Selenoprotein I is indispensable for ether lipid homeostasis and proper myelination.

Selenoprotein I (SELENOI) catalyzes the final reaction of the CDP-ethanolamine branch of the Kennedy pathway, generating the phospholipids phosphatidylethanolamine (PE) and plasmenyl-PE. Plasmenyl-PE is a key component of myelin and is characterized by a vinyl ether bond that preferentially reacts with oxidants, thus serves as a sacrificial antioxidant. In humans, multiple loss-of-function mutations in genes affecting plasmenyl-PE metabolism have been implicated in hereditary spastic paraplegia, including SELENOI. Herein, we developed a mouse model of nervous system-restricted SELENOI deficiency that circumvents embryonic lethality caused by constitutive deletion and recapitulates phenotypic features of hereditary spastic paraplegia. Resulting mice exhibited pronounced alterations in brain lipid composition, which coincided with motor deficits and neuropathology including hypomyelination, elevated reactive gliosis, and microcephaly. Further studies revealed increased lipid peroxidation in oligodendrocyte lineage cells and disrupted oligodendrocyte maturation both in vivo and in vitro. Altogether, these findings detail a critical role for SELENOI-derived plasmenyl-PE in myelination that is of paramount importance for neurodevelopment.

Mass spectrometry imaging protocol for spatial mapping of lipids, N-glycans and peptides in murine lung tissue.

RATIONALE: The application of matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) to murine lungs is challenging due to the spongy nature of the tissue. Lungs consist of interconnected air sacs (alveoli) lined by a single layer of flattened epithelial cells, which requires inflation to maintain its natural structure. Therefore, a protocol that is compatible with both lung instillation and high spatial resolution is essential to enable multi-omic studies on murine lung disease models using MALDI-MSI. METHODS AND RESULTS: To maintain the structural integrity of the tissue, murine lungs were inflated with 8% (w/v) gelatin for lipid MSI of fresh frozen tissues or 4% (v/v) paraformaldehyde neutral buffer for N-glycan and peptide MSI of FFPE tissues. Tissues were sectioned and prepared for enzymatic digestion and/or matrix deposition. Glycerol-free PNGase F was applied for N-glycan MSI, while Trypsin Gold was applied for peptide MSI using the iMatrixSpray and ImagePrep Station, respectively. For lipid, N-glycan and peptide MSI, α-cyano-4-hydroxycinnamic acid matrix was deposited using the iMatrixSpray. MS data were acquired with 20 µm spatial resolution using a timsTOF fleX MS instrument followed by MS fragmentation of lipids, N-glycans and peptides. For lipid MSI, trapped ion mobility spectrometry was used to separate isomeric/isobaric lipid species. SCiLS™ Lab was used to visualize all MSI data. For analyte identification, MetaboScape®, GlycoMod and Mascot were used to annotate MS fragmentation spectra of lipids, N-glycans and tryptic peptides, respectively. CONCLUSIONS: Our protocol provides instructions on sample preparation for high spatial resolution MALDI-MSI, MS/MS data acquisition and lipid, N-glycan and peptide annotation and identification from murine lungs. This protocol will allow non-biased analyses of diseased lungs from preclinical murine models and provide further insight into disease models.

Monte-Carlo simulation for calculating phakic supplementary lenses based on a thick and thin lens model using anterior segment OCT data.

