Meibum lipid hydrocarbon chain branching and rheology after hematopoietic stem cell transplantation

Purpose: Meibum from donors who have had hematological stem cell transplantations (MHSCT) are susceptible to severe dry eye symptoms and exhibit very high lipid order (stiffness) compared with meibum from donors without dry eye (Mn). Since lipid order could have functional consequences, we compared the rheology and composition of Mn and MHSCT to measure meibum compositional, structural and functional relationships. Methods: The rheology and composition was measured using Langmuir trough and 1H NMR spectroscopy, respectively. Results: MHSCT and Mn was studied from 16 to 43 donors, respectively, using NMR spectroscopy. MHSCT contained significantly 16% more straight chain and 24% less iso-chain hydrocarbons compared with Mn. The cholesteryl ester to wax ester molar ratio, and hydrocarbon chain unsaturation were not significantly different, for MHSCT compared with Mn. Surface pressure-area isotherms of meibum from 30 donors without dry-eye were grouped into 4 pools (PC) and meibum from 32 donors with dry eye who had hematopoietic stem cell transplantation (PT) were grouped into 3 pools. Above 15 years of age the Пmax and (Cs−1)max increased with age for both the PC and the PT cohorts. (Cs−1)max values were higher for PT samples compared with age matched PC samples, indicating they had higher elasticity and stiffness. A more ordered lipid could contribute to the formation of a discontinuous patchy tear film lipid layer, which in turn results in deteriorated spreading, and decreased surface elasticity. Conclusions: The composition and rheology of meibum from donors with dry eye and who have had HSCT support the idea from other studies that more ordered meibum may contribute to or be a marker of dry eye

Influence of climate on clinical diagnostic dry eye tests: pilot study

Purpose. To analyze dry eye disease (DED) tests and their consistency in similar nonsymptomatic population samples living in two geographic locations with different climates (Continental vs. Atlantic). Methods. This is a pilot study including 14 nonsymptomatic residents from Valladolid (Continental climate, Spain) and 14 sex-matched and similarly aged residents from Braga (Atlantic climate, Portugal); they were assessed during the same season (spring) of two consecutive years. Phenol red thread test, conjunctival hyperemia, fluorescein tear breakup time, corneal and conjunctival staining, and Schirmer test were evaluated on three different consecutive visits. Reliability was assessed using the intraclass correlation coefficient and weighted kappa (J) coefficient for quantitative and ordinal variables, respectively. Results. Fourteen subjects were recruited in each city with a mean (TSD) age of 63.0 (T1.7) and 59.1 (T0.9) years (p = 0.08) in Valladolid and Braga, respectively. Intraclass correlation coefficient and J values of the tests performed were below 0.69 and 0.61, respectively, for both samples, thus showing moderate to poor reliability. Subsequently, comparisons were made between the results corresponding to the middle and higher outdoor relative humidity (RH) visit in each location as there were no differences in mean temperature (p Q 0.75) despite RH values significantly differing (p e 0.005). Significant (p e 0.05) differences were observed between Valladolid and Braga samples on tear breakup time (middle RH visit, 2.76 T 0.60 vs. 5.26 T 0.64 seconds; higher RH visit, 2.61 T 0.32 vs. 5.78 T 0.88 seconds) and corneal (middle RH, 0.64 T 0.17 vs. 0.14 T 0.10; higher RH, 0.60 T 0.22 vs. 0.0 T 0.0) and conjunctival staining (middle RH, 0.61 T 0.17 vs. 0.14 T 0.08; higher RH, 0.57 T 0.15 vs. 0.18 T 0.09). Conclusions. This pilot study provides initial evidence to support that DED test outcomes assessing the ocular surface integrity and tear stability are climate dependent. Future large-sample studies should support these outcomes also in DED patients. This knowledge is fundamental for multicenter clinical trials. Lack of consistency in diagnostic clinical tests for DED was also corroborated. (Optom Vis Sci 2015;92:e284Ye289)The authors thank M. Eugenia Mateo for statistical advice. The present study was supported by grants from Junta de Castilla y Leo´n: Grupos de Excelencia (GR217), EDU-VA145A11-2, and University of Valladolid FPI-UVa-2008. This study was also funded by FEDER through the COMPETE Program and by the Portuguese Foundation for Science and Technology (FCT) in the framework of projects PTDC/SAU-BEB/098391/ 2008, PTDC/SAU-BEB/098392/2008, and the Strategic Project PESTC/FIS/UI607/2011. No funding organizations had a role in the design or conduct of this research

