Study on the interaction between 4-(1H-indol-3-yl)-2-(p-tolyl)quinazoline-3-oxide and human serum albumin

An organic small-molecular drug, 4-(1H-indol-3-yl)-2-(p-tolyl)quinazoline-3-oxide 1a was synthesized. It was employed to investigate the binding interaction and mechanism with human serum albumin (HSA). The experimental results indicated that the fluorescence quenching of HSA by 1a is a static quenching process and formation 1a-HSA complex. The site competition experiments revealed that the combination of 1a on HSA are hydrophobic interactions in the IIA domain and hydrogen bonds in IIIA domain of HSA, and the hydrophobic interactions of 1a on HSA are stronger than that of hydrogen bonds. These results were also confirmed by molecular docking theoretic analysis and ANS-hydrophobic fluorescent probe experiment. Synchronous fluorescence experiments showed that the polarity of HSA microenvironment was increase in the interaction process of 1a with HSA. The results of binding distance explored indicated that the combination distance between 1a and HSA is 3.63 nm, which is between 0.5R₀ and 1.5R₀, revealing the energy transfer between HSA and 1a is non-radiative. These results are very helpful for people to screen out high efficient indoloquinazoline drugs.     

Novel Interaction of the Z-DNA Binding Domain of Human ADAR1 with the Oncogenic c-Myc Promoter G-Quadruplex

Both G-quadruplex and Z-DNA can be formed in G-rich and repetitive sequences on genome, and their formation and biological functions are controlled by specific proteins. Z-DNA binding proteins, such as human ADAR1, have a highly conserved Z-DNA binding domain having selective affinity to Z-DNA. Here, our study identifies the Z-DNA binding domain of human ADAR1 (hZα_ADAR1) as a novel G-quadruplex binding protein that recognizes c-myc promoter G-quadruplex formed in NHEIII₁ region and represses the gene expression. An electrophoretic migration shift assay shows the binding of hZα_ADAR1 to the intramolecular c-myc promoter G-quadruplex-forming DNA oligomer. To corroborate the binding of hZα_ADAR1 to the G-quadruplex, we conducted CD and NMR chemical shift perturbation analyses. CD results indicate that hZα_ADAR1 stabilizes the parallel-stranded conformation of the c-myc G-quadruplex. The NMR chemical shift perturbation data reveal that the G-quadruplex binding region in hZα_ADAR1 was almost identical with the Z-DNA binding region. Finally, promoter assay and Western blot analysis show that hZα_ADAR1 suppresses the c-myc expression promoted by NHEIII₁ region containing the G-quadruplex-forming sequence. This finding suggests a novel function of Z-DNA binding protein as a regulator of G-quadruplex-mediated gene expression.     

Total bile acids in hyperlipidaemic serum determined by bioluminescent and spectrophotometric methods

A simple, rapid and sensitive bioluminescent method has been used to measure total bile acids in hyperlipidaemic serum. We found that the levels of total bile acids in hypertriglyceridaemic and hypercholesterolemic sera determined by a spectrophotometric method were four-fold higher than those measured by the bioluminescent method (6.73 ± 4.07 μmol/l (mean ± SD) by bioluminescent and 26.10 ± 13.42 μmol/l by the spectrophotometric method). There was no difference in total bile acid levels between these two methods for normal serum (4.72 ± 3.38 μmol/l by bioluminescence and 4.49 ± 3.27 μmol/l by the spectrophotometric method).     