BACKGROUND: Phakic lenses (PIOLs, the most common and only disclosed type being the implantable collamer lens, ICL) are used in patients with large or excessive ametropia in cases where laser refractive surgery is contraindicated. The purpose of this study was to present a strategy based on anterior segment OCT data for calculating the refraction correction (REF) and the change in lateral magnification (ΔM) with ICL implantation. METHODS: Based on a dataset (N = 3659) containing Casia 2 measurements, we developed a vergence-based calculation scheme to derive the REF and gain or loss in ΔM on implantation of a PIOL having power PIOLP. The calculation concept is based on either a thick or thin lens model for the cornea and the PIOL. In a Monte-Carlo simulation considering, all PIOL steps listed in the US patent 5,913,898, nonlinear regression models for REF and ΔM were defined for each PIOL datapoint. RESULTS: The calculation shows that simplifying the PIOL to a thin lens could cause some inaccuracies in REF (up to ½ dpt) and ΔM for PIOLs with high positive power. The full range of listed ICL powers (- 17 to 17 dpt) could correct REF in a range from - 17 to 12 dpt with a change in ΔM from 17 to - 25%. The linear regression considering anterior segment biometric data and the PIOLP was not capable of properly characterizing REF and ΔM, whereas the nonlinear model with a quadratic term for the PIOLP showed a good performance for both REF and ΔM prediction. CONCLUSION: Where PIOL design data are available, the calculation concept should consider the PIOL as thick lens model. For daily use, a nonlinear regression model can properly predict REF and ΔM for the entire range of PIOL steps if a vergence calculation is unavailable.

Leaving trusted paths: Estimating corneal keratometric index in cataract surgery eyes with zero-power implants.

PURPOSE: This study aimed to estimate the corneal keratometric index in the eyes of cataract surgery patients who received zero-power intraocular lenses (IOLs). METHODOLOGY: This retrospective study analyzed postoperative equivalent spherical refraction and axial length, mean anterior curvature radius and aqueous humor refractive index to calculate the theoretical corneal keratometric index value (nk). Data was collected from 2 centers located in France and Germany. RESULTS: Thirty-six eyes were analyzed. The results revealed a mean corneal keratometric index of 1.329 ± 0.005 for traditional axial length (AL) and 1.331 ± 0.005 for Cooke modified axial length (CMAL). Results ranged from minimum values of 1.318/1.320 to maximum values of 1.340/1.340. CONCLUSION: The corneal keratometric index is a crucial parameter for ophthalmic procedures and calculations, particularly for IOL power calculation. Notably, the estimated corneal keratometric index value of 1.329/1.331 in this study is lower than the commonly used 1.3375 index. These findings align with recent research demonstrating that the theoretical corneal keratometric index should be approximately 1.329 using traditional AL and 1.331 using CMAL, based on the ratio between the mean anterior and posterior corneal curvature radii (1.22).

Effects of flow-induced electromagnetic field and surface roughness on antifouling activity of phenolic compounds.

Microbial fouling involves the physicochemical interactions between microorganisms and solid surfaces. An electromagnetic field (EMF) may change the diffusion rates of microbial cells and the electrical double layer around the cells and contacting surfaces. In the current study, polycardanol exhibiting antibiofouling activity was modified with ferromagnetic iron oxide (IO) to investigate the EMF effects on bacterial adhesion. When there was a flow of electrolyte that contained bacterial cells, flow-induced EMF was generated according to Faraday's principle. It was observed that the IO-ionic solution (IS)-modified surfaces, with an induced current of 44, 53, 66 nA, showed decreases in the adhesion of bacteria cells more than the unmodified (polycardanol) and IO-nanoparticles-modified ones. In addition to the EMF effects, the nano-scale uniform roughness of the modified surfaces appeared to play an important role in the reduction of cell adhesion. The results demonstrated that the IOIS-modified surface (3.2 × 10-6 mM IO) had the highest antibiofouling activity.

Evaluating keratometry and corneal astigmatism data from biometers and anterior segment tomographers and mapping to reconstructed corneal astigmatism.