Raman tweezers provide the fingerprint of cells supporting the late stages of KSHV reactivation

Kaposi's sarcoma-associated herpesvirus (KSHV) has both latent and lytic phases of replication. The molecular switch that triggers a reactivation is still unclear. Cells from S phase of cell cycle provide apt conditions for an active reactivation. In order to specifically delineate the Raman spectra of cells supporting KSHV reactivation, we followed a novel approach where cells were sorted based on the state of infection (latent Vs lytic) by a flow cytometer and then analyzed by the Raman tweezers. The Raman bands at 785, 813, 830, 1095, and 1128 cm-1 are specifically altered in cells supporting KSHV reactivation. These 5 peaks make up the Raman fingerprint of cells supporting KSHV reactivation. The physiological relevance of the changes in these peaks with respect to KSHV reactivation is discussed in the following report. Originally published Journal of Cellular and Molecular Medicine, Vol. 13, No. 8b, Aug 200

Raman spectroscopy: techniques and applications in the life sciences

Raman spectroscopy is an increasingly popular technique in many areas including biology and medicine. It is based on Raman scattering, a phenomenon in which incident photons lose or gain energy via interactions with vibrating molecules in a sample. These energy shifts can be used to obtain information regarding molecular composition of the sample with very high accuracy. Applications of Raman spectroscopy in the life sciences have included quantification of biomolecules, hyperspectral molecular imaging of cells and tissue, medical diagnosis, and others. This review briefly presents the physical origin of Raman scattering explaining the key classical and quantum mechanical concepts. Variations of the Raman effect will also be considered, including resonance, coherent, and enhanced Raman scattering. We discuss the molecular origins of prominent bands often found in the Raman spectra of biological samples. Finally, we examine several variations of Raman spectroscopy techniques in practice, looking at their applications, strengths, and challenges. This review is intended to be a starting resource for scientists new to Raman spectroscopy, providing theoretical background and practical examples as the foundation for further study and exploration

Deiminated proteins and extracellular vesicles - novel serum biomarkers in whales and Orca

Peptidylarginine deiminases (PADs) are a family of phylogenetically conserved calcium-dependent enzymes which cause post-translational protein deimination. This can result in neoepitope generation, affect gene regulation and allow for protein moonlighting via functional and structural changes in target proteins. Extracellular vesicles (EVs) carry cargo proteins and genetic material and are released from cells as part of cellular communication. EVs are found in most body fluids where they can be useful biomarkers for assessment of health status. Here, serum-derived EVs were profiled, and post-translationally deiminated proteins and EV-related microRNAs are described in 5 ceataceans: minke whale, fin whale, humpback whale, Cuvier's beaked whale and orca. EV-serum profiles were assessed by transmission electron microscopy and nanoparticle tracking analysis. EV profiles varied between the 5 species and were identified to contain deiminated proteins and selected key inflammatory and metabolic microRNAs. A range of proteins, critical for immune responses and metabolism were identified to be deiminated in cetacean sera, with some shared KEGG pathways of deiminated proteins relating to immunity and physiology, while some KEGG pathways were species-specific. This is the first study to characterise and profile EVs and to report deiminated proteins and putative effects of protein-protein interaction networks via such post-translationald deimination in cetaceans, revealing key immune and metabolic factors to undergo this post-translational modification. Deiminated proteins and EVs profiles may possibly be developed as new biomarkers for assessing health status of sea mammals