Can the problem of hybridization in threatened species be evaluated using a fieldwork research? A case study in snapdragons

Hybridization, natural or artificial, is considered disadvantageous for species biodiversity when it threatens the population integrity of endangered species. Frequently, studies investigating whether hybridization poses a legitimate risk to rare species are based on genetic data obtained in molecular biology laboratories. In this study, we used field research to approach the problem that hybridization could cause for the viability of a population of a rare species and to be able to propose the most appropriate initial conservation strategy. Specifically, using the model genus Antirrhinum, the reproductive barriers between the rare A. pulverulentum and its common congener A. litigiosum have been analysed under the reproductive isolation index (RI). A. pulverulentum had a high value of total RI, indicating that there are barriers to gene flow from A. litigiosum towards this species, and also had a high value for the intrinsic RI, reflecting a low inherent capacity for production of hybrid plants; in addition, the possibility of successful backcrosses between this species with hybrids produced from A. litigiosum ovules were low, given the high intrinsic RI of A. litigiosum. These data indicate the current existence of strong and permanent barriers to hybridization between the two species, suggesting that hybridization does not seem to be a serious problem for the conservation of A. pulverulentum in the studied population, nor for the near future. This study shows how the RI index can provide useful information for conservation purposes and proposes different management recommendations.     

Unequivocal determination of sire allele origin for multiallelic microsatellites when only the sire and progeny are genotyped

The effect of a segregating economic trait locus (ETL) can be detected with the aid of a linked genetic marker, if specific alleles of each locus are in association among the individuals genotyped for the genetic marker. For dairy cattle this can be achieved by application of the ‘granddaughter design’. If only the sires and their sons are genotyped for the genetic markers, then the allele origin of sons having the same genotypes as their sires cannot be determined. Seven sires and 101 sons were genotyped for five microsatellites. The mean frequency of heterozygous sires was 77%. The mean number of alleles per locus was 8.2. Frequency of informative sons per locus ranged from 60% to 80% with a mean of 72%. With highly polymorphic microsatellites, at least 60% more grandsire families can be included in the analysis, and the number of sons assayed can be reduced by 40%, as compared to diallelic markers.     

Genetic and allelopathic differences between populations of daisy fleabane Erigeron annuus (L.) Pers. (Asteraceae) from disturbed and stable habitats

The interactions between invasive plants and their habitats may vary at different phases of the invasion process and depend on the phenotypic plasticity or local adaptations of each species. In this study, we investigated whether habitat changes during the invasion process are related to variations in the physiological traits (allelopathic properties) and genetic differentiation of daisy fleabane (Erigeron annuus (L.) Pers.). E. annuus is a winter annual invasive species that originated in North America and is now distributed throughout Europe. Genetic and genotypic diversity analyses were performed for 37 populations of E. annuus based on inter simple sequence repeat (ISSR) polymorphisms. In total, 684 plants were analyzed; 342 were from stable habitats and 342 were from disturbed habitats. The genetic differences among the populations from the different habitats were studied using a Bayesian cluster analysis and an analysis of molecular variance (AMOVA) and by calculating the genetic and genotypic diversity parameters. A germination test using the juglone index was employed to examine the potential allelopathic properties of the plants from the different habitats. Bayesian cluster analysis, AMOVA and allelopathic effects evaluation revealed differences in the allelopathic potential and genetic structure of the E. annuus populations from the disturbed and stable habitats. This differentiation of populations could be associated with founder effects or with different selection pressures among habitats.     

Sampling Blood from the Lateral Tail Vein of the Rat

Blood samples are commonly obtained in many experimental contexts to measure targets of interest, including hormones, immune factors, growth factors, proteins, and glucose, yet the composition of the blood is dynamically regulated and easily perturbed. One factor that can change the blood composition is the stress response triggered by the sampling procedure, which can contribute to variability in the measures of interest. Here we describe a procedure for blood sampling from the lateral tail vein in the rat. This procedure offers significant advantages over other more commonly used techniques. It permits rapid sampling with minimal pain or invasiveness, without anesthesia or analgesia. Additionally, it can be used to obtain large volume samples (upwards of 1 ml in some rats), and it may be used repeatedly across experimental days. By minimizing the stress response and pain resulting from blood sampling, measures can more accurately reflect the true basal state of the animal, with minimal influence from the sampling procedure itself.     

