In vivo monitoring blood‐brain barrier permeability using spectral imaging through optical clearing skull window

The blood‐brain barrier (BBB) plays a key role in the health of the central nervous system. Opening the BBB is very important for drug delivery to brain tissues to enhance the therapeutic effect on brain diseases. It is necessary to in vivo monitor the BBB permeability for assessing drug release with high resolution; however, an effective method is lacking. In this work, we developed a new method that combined spectral imaging with an optical clearing skull window to in vivo dynamically monitor BBB opening caused by 5‐aminolevulinic acid (5‐ALA)‐mediated photodynamic therapy (PDT), in which the Evans blue dye (EBd) acted as an indicator of the BBB permeability. Using this method, we effectively monitored the cerebrovascular EBd leakage process. Moreover, the analysis of changes in the vascular and extravascular EBd concentrations demonstrated that the PDT‐induced BBB opening exhibited spatiotemporal differences in the cortex. This spectral imaging method based on the optical clearing skull window provides a low‐cost and simply operated tool for in vivo monitoring BBB opening process. This has a high potential for the visualization of drug delivery to the central nervous system. Thus, it is of tremendous significance in brain disease therapy. Monitoring the changes in PDT‐induced BBB permeability by evaluating the EBd concentration using an optical clearing skull window. (A) Entire brains and coronal sections following treatment of PDT with/without an optical clearing skull window after injection of EBd. (B) Typical EBd distribution maps before and after laser irradiation captured by the spectral imaging method. (Colorbar represents the EBd concentration).      

The peculiarities of polymorphism of XPD and XRCC1 repair genes in cells of down and Ehlers-Danlo syndrome patients characterized by increased radiosensitivity

Codon 312 and 751 polymorphisms of XPD gene and codon 399 polymorphism of XRCC1 gene of peripheral blood lymphocytes in patients with Down syndrome (DS) (46 individuals) and Ehlers-Danlo syndrome (EDS) (47 individuals) and in a group of healthy donors (control) (40 individuals) were studied. The frequency of XPD genotype (G312G) coding for the most effectively functioning form of XPD protein was lower in patients with DS (26%) than in the group of healthy donors (42.5%) (p = 0.035), whereas no significant differences with the control were revealed for this codon in patients with EDS. No patients with XPD genotype (C751C) (p = 0.036) were revealed in the group of EDS patients, while this genotype was found in 16% of the group of healthy donors and in 17% of patients with DS. A trend of XRCC1 genotype frequency reduction (A399A) (p = 0.085) in EDS patients (3.9%) compared with the group of healthy donors (13.5%) and DS patients (13.3%) was obtained. These data showed that polymorphisms of the excision repair genes under study were accompanied by an elevated individual radiosensitivity in patients with DS. Genes investigated (their polymorphic variants) did not participate in the mechanisms for radiosensitive phenotype formation in EDS patients.     

Differential Activity of Superoxide Dismutase Inhibitor in Human Cells under the Action of Radiation, Chemical Mutagens, and upon Radioadaptive Response

The superoxide dismutase (SOD) inhibitor, TRIEN, which enhanced the formation of -induced DNA breaks in cells of healthy donors and patients with Marfan syndrome and Bloom syndrome (repair-defective hereditary diseases), had virtually no effect on the formation of radioadaptive response (RAR) in these systems. Similar results were obtained in studies on cell survival: TRIEN facilitated mortality in cells irradiated with -rays but did not affect RAR formation. TRIEN also increased the deleterious effect of CdCl₂, which indicates that SOD apparently plays a certain role in cell defence against this mutagen.     

Evaluation of clinical significance of с.2956G&gt;A (rs112287730) polymorphism in <Emphasis Type="Italic">FBN1</Emphasis> gene in Marfan syndrome

Marfan syndrome (MFS) is an autosomal dominant inherited systemic disorder of connective tissue with many clinical manifestations in the cardiovascular, skeletal, and ocular systems. MFS is caused by mutations in the fibrillin-1 (FBN1) gene. To date, about 2000 FBN1 pathogenic variants that cause MFS or related phenotypes have been described. The c.2956G>A, p.Ala986Thr substitution (exon 25) in the FBN1 gene is described in the SNP database as rs112287730 with allele frequency of 0.02%. Although numerous published data exist, the clinical significance of this variant is unknown. Some studies identify this substitution as probably a pathogenic mutation, and others, as a polymorphism. Among Russian Marfan patients, the heterozygous c.2956G>A substitution was identified in four probands; three of them had familial history. To determine the clinical significance of this substitution, a segregation analysis of DNA samples of affected and unaffected family members was conducted. In the first case, a segregation of the c.2956G>A substitution with the disease was observed in the family: this substitution was detected in the heterozygous state in the three affected members, but not in the one unaffected member. However, the opposite observation occurred in the second familial case: three affected members did not have the c.2956G>A substitution, whereas it was found in one unaffected member. In addition, the molecular-genetic analysis of 110 ethnically unrelated unexplored individuals was performed. The c.2956G>A substitution was identified in two of 220 examined chromosomes (allele frequency 0.9%). Thus, it was established that the c.2956G>A substitution appears to be a polymorphism (nonpathogenic variant) and cannot cause MFS.