BACKGROUND: To compare results from different corneal astigmatism measurement instruments; to reconstruct corneal astigmatism from the postimplantation spectacle refraction and toric intraocular lens (IOL) power; and to derive models for mapping measured corneal astigmatism to reconstructed corneal astigmatism. METHODS: Retrospective single centre study involving 150 eyes treated with a toric IOL (Alcon SN6AT, DFT or TFNT). Measurements included IOLMaster 700 keratometry (IOLMK) and total keratometry (IOLMTK), Pentacam keratometry (PK) and total corneal refractive power in 3 and 4 mm zones (PTCRP3 and PTCRP4), and Aladdin keratometry (AK). Regression-based models mapping the measured C0 and C45 components (Alpin's method) to reconstructed corneal astigmatism were derived. RESULTS: Mean C0 components were 0.50/0.59/0.51 dioptres (D) for IOLMK/PK/AK; 0.2/0.26/0.31 D for IOLMTK/PTCRP3/PTCRP4; and 0.26 D for reconstructed corneal astigmatism. All corresponding C45 components ranged around 0. The prediction models had main diagonal elements lower than 1 with some crosstalk between C0 and C45 (nonzero off-diagonal elements). Root-mean-squared residuals were 0.44/0.45/0.48/0.51/0.50/0.47 D for IOLMK/IOLMTK/PK/PTCRP3/PTCRP4/AK. CONCLUSIONS: Results from the different modalities are not consistent. On average IOLMTK/PTCRP3/PTCRP4 match reconstructed corneal astigmatism, whereas IOLMK/PK/AK show systematic C0 offsets of around 0.25 D. IOLMTK/PTCRP3/PTCRP4. Prediction models can reduce but not fully eliminate residual astigmatism after toric IOL implantation.

Vector analysis of corneal astigmatism in cataractous eyes based on IOLMaster 700 biometry.

PURPOSE: The purpose of this study was to investigate the effect of the corneal back surface by comparing the keratometric astigmatism (K, derived from the corneal front surface) of a modern optical biometer against astigmatism of Total Keratometry (TK, derived from both corneal surfaces) in a large population with cataractous eyes. The results were then used to define linear prediction models to map K to TK. METHODS: From a large dataset containing bilateral biometric measurements (IOLMaster 700) in 9736 patients prior to cataract surgery, the total corneal astigmatism was decomposed into vectors for K, corneal back surface (BS), and TK. A multivariate prediction model (MV), simplified model with separation of vector components (SM) and a constant model (CM) were defined to map K to TK vector components. RESULTS: The K centroid (X/Y) showed some astigmatism with-the-rule (0.1981/-0.0211 dioptre (dpt)) whereas the TK centroid was located around zero (-0.0071/-0.0381 dpt against-the-rule) and the BS centroid showed systematic astigmatism against-the-rule (-0.2367/-0.0145 dpt). The respective TK-K centroid was located at -0.2052/-0.0302 dpt. The MV model showed the same performance (i.e. mean absolute residuum) as the SM did (0.1098 and 0.1099 dpt respectively) while the CM performed only slightly worse (0.1121 dpt mean absolute residuum). CONCLUSION: In cases where tomographic data are unavailable statistical models could be used to consider the overall contribution of the back surface to the total corneal astigmatism. Since the performance of the CM is sufficiently close to that of MV and SM we recommend using the CM which can be directly considered e.g. as surgically induced astigmatism.

The Homburg-Adelaide toric IOL nomogram: How to predict corneal power vectors from preoperative IOLMaster 700 keratometry and total corneal power in toric IOL implantation.

