Novel Actor in Human Glioblastoma Stem Cell Survival

Glioblastomas are the most frequent and aggressive intracranial neoplasms in humans, and despite advances and the introduction of the alkylating agent temozolomide in therapy have improved patient survival, resistance mechanisms limit benefits. Recent studies support that glioblastoma stem-like cells (GSCs), a cell subpopulation within the tumour, are involved in the aberrant expansion and therapy resistance properties of glioblastomas, through still unclear mechanisms. Emerging evidence suggests that sphingosine-1-phosphate (S1P) a potent onco-promoter able to act as extracellular signal, favours malignant and chemoresistance properties in GSCs. Notwithstanding, the origin of S1P in the GSC environment remains unknown. We investigated S1P metabolism, release, and role in cell survival properties of GSCs isolated from either U87-MG cell line or a primary culture of human glioblastoma. We show that both GSC models, grown as neurospheres and expressing GSC markers, are resistant to temozolomide, despite not expressing the DNA repair protein MGMT, a major contributor to temozolomide-resistance. Pulse experiments with labelled sphingosine revealed that both GSC types are able to rapidly phosphorylate the long-chain base, and that the newly produced S1P is efficiently degraded. Of relevance, we found that S1P was present in GSC extracellular medium, its level being significantly higher than in U87-MG cells, and that the extracellular/intracellular ratio of S1P was about ten-fold higher in GSCs. The activity of sphingosine kinases was undetectable in GSC media, suggesting that mechanisms of S1P transport to the extracellular environment are constitutive in GSCs. In addition we found that an inhibitor of S1P biosynthesis made GSCs sensitive to temozolomide (TMZ), and that exogenous S1P reverted this effect, thus involving extracellular S1P as a GSC survival signal in TMZ resistance. Altogether our data implicate for the first time GSCs as a pivotal source of extracellular S1P, which might act as an autocrine/paracrine signal contributing to their malignant properties.     

Landscape distribution of food and nesting sites affect larval diet and nest size,but not abundance of Osmia bicornis

Habitat fragmentation is a major threat for beneficial organisms and the ecosystem services they provide. Multiple‐habitat users such as wild bees depend on both nesting and foraging habitat. Thus, they may be affected by the fragmentation of at least two habitat types. We investigated the effects of landscape‐scale amount of and patch isolation from both nesting habitat (woody plants) and foraging habitat (specific pollen sources) on the abundance and diet of Osmia bicornis L. Trap‐nests of O. bicornis were studied in 30 agricultural landscapes of the Swiss Plateau. Nesting and foraging habitats were mapped in a radius of 500 m around the sites. Pollen composition of larval diet changed as isolation to the main pollen source, Ranunculus, increased, suggesting that O. bicornis adapted its foraging strategy in function of the nest proximity to main pollen sources. Abundance of O. bicornis was neither related to isolation or amount of nesting habitat nor to isolation or abundance of food plants. Surprisingly, nests of O. bicornis contained fewer larvae in sites at forest edge compared to isolated sites, possibly due to higher parasitism risk. This study indicates that O. bicornis can nest in a variety of situations by compensating scarcity of its main larval food by exploiting alternative food sources.     

Quantification of Anti‐Addictive Alkaloids Ibogaine and Voacangine in In Vivo‐ and In Vitro‐Grown Plants of Two Mexican Tabernaemontana Species

Tabernaemontana alba and Tabernaemontana arborea are Apocynaceae species used in Mexican traditional medicine for which little phytochemical information exists. In this study, preliminary gas chromatography/mass spectrometry analyses of different organs obtained from wild plants of both species identified a total of 10 monoterpenoid indole alkaloids (MIAs) and one simple indole alkaloid, nine of which were reported for the first time in these species. Furthermore, callus cultures were established from T. alba leaf explants and regeneration of whole plants was accomplished via somatic embryogenesis. The anti‐addictive MIAs ibogaine and voacangine were then quantified by gas chromatography with flame ionization detection in wild plants of both species, as well as greenhouse‐grown plants, in vitro‐grown plantlets and embryogenic callus of T. alba. Ibogaine and voacangine were present in most samples taken from the whole plants of both species, with stem and root barks showing the highest concentrations. No alkaloids were detected in callus samples. It was concluded that T. alba and T. arborea are potentially viable sources of ibogaine and voacangine, and that these MIAs can be produced through somatic embryogenesis and whole plant regeneration of T. alba. Approaches to increase MIA yields in whole plants and to achieve alkaloid production directly in cell cultures are discussed.     

