THE PROTECTIVE EFFECT OF THYMOQUINONE AGAINST LEAD ACETATE INDUCED DNA DAMAGE AND ALTERATIONS IN TUMOR INITIATION GENES

objective: Several pollutants represent a significant ecological and public health concern due to their toxicity and their ability to accumulate in livingorganisms in which lead is one of them. The present investigation was designated to assess the modulating effect of thymoquinone (TQ) against leadacetate (LA) toxicity.Methods: Several endpoints were considered to design this study such as: The gene expression of tumor initiation genes (cytochrome P450 3A[CYP3A], cyclooxygenase 2 [COX2], BAX and Bcl), DNA damage and alterations in the levels of glutathione (GSH), lipid oxidation (malondialdehyde[MDA]), and protein oxidation (protein carbonyl [PC]) in male rats. About 60 male rats were used in this study which allocated in six groups (10 animaleach) and treated with LA (200 mg/kg diet), TQ (5 and 10 mg/kg b.wt.), and LA + TQ.2Results: The results revealed that LA induced significant DNA damage and alteration in the expression of CYP3A, COX2, BAX, and Bcl as well asinduced changes in GSH content and MDA and PC levels in male rats. Meanwhile, TQ was decreased significantly the toxic effect of LA in male ratswhich decreased the alterations in the gene expression and DNA damage as well as GSH content and MDA and PC levels.Conclusion: The results suggested that TQ treatment confers protection against toxicity inflicted by LA and support the contention that TQ protectionis achieved by its ability as a scavenger for free radicals generated by LA.Keywords: Thymoquinone, Lead acetate, Gene expression, DNA damage, Rats

Genetic polymorphism of five genes associated with growth traits in goat

Genetic polymorphism studies in domestic animals aim at evaluating genetic variations within and across breeds mainly for conservation purposes. In this study, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to detect polymorphisms of five candidate genes in four Egyptian and Saudi goat breeds (Barki, Zaribi, Ardi and Masri), to detect the genotype of GH, IGF1, POUIF1, MSTN and BMP15 genes in the goat breeds and their allele frequencies. Results of GH gene which encloses a Haelll endonuclease restriction site show four unique PCR-RFLP banding patterns (genotypes AA, AB, CC and CD). The frequencies of the A allele in the samples from the goat breeds varied from 0.410 to 0.620. While  IGF-1gene revealed three fragments after digestion with Haelll with genotype AA, AB and BB and the  frequencies of allele A varied from 0.432 to 0.731. Furthermore, PCR-RFLP of POUIF1 gene showed two  fragments after digestion by Pst1 endonuclease with genotype TT and CC and the frequencies of allele T varied from 0.250 to 0.840. The MSTN gene revealed three fragments after digestion with DraI with genotype AA, BB and AB and the frequencies of allele A varied from 0.240 to 0.630. Meanwhile, the BMP15 gene revealed one fragments of 112 bp for AA after digestion with Hinf1 enzyme.Key words: Goats, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), GH, IGF-1, POUIF1, MSTN, BMP-15

Mitochondrial DNA genetic variations among four horse populations in Egypt

Horses are one of the early domesticated animals in the world that changed societies and civilizations on a continent-wide scale. Due to the rare information about the genetic characterization of different horse populations in Egypt, this study aimed to identify the genetic biodiversity and relationships between four horse populations reared in Egypt. Genomic DNA was extracted and mtDNA region was amplified using polymerase chain reaction (PCR). The alignment of 384-bp amplified fragments showed the presence of 41 polymorphic sites resulting in 29 haplotypes which their sequences were submitted to GenBank under the accession numbers: KX909898-KX909926. The phylogeny tree for tested horses declared the presence of mixing maternal lineages between the four tested populations but still there are some separated lineages especially for Arabian and Thoroughbred horses. The sequences of 72 tested sequences were aligned with 13 published sequences as references, 11 of them for different Equus caballus whereas the other two reference sequences for Equus burchellii and Equus asinus. The results showed that all tested horses from the four populations are grouped with reference sequences of Equus caballus and separated from the other two reference sequences of Equus burchellii and Equus asinus. It is concluded that sequence analysis of mtDNA control region is still the most informative tool for the identification of genetic biodiversity and phylogeny of different horse breeds and populations. The horse populations reared in Egypt possess low genetic diversity and all of them are belonged to Equus caballus breed

Single nucleotide polymorphisms in the growth hormone receptor gene and Alu1 polymorphisms in the diacylglycerol acyltransferase 1 gene as related to meat production in sheep