PURPOSE: The purpose of this study is to compare the reconstructed corneal power (RCP) by working backwards from the post-implantation spectacle refraction and toric intraocular lens power and to develop the models for mapping preoperative keratometry and total corneal power to RCP. METHODS: Retrospective single-centre study involving 442 eyes treated with a monofocal and trifocal toric IOL (Zeiss TORBI and LISA). Keratometry and total corneal power were measured preoperatively and postoperatively using IOLMaster 700. Feedforward neural network and multilinear regression models were derived to map keratometry and total corneal power vector components (equivalent power EQ and astigmatism components C0 and C45) to the respective RCP components. RESULTS: Mean preoperative/postoperative C0 for keratometry and total corneal power was -0.14/-0.08 dioptres and -0.30/-0.24 dioptres. All mean C45 components ranged between -0.11 and -0.20 dioptres. With crossvalidation, the neural network and regression models showed comparable results on the test data with a mean squared prediction error of 0.20/0.18 and 0.22/0.22 dioptres2 and on the training data the neural network models outperformed the regression models with 0.11/0.12 and 0.22/0.22 dioptres2 for predicting RCP from preoperative keratometry/total corneal power. CONCLUSIONS: Based on our dataset, both the feedforward neural network and multilinear regression models showed good precision in predicting the power vector components of RCP from preoperative keratometry or total corneal power. With a similar performance in crossvalidation and a simple implementation in consumer software, we recommend implementation of regression models in clinical practice.

Repeatability of biometric measures from the IOLMaster 700 in a cataractous population.

PURPOSE: The purpose of this study was to investigate the repeatability of biometric measures and also to assess the interactions between the uncertainties in these measures for use in an error propagation model, using data from a large patient cohort. METHODS: In this cross-sectional non-randomised study we evaluated a dataset containing 3379 IOLMaster 700 biometric measurements taken prior to cataract surgery. Only complete scans with at least 3 successful measurements for each eye performed on the same day were considered. The mean (Mean) and standard deviations (SD) for each sequence of measurements were derived and analysed. Correlations between the uncertainties were assessed using Spearman rank correlations. RESULTS: In the dataset with 677 eyes matching the inclusion criteria, the within subject standard deviation and repeatability for all parameters match previously published data. The SD of the axial length (AL) increased with the Mean AL, but there was no noticeable dependency of the SD of any of the other parameters on their corresponding Mean value. The SDs of the parameters are not independent of one another, and in particular we observe correlations between those for AL, anterior chamber depth, aqueous depth, lens thickness and corneal thickness. CONCLUSIONS: The SD change over Mean for AL measurement and the correlations between the uncertainties of several biometric parameters mean that a simple Gaussian error propagation model cannot be used to derive the effect of biometric uncertainties on the predicted intraocular lens power and refraction after cataract surgery.

Surrogate optimisation strategies for intraocular lens formula constant optimisation.

PURPOSE: To investigate surrogate optimisation (SO) as a modern, purely data-driven, nonlinear adaptive iterative strategy for lens formula constant optimisation in intraocular lens power calculation. METHODS: A SO algorithm was implemented for optimising the root mean squared formula prediction error (rmsPE, defined as predicted refraction minus achieved refraction) for the SRKT, Hoffer Q, Holladay, Haigis and Castrop formulae in a dataset of N = 888 cataractous eyes with implantation of the Hoya Vivinex hydrophobic acrylic aspheric lens. A Gaussian Process estimator was used as the model, and the SO was initialised with equidistant datapoints within box constraints, and the number of iterations restricted to either 200 (SRKT, Hoffer Q, Holladay) or 700 (Haigis, Castrop). The performance of the algorithm was compared to the classical gradient-based Levenberg-Marquardt algorithm. RESULTS: The SO algorithm showed stable convergence after fewer than 50/150 iterations (SRKT, HofferQ, Holladay, Haigis, Castrop). The rmsPE was reduced systematically to 0.4407/0.4288/0.4265/0.3711/0.3449 dioptres. The final constants were A = 119.2709, pACD = 5.7359, SF = 1.9688, -a0 = 0.5914/a1 = 0.3570/a2 = 0.1970, C = 0.3171/H = 0.2053/R = 0.0947 for the SRKT, Hoffer Q, Holladay, Haigis and Castrop formula and matched the respective constants optimised in previous studies. CONCLUSION: The SO proves to be a powerful adaptive nonlinear iteration algorithm for formula constant optimisation, even in formulae with one or more constants. It acts independently of a gradient and is in general able to search within a (box) constrained parameter space for the best solution, even where there are multiple local minima of the target function.