Polyclonal antibodies for specific detection of tobacco host cell proteins can be efficiently generated following RuBisCO depletion and the removal of endotoxins

The production of biopharmaceutical proteins in plants requires efficient downstream processing steps that remove impurities such as host cell proteins (HCPs) and adventitious endotoxins produced by bacteria during transient expression. We therefore strived to develop effective routines for endotoxin removal from plant extracts and the subsequent use of the extracts to generate antibodies detecting a broad set of HCPs. At first, we depleted the superabundant protein ribulose‐1,5‐bisphosphate carboxylase/oxygenase (RuBisCO) for which PEG precipitation achieved the best results, preventing a dominant immune reaction against this protein. We found that a mixture of sera from rabbits immunized with pre‐depleted or post‐depleted extracts detected more HCPs than the individual sera used alone. We also developed a powerful endotoxin removal procedure using Polymyxin B for extracts from wild type plants or a combination of fiber‐flow filtration and EndoTrap Blue for tobacco plants infiltrated with Agrobacterium tumefaciens. The antibodies we generated will be useful for quality and performance assessment in future process development and the methods we present can easily be transferred to other expression systems rendering them useful in the field of plant molecular farming.     

BAG3 regulates total MAP1LC3B protein levels through a translational but not transcriptional mechanism

Autophagy is mainly regulated by post-translational and lipid modifications of ATG proteins. In some scenarios, the induction of autophagy is accompanied by increased levels of certain ATG mRNAs such as MAP1LC3B/LC3B, ATG5 or ATG12. However, little is known about the regulation of ATG protein synthesis at the translational level. The cochaperone of the HSP70 system BAG3 (BCL2-associated athanogene 3) has been associated to LC3B lipidation through an unknown mechanism. In the present work, we studied how BAG3 controls autophagy in HeLa and HEK293 cells. Our results showed that BAG3 regulates the basal amount of total cellular LC3B protein by controlling its mRNA translation. This effect was apparently specific to LC3B because other ATG protein levels were not affected. BAG3 knockdown did not affect LC3B lipidation induced by nutrient deprivation or proteasome inhibition. We concluded that BAG3 maintains the basal amount of LC3B protein by controlling the translation of its mRNA in HeLa and HEK293 cells.     

The antigenotoxic potential of dietary flavonoids

Human exposure to genotoxic agents has dramatically increased. Both endogenous (reactive species generated during physiological and pathological processes) and exogenous (UV light, ionizing radiation, alkylating agents, antimetabolites and topoisomerase inhibitors, air, water and food pollutants) factors can impair genomic stability. The cumulative DNA damage causes mutations involved in the development of cancer and other disorders (neuromuscular and neurodegenerative diseases, immune deficiencies, infertility, cardiovascular diseases, metabolic syndrome and aging). Dietary flavonoids have protective effects against DNA damage induced by different genotoxic agents such as mycotoxins, food processing-derived contaminants (polycyclic aromatic hydrocarbons, N-nitrosamines), cytostatic agents, other medications (estrogenic and androgenic hormones), nicotine, metal ions (Cd²⁺, Cr⁶⁺), radiopharmaceuticals and ionizing radiation. Dietary flavonoids exert their genoprotection by reducing oxidative stress and modulation of enzymes responsible for bioactivation of genotoxic agents and detoxification of their reactive metabolites. Data on structure–activity relationship is sometimes contradictory. Free hydroxyl groups on the B ring (catechol moiety) and C-3 position of the C ring are important structural features for the antigenotoxic activity. As dietary flavonoids are extensively metabolized, more in vivo studies are needed for a better characterization of their antigenotoxic potential.     

Mechanistic Mathematical Modeling Tests Hypotheses of the Neurovascular Coupling in fMRI

Functional magnetic resonance imaging (fMRI) measures brain activity by detecting the blood-oxygen-level dependent (BOLD) response to neural activity. The BOLD response depends on the neurovascular coupling, which connects cerebral blood flow, cerebral blood volume, and deoxyhemoglobin level to neuronal activity. The exact mechanisms behind this neurovascular coupling are not yet fully investigated. There are at least three different ways in which these mechanisms are being discussed. Firstly, mathematical models involving the so-called Balloon model describes the relation between oxygen metabolism, cerebral blood volume, and cerebral blood flow. However, the Balloon model does not describe cellular and biochemical mechanisms. Secondly, the metabolic feedback hypothesis, which is based on experimental findings on metabolism associated with brain activation, and thirdly, the neurotransmitter feed-forward hypothesis which describes intracellular pathways leading to vasoactive substance release. Both the metabolic feedback and the neurotransmitter feed-forward hypotheses have been extensively studied, but only experimentally. These two hypotheses have never been implemented as mathematical models. Here we investigate these two hypotheses by mechanistic mathematical modeling using a systems biology approach; these methods have been used in biological research for many years but never been applied to the BOLD response in fMRI. In the current work, model structures describing the metabolic feedback and the neurotransmitter feed-forward hypotheses were applied to measured BOLD responses in the visual cortex of 12 healthy volunteers. Evaluating each hypothesis separately shows that neither hypothesis alone can describe the data in a biologically plausible way. However, by adding metabolism to the neurotransmitter feed-forward model structure, we obtained a new model structure which is able to fit the estimation data and successfully predict new, independent validation data. These results open the door to a new type of fMRI analysis that more accurately reflects the true neuronal activity.     

