Assessment of Hepatic Detoxification Activity: Proposal of an Improved Variant of the 13C-Methacetin Breath Test

Breath tests based on the administration of a ¹³C-labeled drug and subsequent monitoring of ¹³CO₂ in the breath (quantified as DOB – delta over baseline) liberated from the drug during hepatic CPY-dependent detoxification are important tools in liver function diagnostics. The capability of such breath tests to reliably indicate hepatic CYP performance is limited by the fact that ¹³CO₂ is not exclusively exhaled but also exchanged with other compartments of the body. In order to assess this bias caused by variations of individual systemic CO₂ kinetics we administered intravenously the test drug ¹³C-methacetin to 25 clinically liver-healthy individuals and monitored progress curves of DOB and the plasma concentration of ¹³C-methacetin. Applying compartment modelling we estimated for each individual a set of kinetic parameters characterizing the time-dependent exchange of the drug and of CO₂ with the liver and non-hepatic body compartments. This analysis revealed that individual variations in the kinetics of CO₂ may account for up to 30% deviation of DOB curve parameters from their mean at otherwise identical ¹³C-methacetin metabolization rates. In order to correct for this bias we introduced a novel detoxification score which ideally should be assessed from the DOB curve of a 2-step test (“2DOB”) which is initialized with the injection of a standard dose of ¹³C-labeled bicarbonate (in order to provide information on the actual CO₂ status of the individual) followed by injection of the ¹³C-labeled test drug (the common procedure). Computer simulations suggest that the predictive power of the proposed 2DOB breath test to reliably quantity the CYP-specific hepatic detoxification activity should be significantly higher compared to the conventional breath test.     

Effects of crocin in reducing DNA damage,inflammation, and oxidative stress in multiple sclerosis patients: A double‐blind,randomized, and placebo‐controlled trial

Multiple sclerosis (MS) is an autoimmune disease in which the immune system attacks the nerve cells, resulting in neurological disorders. Oxidative stress, free radicals, and neuritis have important roles in MS pathogenesis. Here, we aim to evaluate the effect of crocin on inflammatory markers, oxidative damage, and deoxyribonucleic acid (DNA) damage in the blood of patients with MS. A total of 40 patients were divided into two groups, drug and placebo‐treated groups, using random assignment. Participants of the intervention and control groups received two crocin capsules or placebo per day for 28 days, respectively. Findings revealed a significant decrease in the level of important pathogenic factors in MS, including lipid peroxidation, DNA damage, tumor necrosis factor‐alpha, and interleukin 17 as well as a significant increase in the total antioxidant capacity in the serum of patients treated with crocin compared with the placebo group. Our results suggest the beneficial and therapeutic effects of crocin in MS.     

Purification of a conjugated polysaccharide vaccine using tangential flow diafiltration

Conjugated vaccines prepared from the capsular polysaccharide of Streptococcus pneumoniae can provide immunization against invasive pneumococcal disease, meningitis, and otitis media. One of the critical steps in the production of these vaccines is the removal of free (unreacted) polysaccharides from the protein-polysaccharide conjugate. Experimental studies were performed to evaluate the effects of membrane pore size, filtrate flux, and solution conditions on the transmission of both the conjugate and free polysaccharide through different ultrafiltration membranes. Conjugate purification was done using diafiltration performed in a linearly-scalable tangential flow filtration cassette. More than 98% of the free polysaccharide was removed within a 5-diavolume diafiltration process, which is a significant improvement over previously reported results for purification of similar conjugated vaccines. These results clearly demonstrate the opportunities for using ultrafiltration/diafiltration for the final purification of conjugated vaccine products.     

Mechanical difference between white and gray matter in the rat cerebellum measured by scanning force microscopy

The mechanical properties of tissues are increasingly recognized as important cues for cell physiology and pathology. Nevertheless, there is a sparsity of quantitative, high-resolution data on mechanical properties of specific tissues. This is especially true for the central nervous system (CNS), which poses particular difficulties in terms of preparation and measurement. We have prepared thin slices of brain tissue suited for indentation measurements on the micrometer scale in a near-native state. Using a scanning force microscope with a spherical indenter of radius ～20 μm we have mapped the effective elastic modulus of rat cerebellum with a spatial resolution of 100 μm. We found significant differences between white and gray matter, having effective elastic moduli of K=294±74 and 454±53 Pa, respectively, at 3 μm indentation depth (n_g=245, n_w=150 in four animals, p<0.05; errors are SD). In contrast to many other measurements on larger length scales, our results were constant for indentation depths of 2–4 μm indicating a regime of linear effective elastic modulus. These data, assessed with a direct mechanical measurement, provide reliable high-resolution information and serve as a quantitative basis for further neuromechanical investigations on the mechanical properties of developing, adult and damaged CNS tissue.     