Performance of a simplified strategy for formula constant optimisation in intraocular lens power calculation.

PURPOSE: To investigate the performance of a simple prediction scheme for the formula constants optimised for a mean refractive prediction error. METHODS: Analysis based on a dataset of 888 eyes before and after cataract surgery with IOL implantation (Hoya Vivinex). IOLMaster 700 biometric data, power of the implanted lens and postoperative spherical equivalent refraction were used to calculate the optimised constants (.)opt for SRKT, HofferQ, Holladay and Haigis formula with an iterative nonlinear optimisation. For detuning start values by ±1.5 from (.)opt, the predicted formula constants (.)pred were calculated and compared with (.)opt. Formula performance metrics mean (MPE), median (MEDPE), mean absolute (MAPE), median absolute (MEDAPE), root mean squared (RMSPE) and standard deviation (SDPE) of the formula prediction error were analysed for (.)opt and (.)pred. RESULTS: (.)pred - (.)opt showed a 2nd order parabolic behaviour with maximal deviations up to 0.09 at the tails of detuning and a minimal deviation up to -0.01 for all formulae. The performance curves of different metrics of PE as functions of detuning variations show that the formula constants for zeroing MPE and MEDPE yield almost identical formula constants, optimisation for MAPE, MEDAPE and RMSPE yielded formula constants very close to (.)opt, and optimisation for SDPE could result in formula constants up to 0.5 off (.)opt which is unacceptable for clinical use. CONCLUSION: This simple prediction scheme for formula constant optimisation for zero mean refraction error performs excellently in our monocentric dataset, even for larger deviations of the start value from (.)opt. Further studies with multicentric data and larger sample sizes are required to investigate the performance in a clinical setting further.

A systematic review and in silico analysis of studies investigating the ischaemic penumbra proteome in animal models of experimental stroke.

Ischaemic stroke results in the formation of a cerebral infarction bordered by an ischaemic penumbra. Characterising the proteins within the ischaemic penumbra may identify neuro-protective targets and novel circulating markers to improve patient care. This review assessed data from studies using proteomic platforms to compare ischaemic penumbra tissues to controls following experimental stroke in animal models. Proteins reported to differ significantly between penumbra and control tissues were analysed in silico to identify protein-protein interactions and over-represented pathways. Sixteen studies using rat (n = 12), mouse (n = 2) or primate (n = 2) models were included. Heterogeneity in the design of the studies and definition of the penumbra were observed. Analyses showed high abundance of p53 in the penumbra within 24 hours of permanent ischaemic stroke and was implicated in driving apoptosis, cell cycle progression, and ATM- MAPK- and p53- signalling. Between 1 and 7 days after stroke there were changes in the abundance of proteins involved in the complement and coagulation pathways. Favourable recovery 1 month after stroke was associated with an increase in the abundance of proteins involved in wound healing. Poor recovery was associated with increases in prostaglandin signalling. Findings suggest that p53 may be a target for novel therapeutics for ischaemic stroke.

Upregulated selenoprotein I during lipopolysaccharide-induced B cell activation promotes lipidomic changes and is required for effective differentiation into IgM-secreting plasma B cells.