A mannose-specific lectin from Vicia villosa seeds

A lectin specific to mannose has been purified from Vicia villosa seed by (NH₄)₂SO₄ fractionation, GalNAc-Sepharose and Man-Sepharose affinity chromatography. It was defined as VVL_M, which showed a single band on an acidic-PAGE stained with Coosmassie brilliant blue. The molecular weight of VVL_M was 50 kDa as determined by gel filtration on Biogel P-100 column. The VVL_M molecule consists of 2 distinct subunits with apparent molecular weight of 30 kDa and 22kDa determined by SDS-PAGE. VVL_M has at least four isolectins with similar haemagglutinating activity. Its extinction coefficient is calculated as A_1cm¹ = 16.4 at 280 nm. Sugars could not be detected phenol-sulfuric acid method. The circular dichroism analysis at far UV indicated that VVL_M was a β-sheet-rich protein, and gave no α-helix, 69% β-sheet, 14% β-turn by Provencher and Glockner method. The lectin was inhibited by α-methyl-d-mannose at 12.5 mM and glucose or GlcNAc at 50 mM. The carbohydrate binding specificity of VVL_M was investigated by using affinity chromatography on a VVL_M-Sepharose column. Among various Asn-linked oligosaccharides, core structure Manα1→3(Manα1→6)Manβ1→4GlcNAcβ1→4GlcNAc_OT were found to have high affinity for VVL_M-Sepharose. The antisera of VVL_M did not produce precipitin line with VVL_G in agar double diffusion plate indicating so serological relationship between VVL_M and VVL_G. However VVL_M showed similar immunodeterminants of some other lectins of mannose specificity such as Con A, PSL, LCA and VFL.     

Doxorubicin-induced alterations in kidney functioning,oxidative stress,DNA damage,and renal tissue morphology; Improvement by Acacia hydaspica tannin-rich ethyl acetate fraction

Doxorubicin (DOX) is an anthracycline drug used for cancer treatment. However, its treatment is contiguous with toxic effects. We examined the nephroprotective potential of A. hydaspica polyphenol-rich ethyl acetate extract (AHE) against DOX persuaded nephrotoxicity. 36 male Sprague Dawley rats were randomly assorted into 6 groups. Control group received saline; DOX group: 3 mg/kg b.w. dosage of DOX intraperitoneally for 6 weeks (single dose/week). In co-treatment groups, 200 and 400 mg/kg b.w AHE was given orally for 6 weeks in concomitant with DOX (3 mg/kg b.w, i.p. injection per week) respectively. Standard group received silymarin 400 mg/kg b.w daily + DOX (single dose/week). Biochemical kidney function tests, oxidative stress markers, genotoxicity, antioxidant enzyme status, and histopathological changes were examined. DOX caused significant body weight loss and decrease kidney weight. DOX-induced marked deterioration in renal function indicators in both urine and serum, i.e., PH, specific gravity, total protein, albumin, urea, creatinine, uric acid, globulin, blood urea nitrogen, etc. Also, DOX treatment increases renal tissue oxidative stress markers, while lower antioxidant enzymes in tissue along with degenerative alterations in the renal tissue compared to control rats. AHE co-treatment ameliorates DOX-prompted changes in serum and urine chemistry. Likewise, AHE treatment decreases sensitive markers of oxidative stress and prevented DNA damages by enhancing antioxidant enzyme levels. DOX induction in rats also caused DNA fragmentation which was restored by AHE co-treatment. Moreover, the histological observations evidenced that AHE effectively rescued the kidney tissue from DOX interceded oxidative damage. Our results suggest that co-treatment of AHE markedly improve DOX-induced deleterious effects in a dose-dependent manner. The potency of AHE co-treatment at 400 mg/kg dose is similar to silymarin. These outcomes revealed that A. hydaspica AHE extract might serve as a potential adjuvant that avoids DOX-induced nephrotoxicity.