Bridging the gap between chemistry, physiology, and evolution: quantifying the functionality of sperm whale myoglobin mutants

This work merges a large set of previously reported thermochemical data for myoglobin (Mb) mutants with a physiological model of O₂-transport and -storage. The model allows a quantification of the functional proficiency of myoglobin (Mb) mutants under various physiological conditions, i.e. O₂-consumption rate resembling workload, O₂ partial pressure resembling hypoxic stress, muscle cell size, and Mb concentration, resembling different organism-specific and compensatory variables. We find that O₂-storage and -transport are distinct functions that rank mutants and wild type differently depending on O₂ partial pressure. Specifically, the wild type is near-optimal for storage at all conditions, but for transport only at severely hypoxic conditions. At normoxic conditions, low-affinity mutants are in fact better O₂-transporters because they still have empty sites for O₂, giving rise to a larger [MbO₂] gradient (more varying saturation curve). The distributions of functionality reveal that many mutants are near-neutral with respect to function, whereas only a few are strongly affected, and the variation in functionality increases dramatically at lower O₂ pressure. These results together show that conserved residues in wild type (WT) Mb were fixated under a selection pressure of low P_O2.     

Let-7e enhances the radiosensitivity of colorectal cancer cells by directly targeting insulin-like growth factor 1 receptor

Abnormal expression of various microRNAs (miRNAs), as regulators of biological signaling pathways, has a strong association with cancer resistance to chemotherapy and radiotherapy. The let-7 family of miRNAs as tumor suppressors have shown to be downregulated in different types of human malignancies including colorectal cancer (CRC). However, the biological function of let-7 members in the processes of resistance to radiation in CRC has not yet been completely elucidated. Insulin-like growth factor 1 receptor (IGF-1R) signaling pathway is amplified in CRC and leads to its progression, development, and also radiation resistance. So, it seems like an attractive target for anticancer therapy. In this study, by using bioinformatics analysis, it has been revealed that IGF-1R is a direct target of the let-7e member. Consistent with this, we identified that increased levels of let-7e in CRC cells reduced IGF-1R protein level and subsequently its downstream signaling pathways, which resulted in the G1 cell cycle arrest and a significant reduction in the proliferation, survival and also resistance to radiation of CRC cells. Altogether, these results suggested that let-7e by targeting the IGF-1R signaling pathway might serve as therapeutics in anticancer therapy.     

Natural Selection on the Phase-Separation Properties of FUS during 160 My of Mammalian Evolution

Protein phase separation can help explain the formation of many nonmembranous organelles. However, we know little about its ability to change in evolution. Here we studied the evolution of the mammalian RNA-binding protein Fused in Sarcoma (FUS), a protein whose prion-like domain (PLD) contributes to the formation of stress granules through liquid–liquid phase separation. Although the PLD evolves three times as rapidly as the remainder of FUS, it harbors absolutely conserved tyrosine residues that are crucial for phase separation. Ancestral reconstruction shows that the phosphorylation sites within the PLD are subject to stabilizing selection. They toggle among a small number of amino acid states. One exception to this pattern is primates, where the number of such phosphosites has increased through positive selection. In addition, we find frequent glutamine to proline changes that help maintain the unstructured state of FUS that is necessary for phase separation. Our work provides evidence that natural selection has stabilized the liquid forming potential of FUS and minimized the propensity of cytotoxic liquid-to-solid phase transitions during 160 My of mammalian evolution.     

Bilateral symmetrical lymphangiomas of the gingiva: case report

The Dabska tumor also known as Endovascular papillary angioendothelioma is a rare type of hemangioendothelioma characterized by intraluminal papillary endothelial structures. Most of these are superficial in location but occurrence in deeper tissues is also known. We describe case report of testicular Dabska tumor in a child presenting as inguinal hernia. To the best of our knowledge this is the first case report describing the occurrence of this rare entity in testis.     

Correction of PCR-bias in quantitative DNA methylation studies by means of cubic polynomial regression

DNA methylation profiling has become an important aspect of biomedical molecular analysis. Polymerase chain reaction (PCR) amplification of bisulphite-treated DNA is a processing step that is common to many currently used methods of quantitative methylation analysis. Preferential amplification of unmethylated alleles-known as PCR-bias-may significantly affect the accuracy of quantification. To date, no universal experimental approach has been reported to overcome the problem. This study presents an effective method of correcting biased methylation data. The procedure includes a calibration performed in parallel to the analysis of the samples under investigation. DNA samples with defined degrees of methylation are analysed. The observed deviation of the experimental results from the expected values is used for calculating a regression curve. The equation of the best-fitting curve is then used for correction of the data obtained from the samples of interest. The process can be applied irrespective of the locus interrogated and the number of sites analysed, avoiding an optimization of the amplification conditions for each individual locus.