Hydrophilic Interaction Liquid Chromatography-Hydrogen/Deuterium Exchange-Mass Spectrometry (HILIC-HDX-MS) for Untargeted Metabolomics.

Liquid chromatography with mass spectrometry (LC-MS)-based metabolomics detects thousands of molecular features (retention time-/ pairs) in biological samples per analysis, yet the metabolite annotation rate remains low, with 90% of signals classified as unknowns. To enhance the metabolite annotation rates, researchers employ tandem mass spectral libraries and challenging in silico fragmentation software. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) may offer an additional layer of structural information in untargeted metabolomics, especially for identifying specific unidentified metabolites that are revealed to be statistically significant.

Triacylglycerols containing branched palmitic acid ester of hydroxystearic acid (PAHSA) are present in the breast milk and hydrolyzed by carboxyl ester lipase.

Breast milk is a complex mixture containing underexplored bioactive lipids. We performed an observational case-control study to compare the impact of delivery mode: caesarean section (CS) and vaginal birth (VB); and term (preterm and term delivery) on the levels of lipokines in human milk at different stages of lactation. Metabolomic analysis of the milk identified triacylglycerol estolides as a metabolic reservoir of the anti-inflammatory lipid mediator 5-palmitic acid ester of hydroxystearic acid (5-PAHSA).

Metabolomics atlas of oral 13C-glucose tolerance test in mice.

Glucose tolerance represents a complex phenotype in which many tissues play important roles and interact to regulate metabolic homeostasis. Here, we perform an analysis of C-glucose tissue distribution, which maps the metabolome and lipidome across 12 metabolically relevant mouse organs and plasma, with integrated C-glucose-derived carbon tracing during oral glucose tolerance test (OGTT). We measure time profiles of water-soluble metabolites and lipids and integrate the global metabolite response into metabolic pathways.

Branched and linear fatty acid esters of hydroxy fatty acids (FAHFA) relevant to human health.

Fatty acid esters of hydroxy fatty acids (FAHFAs) represent a complex lipid class that contains both signaling mediators and structural components of lipid biofilms in humans. The majority of endogenous FAHFAs share a common chemical architecture, characterized by an estolide bond that links the hydroxy fatty acid (HFA) backbone and the fatty acid (FA). Two structurally and functionally distinct FAHFA superfamilies are recognized based on the position of the estolide bond: omega-FAHFAs and in-chain branched FAHFAs.

Distinct roles of adipose triglyceride lipase and hormone-sensitive lipase in the catabolism of triacylglycerol estolides.

Branched esters of palmitic acid and hydroxy stearic acid are antiinflammatory and antidiabetic lipokines that belong to a family of fatty acid (FA) esters of hydroxy fatty acids (HFAs) called FAHFAs. FAHFAs themselves belong to oligomeric FA esters, known as estolides. Glycerol-bound FAHFAs in triacylglycerols (TAGs), named TAG estolides, serve as metabolite reservoir of FAHFAs mobilized by lipases upon demand.

Understanding FAHFAs: From structure to metabolic regulation.

The discovery of branched fatty acid esters of hydroxy fatty acids (FAHFAs) in humans draw attention of many researches to their biological effects. Although FAHFAs were originally discovered in insects and plants, their introduction into the mammalian realm opened new horizons in bioactive lipid research. Hundreds of isomers from different families have been identified so far and their role in (patho) physiological processes is currently being explored.

Triacylglycerol-Rich Oils of Marine Origin are Optimal Nutrients for Induction of Polyunsaturated Docosahexaenoic Acid Ester of Hydroxy Linoleic Acid (13-DHAHLA) with Anti-Inflammatory Properties in Mice.

Scope: The docosahexaenoic acid ester of hydroxy linoleic acid (13-DHAHLA) is a bioactive lipid with anti-inflammatory properties from the family of fatty acid esters of hydroxy fatty acids (FAHFA).

Methods And Results: To explore the biosynthesis of 13-DHAHLA from dietary oils, C57BL/6N mice are gavaged for 8 days with various corn oil/marine oil mixtures containing the same amount of DHA. Plasma levels of omega-3 FAHFAs are influenced by the lipid composition of the mixtures but do not reflect the changes in bioavailability of polyunsaturated fatty acids in plasma.

Lipokine 5-PAHSA Is Regulated by Adipose Triglyceride Lipase and Primes Adipocytes for De Novo Lipogenesis in Mice.