Aim: This study aimed to investigate the polymorphisms in genes related to meat production, including growth hormone receptor (GHR) and diacylglycerol acyltransferase 1 (DGAT1) genes, in different breeds of sheep, including Barki, Najdi, and Harri. Materials and Methods: Blood samples were collected from 75 randomly selected healthy Barki, Najdi, and Harri breeds of sheep, with 25 samples per breed. GHR and DGAT1 genes were identified using a single nucleotide polymorphism assay followed by digestion with the restriction enzyme Alu1. Results: The analysis of the GHR gene sequence showed nucleotide substitutions at nt 69 in exon 10 (c.69 G > A); this mutation is considered a transition mutation. The sequences of detected SNPs in the GHR gene in the different sheep breeds were submitted to the GenBank database with accession numbers MG906773 to MG906781. The substitutions at exon 10 (c.69 G > A) results in an alteration to the amino acid (p. Lysine > Arginine). At c.69, the A allele frequency was 0.61, 0.59, and 0.54, while the G allele frequency was 0.39, 0.41, and 0.46, for Barki, Najdi, and Harri breeds, respectively. The genotype AG at nt 69 locus had the highest frequency in the Najdi and Harri sheep. The frequency of AG was 0.62, 0.61, and 0.64, while the frequency of AA was 0.30, 0.28, and 0.22, for Barki, Najdi, and Harri sheep, respectively. After digestion with the restriction enzyme AluI, the DGAT1 locus had two genotypes, CC and CT. The highest frequency, 0.88, was found for allele C, which was detected in Barki breed. The lowest frequency, 0.75, for the same allele was found for Harri. Conclusion: The detected CT genotype may explain the moderate intramuscular fat content and muscle marbling in the Barki sheep breed

Effect of Diet Restriction and Polymorphism of Bone Morphogenetic Protein-15 and Growth Differentiation Factor-9 (GDF9) on Reproductive Performance of Three Egyptian Fat Tail Sheep Breeds

Several genes are controlling prolificay of sheep. The bone morphogenetic protein-15 and growth differentiation factor-9 (GDF9) are control reproductivity of sheep. This study aimed to eplore diet restriction and the polymorphism in the bone morphogenetic protein-15 (BMP15) and growth differentiation factor-9 (GDF9) genes on ovulation and reproductive hormones in three sheep breeds. Ovaries of Rahmani ewes (R, n=56), Barki (B, n=56), and Ossimi ewes (O, n=50) were scanned to determine the preovulatory follicles and ovulation. The harvested sera were used for assaying progesterone, leptin, insulin-like growth Factor-I, and insulin. Whole blood samples were used for gene analysis using two PCR primers for amplifying the fragment 141-bp for FecXG site (exon-2) of the BMP15 gene and 139-bp for FecGH locus (exon-1) of the GDF9 gene. The output amplicons were digested (RFLP) with HinfI and DdeI endonucleases. Results revealed no mutation in the FecXG locus in all breeds. FecGH showed mutations but not in all breeds. Treated O received half dietary requirements for four weeks were the youngest (P=0.0001). Barki had the lightest bodyweight (P=0.017). Treated ewes had a higher (P=0.047) number of large follicles compared to their controls. Treated B and R got larger (P=0.0001) dominant follicles. The ovulation rate did not vary within the diet treated breeds but was lower than their control. Control O and R had higher (P=0.0001) ovulation rate, compared to B. Leptin concentrations were low (P=0.047) in treated B and R. The lowest (P=0.031) insulin concentrations were observed in treated O. In conclusion, the BMP15 loci showed no polymorphism, while the GDF9 loci were polymorphic in all breeds and treatments. Nutritional status and age modified ovulation rate in sheep

Coculture of bacterial levans and evaluation of its anti-cancer activity against hepatocellular carcinoma cell lines

Abstract This research represents a novel study to assess how coculture affects levan yield, structure, bioactivities, and molecular weight. Among the 16 honey isolates, four bacterial strains recorded the highest levan yield. The Plackett–Burman design showed that the coculture (M) of isolates G2 and K2 had the maximum levan yield (52 g/L) and the effective factors were sucrose, incubation time, and sugarcane bagasse. The CCD showed that the most proper concentrations for maximum levan yield (81 g/L): were 130 g/L of sucrose and 6 g/f of sugarcane bagasse. Levan’s backbone was characterized, and the molecular weight was determined. G2 and K2 isolates were identified based on 16 sRNA as Bacillus megaterium strain YM1C10 and Rhizobium sp. G6-1. M levan had promising antioxidant activity (99.66%), slowed the migration activity to a great extent, and recorded 70.70% inhibition against the hepatoblastoma cell line (HepG2) at 1000 µg/mL. Gene expression analysis in liver cancer cell lines (HePG2) revealed that M levan decreased the expression of CCL20), 2GRB2, and CCR6) genes and was superior to Doxo. While increasing the expression of the IL4R and IL-10 genes. The DNA damage values were significantly increased (P < 0.01) in treated liver cancer cell lines with levan M and Doxo. The results referred to the importance of each of the hydroxyl and carboxyl groups and the molecular weight in levans bioactivities

Impact of trifluoromethyl and sulfonyl groups on the biological activity of novel aryl-urea derivatives: synthesis, in-vitro, in-silico and SAR studies