The mechanisms driving metabolic reprogramming during B cell activation are unclear, particularly roles for enzymatic pathways involved in lipid remodeling. We found that murine B cell activation with lipopolysaccharide (LPS) led to a 1.6-fold increase in total lipids that included higher levels of phosphatidylethanolamine (PE) and plasmenyl PE. Selenoprotein I (SELENOI) is an ethanolamine phospholipid transferase involved in the synthesis of both PE and plasmenyl PE, and SELENOI expression was also upregulated during activation. Selenoi knockout (KO) B cells exhibited decreased levels of plasmenyl PE, which plays an important antioxidant role. Lipid peroxidation was measured and found to increase ∼2-fold in KO vs. wild-type (WT) B cells. Cell death was not impacted by KO in LPS-treated B cells and proliferation was only slightly reduced, but differentiation into CD138 + Blimp-1+ plasma B cells was decreased ∼2-fold. This led to examination of B cell receptors important for differentiation that recognize the ligand B cell activating factor, and levels of TACI (transmembrane activator, calcium-modulator, and cytophilin ligand interactor) (CD267) were significantly decreased on KO B cells compared with WT control cells. Vaccination with ovalbumin/adjuvant led to decreased ovalbumin-specific immunoglobulin M (IgM) levels in sera of KO mice compared with WT mice. Real-time polymerase chain reaction analyses revealed a decreased switch from surface to secreted IgM in spleens of KO mice induced by vaccination or LP-BM5 retrovirus infection. Overall, these findings detail the lipidomic response of B cells to LPS activation and reveal the importance of upregulated SELENOI for promoting differentiation into IgM-secreting plasma B cells.

Bone Marrow-Derived ABCC6 Is an Essential Regulator of Ectopic Calcification In Pseudoxanthoma Elasticum.

Physiological calcification of soft tissues is a common occurrence in aging and various acquired and inherited disorders. ABCC6 sequence variations cause the calcification phenotype of pseudoxanthoma elasticum (PXE) as well as some cases of generalized arterial calcification of infancy, which is otherwise caused by defective ENPP1. ABCC6 is primarily expressed in the liver, which has given the impression that the liver is central to the pathophysiology of PXE/generalized arterial calcification of infancy. The emergence of inflammation as a contributor to the calcification in PXE suggested that peripheral tissues play a larger role than expected. In this study, we investigated whether bone marrow-derived ABCC6 contributes to the calcification in PXE. In Abcc6‒/‒ mice, we observed prevalent mineralization in several lymph nodes and surrounding connective tissues and an extensive network of lymphatic vessels within vibrissae, a calcified tissue in Abcc6‒/‒ mice. Furthermore, we found evidence of lymphangiogenesis in patients with PXE and mouse skin, suggesting an inflammatory process. Finally, restoring wild-type bone marrow in Abcc6‒/‒ mice produced a significant reduction of calcification, suggesting that the liver alone is not sufficient to fully inhibit mineralization. With evidence that ABCC6 is expressed in lymphocytes, we suggest that the adaptative immune system and inflammation largely contribute to the calcification in PXE/generalized arterial calcification of infancy.

Use of tryptic peptide MALDI mass spectrometry imaging to identify the spatial proteomic landscape of colorectal cancer liver metastases.

Colorectal cancer (CRC) is the second leading cause of cancer-related deaths worldwide. CRC liver metastases (CRLM) are often resistant to conventional treatments, with high rates of recurrence. Therefore, it is crucial to identify biomarkers for CRLM patients that predict cancer progression. This study utilised matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) in combination with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to spatially map the CRLM tumour proteome. CRLM tissue microarrays (TMAs) of 84 patients were analysed using tryptic peptide MALDI-MSI to spatially monitor peptide abundances across CRLM tissues. Abundance of peptides was compared between tumour vs stroma, male vs female and across three groups of patients based on overall survival (0-3 years, 4-6 years, and 7+ years). Peptides were then characterised and matched using LC-MS/MS. A total of 471 potential peptides were identified by MALDI-MSI. Our results show that two unidentified m/z values (1589.876 and 1092.727) had significantly higher intensities in tumours compared to stroma. Ten m/z values were identified to have correlation with biological sex. Survival analysis identified three peptides (Histone H4, Haemoglobin subunit alpha, and Inosine-5'-monophosphate dehydrogenase 2) and two unidentified m/z values (1305.840 and 1661.060) that were significantly higher in patients with shorter survival (0-3 years relative to 4-6 years and 7+ years). This is the first study using MALDI-MSI, combined with LC-MS/MS, on a large cohort of CRLM patients to identify the spatial proteome in this malignancy. Further, we identify several protein candidates that may be suitable for drug targeting or for future prognostic biomarker development.