Branched esters of palmitic acid and hydroxystearic acid (PAHSA) are anti-inflammatory and antidiabetic lipokines that connect glucose and lipid metabolism. We aimed to characterize involvement of the 5-PAHSA regioisomer in the adaptive metabolic response of white adipose tissue (WAT) to cold exposure (CE) in mice, exploring the cross talk between glucose utilization and lipid metabolism. CE promoted local production of 5- and 9-PAHSAs in WAT.

IPSC-Derived Corneal Endothelial-like Cells Act as an Appropriate Model System to Assess the Impact of SLC4A11 Variants on Pre-mRNA Splicing.

Purpose: To report molecular genetic findings in six probands with congenital hereditary endothelial dystrophy (CHED) variably associated with hearing loss (also known as Harboyan syndrome). Furthermore, we developed a cellular model to determine if disease-associated variants induce aberrant SLC4A11 pre-mRNA splicing.

Methods: Direct sequencing of the entire SLC4A11 coding region was performed in five probands.

Brittle cornea syndrome: Disease-causing mutations in ZNF469 and two novel variants identified in a patient followed for 26 years.

Aims: Brittle cornea syndrome (BCS) is a rare autosomal recessive disorder. The aim of this study was to review ZNF469 mutations associated with BCS type 1 to date and to describe an additional case of Czech/Polish background.

Methods: Whole genome sequencing was undertaken to identify the molecular genetic cause of disease in the proband.

Characterization and comparison of human limbal explant cultures grown under defined and xeno-free conditions.

Human limbal epithelial cells (LECs) intended for treatment of limbal stem cell deficiency are commonly cultivated on a 3T3 feeder layer with complex culture medium supplemented with fetal bovine serum (FBS). However, FBS is a xenogeneic component containing poorly characterised constituents and exhibits quantitative and qualitative lot-to-lot variations. Human limbal explants were plated on untreated or fibrin coated plastic plates and cultured in two non-xenogeneic media (supplemented with either human serum or platelet lysate only).

The enzymatic de-epithelialization technique determines denuded amniotic membrane integrity and viability of harvested epithelial cells.

The human amniotic membrane (HAM) is widely used for its wound healing effect in clinical practice, as a feeder for the cell cultivation, or a source of cells to be used in cell therapy. The aim of this study was to find effective and safe enzymatic HAM de-epithelialization method leading to harvesting of both denuded undamaged HAM and viable human amniotic epithelial cells (hAECs). The efficiency of de-epithelialization using TrypLE Express, trypsin/ ethylenediaminetetraacetic (EDTA), and thermolysin was monitored by hematoxylin and eosin staining and by the measurement of DNA concentration.

The application of autologous serum eye drops in severe dry eye patients; subjective and objective parameters before and after treatment.

Aim: To assess the impact of autologous serum (AS) eye drops on the ocular surface of patients with bilateral severe dry eye and to draw a comparison between the clinical and laboratory examinations and the degree of subjective symptoms before and after serum treatment.

Materials And Methods: A three-month prospective study was conducted on 17 patients with severe dry eye. AS eye drops were applied a maximum of 12 times a day together with regular therapy.

Cytokeratin 8 is expressed in human corneoconjunctival epithelium, particularly in limbal epithelial cells.

Purpose: The purpose of this study was to investigate the expression of cytokeratin (CK) 8 in the corneoconjunctival epithelium.

Methods: In 17 cadaveric corneoscleral discs and 3 other discs, the presence of CK8 alone or CK8, together with CK3, CK15, vimentin, and integrin α6, was investigated by using indirect immunohistochemistry on radial cryosections. Four corneoscleral discs stored in organ culture were used for the preparation of tangential sections of the limbus and for the isolation of limbal epithelial cells and their subsequent cultivation.

Role of matrix metalloproteinases in recurrent corneal melting.

The aim of this study was to compare the presence and activity of matrix metalloproteinases (MMPs) 1, 2, 3, 7, 8, 9 and 13 in human melted and cadaverous corneas. Twelve melted corneal specimens from three patients with rheumatoid arthritis, one patient with ocular cicatricial pemphigoid and one patient with melting attributed to spastic entropion and ten control corneal buttons were used. The presence of MMPs was detected using indirect enzyme immunohistochemistry.

Matrix metalloproteinases in recurrent corneal melting associated with primary Sjörgen's syndrome.

Purpose: To investigate the contribution of matrix metalloproteinases (MMPs) to recurrent corneal melting in keratoconjunctivitis sicca associated with primary Sjörgen's syndrome (pSS).

Methods: One native melted cornea and ten melted corneal grafts from two patients with severe pSS were used. The presence of MMPs (1, 2, 3, 7, 8, 9, and 13) was detected using indirect enzyme immunohistochemistry.