Abstract We designed and prepared a novel series of urea derivatives with/without sulfonyl group in their structures to investigate the impact of the sulfonyl group on the biological activity of the evaluated compounds. Antibacterial investigations indicated that derivatives 7, 8, 9, and 11 had the most antibacterial property of all the compounds examined, their minimum inhibitory concentrations (MICs) determined against B. mycoides, E. coli, and C. albicans, with compound 8 being the most active at a MIC value of 4.88 µg/mL. Anti-cancer activity has been tested against eight human cancer cell lines; A549, HCT116, PC3, A431, HePG2, HOS, PACA2 and BJ1. Compounds 7, 8 and 9 emerged IC50 values better than Doxorubicin as a reference drug. Compounds 7 and 8 showed IC50 = 44.4 and 22.4 μM respectively against PACA2 compared to Doxorubicin (IC50 = 52.1 μM). Compound 9 showed IC50 = 17.8, 12.4, and 17.6 μM against HCT116, HePG2, and HOS, respectively. qRT-PCR revealed the down-regulation of PALB2 in compounds 7 and 15 treated PACA2 cells. Also, the down-regulation of BRCA1 and BRCA2 was shown in compound 7 treated PC3 cells. As regard A549 cells, compound 8 decreased the expression level of EGFR and KRAS genes. While compounds 7 and 9 down-regulated TP53 and FASN in HCT116 cells. Molecular docking was done against Escherichia coli enoyl reductase and human Son of sevenless homolog 1 (SOS1) and the results showed the promising inhibition of the studied proteins

Design, synthesis, in vitro anticancer, molecular docking and SAR studies of new series of pyrrolo[2,3-d]pyrimidine derivatives

Abstract The current study involves the design and synthesis of a newly synthesized pyrrolo[2,3-d]pyrimidine derivatives to contain chlorine atoms in positions 4 and 6 and trichloromethyl group in position 2 using microwave technique as a new and robust approach for preparation of this type of pyrrolo[2,3-d]pyrimidine derivatives. The chemical structure of the synthesized pyrrolo[2,3-d]pyrimidine derivatives 3–19 was well-characterized using spectral and elemental analyses as well as single-crystal X-ray diffraction. All compounds were tested in vitro against seven selected human cancer cell lines, namely, MCF7, A549, HCT116, PC3, HePG2, PACA2 and BJ1 using MTT assay. It was found that compounds 14a, 16b and 18b were the most active toward MCF7 with IC50 (1.7, 5.7, and 3.4 μg/ml, respectively) relative to doxorubicin (Dox.) (26.1 μg/ml). Additionally, compound 17 exerted promising cytotoxic effects against HePG2 and PACA2 with IC50 (8.7 and 6.4 μg/ml, respectively) relative to Dox. (21.6 and 28.3 μg/ml, respectively). The molecular docking study confirmed our ELISA result which showed the promising binding affinities of compounds 14a and 17 against Bcl2 anti-apoptotic protein. At the gene expression level, P53, BAX, DR4 and DR5 were up-regulated, while Bcl2, Il-8, and CDK4 were down-regulated in 14a, 14b and 18b treated MCF7 cells. At the protein level, compound 14b increased the activity of Caspase 8 and BAX (18.263 and 14.25 pg/ml) relative to Dox. (3.99 and 4.92 pg/ml, respectively), while the activity of Bcl2 was greatly decreased in 14a treated MCF7 (2.4 pg/ml) compared with Dox. (14.37 pg/ml). Compounds 14a and 14b caused cell cycle arrest at the G1/S phase in MCF7. Compounds 16b and 18b induced the apoptotic death of MCF7 cells. In addition, the percentage of fragmented DNA was increased significantly in 14a treated MCF7 cells

Genetic structure of some candidate genes of repeat breeder syndrome in Egyptian buffaloes

Abstract Background This study aimed to explore the association between polymorphisms in three genes: leptin (LEP), leptin receptor (LEPR), and BMP4, and incidence of repeat breeding in Egyptian buffaloes. Methods DNA was extracted from 160 female buffaloes, involving 108 fertile and 52 repeat breeders. Genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Sequence analysis and alignment were performed by employing NCBI/BLAST/blastn suite, to identify SNPs among different patterns and alleles. We utilized PredictSNP software to predict the non-synonymous SNPs influences on protein function. Moreover, the conservation score of the amino acids within the target proteins was computed by ConSurf server. Results The genotyping results showed that LEP and BMP4 genes were monomorphic (CC, GG) in all tested fertile and repeat breeder buffaloes. Leptin gene sequencing showed a non-synonymous C73T SNP, replacing R to C at position 25 within the leptin polypeptide (position 4 in the mature form; R4C) which is a neutral mutation, not affecting function or structure of LEP protein. For LEPR, one synonymous SNP (T102C) and two non-synonymous SNPs (A106G and C146A), triggering V967A and G954C replacements, respectively in LEPR protein. Moreover, they are neutral mutations. Sequencing results of BMP4 showed HinfI restriction site indicate fixed GG genotype (CC genotype in the anti-sense strand) in all sequenced samples. No SNPs were observed within the amplified region. Conclusion Genotyping and sequencing results of the surveyed three genes revealed that there is no association between these genes mutations and the incidence of repeat breeding in Egyptian buffaloes