Methods for making and observing model lipid droplets.

We present methods for making and testing the membrane biophysics of model lipid droplets (LDs). Methods are described for imaging LDs ranging in size from 0.1 to 40 µm in diameter with high-resolution microscopy and spectroscopy. With known LD compositions, membrane binding, sorting, diffusion, and tension were measured via fluorescence correlation spectroscopy (FCS), fluorescence recovery after photobleaching (FRAP), fluorescence lifetime imaging microscopy (FLIM), atomic force microscopy (AFM), and imaging flow cytometry. Additionally, a custom, small-volume pendant droplet tensiometer is described and used to measure the association of phospholipids to the LD surface. These complementary, cross-validating methods of measuring LD membrane behavior reveal the interplay of biophysical processes on lipid droplet monolayers.

Development of a pharmaceutical database as an aid to the nonclinical detection of drug-induced cardiac toxicity.

The Health and Environmental Sciences Institute (HESI) Cardiac Safety Committee designed and created a publicly accessible database with an initial set of 128 pharmacologically defined pharmaceutical agents, many with known cardiotoxic properties. The database includes specific information about each compound that could be useful in evaluating hypotheses around mechanisms of drug-induced cardiac toxicity or for development of novel cardiovascular safety assays. Data on each of the compounds was obtained from published literature and online sources (e.g., DrugBank.ca and International Union of Basic and Clinical Pharmacology (IUPHAR) / British Pharmacological Society (BPS) Guide to PHARMACOLOGY) and was curated by 10 subject matter experts. The database includes information such as compound name, pharmacological mode of action, characterized cardiac mode of action, type of cardiac toxicity, known clinical cardiac toxicity profile, animal models used to evaluate the cardiotoxicity profile, routes of administration, and toxicokinetic parameters (i.e., Cmax). Data from both nonclinical and clinical studies are included for each compound. The user-friendly web interface allows for multiple approaches to search the database and is also intended to provide a means for the submission of new data/compounds from relevant users. This will ensure that the database is constantly updated and remains current. Such a data repository will not only aid the HESI working groups in defining drugs for use in any future studies, but safety scientists can also use the database as a vehicle of support for broader cardiovascular safety studies or exploring mechanisms of toxicity associated with certain pharmacological modes of action.

Proteomic analysis of holocarboxylase synthetase deficient-MDA-MB-231 breast cancer cells revealed the biochemical changes associated with cell death, impaired growth signaling, and metabolism.

We have previously shown that the holocarboxylase synthetase (HLCS) is overexpressed in breast cancer tissue of patients, and silencing of its expression in triple-negative cancer cell line inhibits growth and migration. Here we investigated the global biochemical changes associated with HLCS knockdown in MDA-MB-231 cells to discern the pathways that involve HLCS. Proteomic analysis of two independent HLCS knockdown cell lines identified 347 differentially expressed proteins (DEPs) whose expression change > 2-fold (p < 0.05) relative to the control cell line. GO enrichment analysis showed that these DEPs were mainly associated with the cellular process such as cellular metabolic process, cellular response to stimulus, and cellular component organization or biogenesis, metabolic process, biological regulation, response to stimuli, localization, and signaling. Among the 347 identified DEPs, 64 proteins were commonly found in both HLCS knockdown clones, confirming their authenticity. Validation of some of these DEPs by Western blot analysis showed that plasminogen activator inhibitor type 2 (SerpinB2) and interstitial collagenase (MMP1) were approximately 90% decreased in HLCS knockdown cells, consistent with a 50%-60% decrease in invasion ability of knockdown cells. Notably, argininosuccinate synthase 1 (ASS1), one of the enzymes in the urea cycle, showed approximately a 10-fold increase in the knockdown cells, suggesting the crucial role of HLCS in supporting the urea cycle in the triple-negative cancer cell line. Collectively, our proteomic data provide biochemical insights into how suppression of HLCS expression perturbs global changes in cellular processes and metabolic pathways, impairing cell growth and invasion.