Signal Intensities in Preoperative MRI Do Not Reflect Proliferative Activity in Meningioma

BACKGROUND: Identification of high-grade meningiomas in preoperative magnetic resonance imaging (MRI) is important for optimized surgical strategy and best possible resection. Numerous studies investigated subjectively determined morphological features as predictors of tumor biology in meningiomas. The aim of this study was to identify the predictive value of more reliable, quantitatively measured signal intensities in MRI for differentiation of high- and low-grade meningiomas and identification of meningiomas with high proliferation rates, respectively. PATIENTS AND METHODS: Sixty-six patients (56 World Health Organization [WHO] grade I, 9 WHO grade II, and 1 WHO grade I) were included in the study. Preoperative MRI signal intensities (fluid-attenuated inversion recovery [FLAIR], T1 precontrast, and T1 postcontrast as genuine and normalized values) were correlated with Ki-67 expression in tissue sections of resected meningiomas. Differences between the groups (analysis of variance) and Spearman rho correlation were computed using SPSS 22. RESULTS: Mean values of genuine signal intensities of meningiomas in FLAIR, T1 native, and T1 postcontrast were 323.9 ± 59, 332.8 ± 67.9, and 768.5 ± 165.3. Mean values of normalized (to the contralateral white matter) signal intensities of meningiomas in FLAIR, T1 native, and T1 postcontrast were 1.5 ± 0.3, 0.8 ± 0.1, and 1.9 ± 0.4. There was no significant correlation between MRI signal intensities and WHO grade or Ki-67 expression. Signal intensities did not differ significantly between WHO grade I and II/III meningiomas. Ki-67 expression was significantly increased in high-grade meningiomas compared with low-grade meningiomas (P < 0.01). Objectively measured values of MRI signal intensities (FLAIR, T1 precontrast, and T1 postcontrast enhancement) did not distinguish between high-grade and low-grade meningiomas. Furthermore, there was no association between MRI signal intensities and Ki-67 expression representing proliferative activity.Meningiomas are among the most common brain tumors. Their incidence is about 1%, and they account for almost one third of all primary intracranial masses. The majority of meningiomas are very slowly growing and nonsymptomatic or minimally symptomatic entities, discovered as incidental findings on neuroimaging [1]. The World Health Organization (WHO) classification system distinguishes 3 histological grades and 15 subtypes and is a well-accepted tool for prediction of prognosis. Although most meningiomas are benign masses, certain histological subtypes reveal very high recurrence rates despite the tumors’ seemingly total removal. Grade II (atypical) and grade III (anaplastic) meningiomas are associated with an increased risk of recurrence, are more aggressive, and show invasive behavior [2]. Grade I meningiomas are generally considered as benign tumors, but recent studies indicate substantial neurological deficits and impaired long-term survival due to tumor recurrence and stroke despite their low histopathological grading in a considerable proportion of cases [3], [4]. Increased mitotic activity (more than 4 mitoses per 10 high-power fields) and elevated Ki-67 expression (Ki-67 index of more than 5% of nuclei) are reliable histopathological markers for tumor recurrence [2].Because histopathological grading alone does not predict outcome satisfyingly, numerous studies investigated the value of preoperative magnetic resonance imaging (MRI) for prognostics. For example, Liu et al. demonstrated that hyperintensity on diffusion-weighted imaging, heterogeneous gadolinium enhancement, disruption of the arachnoid at brain tumor interface, T2 hyperintense peritumoral edema, and irregular tumor shape were independent predictors of non–grade I meningioma [5]. Other works produced comparable results, although some of these studies underline the importance of positive capsular enhancement [6], [7], whereas others emphasize the predictive value of peritumoral edema [5], [8]. All the above-cited works investigated morphological features of meningiomas summarized in subjective scoring systems, but not one of the studies objectively analyzed values of SIs in commonly used preoperative MRI sequences.Therefore, the aim of this study was to investigate the predictive value of genuine and normalized SIs of standardized preoperative MRI (T1 pre- and postcontrast, T2, and fluid-attenuated inversion recovery [FLAIR]) as in vivo predictors of proliferative activity of meningiomas.