Novel technical developments in mass spectrometry imaging in 2020: A mini review.

The applicability of mass spectrometry imaging (MSI) has exponentially increased with the improvement of sample preparation, instrumentation (spatial resolution) and data analysis. The number of MSI publications listed in PubMed continues to grow with 378 published articles in 2020-2021. Initially, MSI was just sensitive enough to identify molecular features correlating with distinct tissue regions, similar to the resolution achieved by visual inspection after standard immunohistochemical staining. Although the spatial resolution was limited compared with other imaging modalities, the molecular intensity mapping added a new exciting capability. Over the past decade, significant improvements in every step of the workflow and most importantly in instrumentation were made, which now enables the molecular analysis at a cellular and even subcellular level. Here, we summarize the latest developments in MSI, with a focus on the latest approaches for tissue-based imaging described in 2020.

Ontogenetic development of heterocharax macrolepis eigenmann (Ostariophysi: characiformes: characidae) with comments on the form of the yolk sac in the heterocharacinae

Fishes in early developmental stages frequently have morphological features that differ from those of adult stages, and many characters found later in ontogeny are not available in initial stages. Hence, morphological descriptions of early stages are useful to provide information for the identification of eggs and larvae, a knowledge still restricted among Neotropical fishes. We studied the development of Heterocharax macrolepis, a heterocharacine whose adult specimens from the aquarium trade were kept and spawned at around 23-24ºC. A developmental series of 51 specimens was preserved, ranging from 3.2 mm notochord length to 18.6 mm standard length, covering approximately the first 73 days post-hatching. We described the development of main morphological features emphasizing those useful in the identification of H. macrolepis larvae (i.e., appearance of preopercle spine and development of the pseudotympanum). We also compared H. macrolepis with photographs taken of live larval specimens of Gnathocharax steindachneri, recently included in the Heterocharacinae. Both species have a yolk sac with a small rounded projection directed posteroventrally. Although this information is not yet available for all pertinent taxa, the different yolk sac shape in other representatives of the Characiformes may indicate that this peculiar yolk sac represents an additional synapomorphy of the Heterocharacini.

Peixes em estágios iniciais de desenvolvimento frequentemente apresentam características morfológicas distintas dos adultos e muitos caracteres presentes em estágios avançados não são disponíveis em estágios iniciais. Assim, descrições morfológicas dos estágios iniciais de desenvolvimento de peixes são úteis por fornecerem subsídios para a identificação de ovos e larvas, um conhecimento ainda escasso entre peixes Neotropicais. Estudamos o desenvolvimento de Heterocharax macrolepis, espécie de Heterocharacinae cujos exemplares adultos provenientes do aquarismo foram mantidos e reproduzidos entre 23-24ºC. Uma série de desenvolvimento de 51 exemplares foi preservada, medindo entre 3,2 mm de comprimento da notocorda e 18,6 mm de comprimento padrão, incluindo os primeiros 73 dias pós-eclosão. Descreveram-se detalhadamente os principais aspectos morfológicos, enfatizando características úteis na identificação de larvas de H. macrolepis (i.e., surgimento do espinho do pré-opérculo, desenvolvimento do pseudotímpano). Comparamos H. macrolepis com fotografias de larvas vivas de Gnathocharax steindachneri, recentemente incluída em Heterocharacinae. Ambas espécies possuem saco vitelínico com pequena projeção arredondada póstero-ventral. Ainda que a informação não esteja disponível para os táxons pertinentes, o distinto formato do saco vitelínico em outras linhagens de Characiformes pode indicar que esta forma do saco vitelínico peculiar represente sinapomorfia adicional de Heterocharacini.