Search results for: beta 2 toxin gene

Authors: Dalal Alghawas, Jetty Lee, Kaisa Poutanen, Hani El-Nezami

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Oat β-glucan a water soluble non starch linear polysaccharide has been approved as a cholesterol lowering agent by various food safety administrations and is commonly used to reduce the risk of heart disease. The molecular weight of oat β-glucan can vary depending on the extraction and fractionation methods. It is not clear whether the molecular weight has a significant impact at reducing the acceleration of atherosclerosis. The aim of this study was to investigate three different oat β-glucan fractionations on the development of atherosclerosis in vivo. With special focus on plaque stability and the intestinal barrier function. To test this, ApoE-/- female mice were fed a high fat diet supplemented with oat bran, high molecular weight (HMW) oat β-glucan fractionate and low molecular weight (LMW) oat β-glucan fractionate for 16 weeks. Atherosclerosis risk markers were measured in the plasma, heart and aortic tree. Plaque size was measured in the aortic root and aortic tree. ICAM-1, VCAM-1, E-Selectin, P-Selectin, protein levels were assessed from the aortic tree to determine plaque stability at 16 weeks. The expression of p22phox at the aortic root was evaluated to study the NADPH oxidase complex involved in nitric oxide bioavailability and vascular elasticity. The tight junction proteins E-cadherin and beta-catenin from western blot analyses were analysed as an intestinal barrier function test. Plasma LPS, intestinal D-lactate levels and hepatic FMO gene expression were carried out to confirm whether the compromised intestinal barrier lead to endotoxemia. The oat bran and HMW oat β-glucan diet groups were more effective than the LMW β-glucan diet group at reducing the plaque size and showed marked improvements in plaque stability. The intestinal barrier was compromised for all the experimental groups however the endotoxemia levels were higher in the LMW β-glucan diet group. The oat bran and HMW oat β-glucan diet groups were more effective at attenuating the development of atherosclerosis. Reasons for this could be due to the LMW oat β-glucan diet group’s low viscosity in the gut and the inability to block the reabsorption of cholesterol. Furthermore the low viscosity may allow more bacterial endotoxin translocation through the impaired intestinal barrier. In future food technologists should carefully consider how to incorporate LMW oat β-glucan as a health promoting food.

Keywords: Atherosclerosis, beta glucan, endotoxemia, intestinal barrier function

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Authors: Priya Arora, Ashutosh Mishra

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Gene identification, towards the pursuit of mutated genes, leading to Parkinson’s disease, puts forward a challenge towards proactive cure of the disorder itself. Computational analysis is an effective technique for exploring genes in the form of protein sequences, as the theoretical and manual analysis is infeasible. The limitations and effectiveness of a particular computational method are entirely dependent on the previous data that is available for disease identification. The article presents a sequence-based classification method for the identification of genes responsible for Parkinson’s disease. During the initiation phase, the physicochemical properties of amino acids transform protein sequences into a feature vector. The second phase of the method employs Jaccard distances to select negative genes from the candidate population. The third phase involves artificial neural networks for making final predictions. The proposed approach is compared with the state of art methods on the basis of F-measure. The results confirm and estimate the efficiency of the method.

Keywords: disease gene identification, Parkinson’s disease, physicochemical properties of amino acid, protein sequences

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Authors: Swati Gautam, Amita Jain, Shyampyari Jaiswar

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Objective: To elucidate the role of (ApaI&TaqI) VDR gene polymorphism in the pathogenesis of female genital tuberculosis (FGTB) cases. Background: Female genital TB represents about 15-20% of total extra-pulmonary TB (EPTB). Female subjects with vitamin D deficiency have been shown to be at higher risk of pulmonary TB as well as FGTB. In same context few functional polymorphism in vitamin D receptor (VDR) gene has been considered as an important genetic risk factor that modulate the development of FGTB. Therefore we aimed, to elucidate the role of (ApaI&TaqI) VDR gene polymorphism in the pathogenesis of FGTB. Study design: Case-Control study. Sample size: Cases (60) and Controls (60). Study site: Department of Obstetrics & Gynecology & Department of Microbiology, K.G.M.U. Lucknow, (UP). Inclusion criteria: Cases: Women with age group 20-35 years, premenstrual endometrial aspiration collected and included in the study, those were positive with acid-fast bacilli (AFB)/ TB-PCR/ LJ culture/ liquid culture. Controls: Women with age group 20-35 years having no history of ATT and all test negative for TB recruited as control. Exclusion criteria: -Women with endometriosis, polycystic ovaries (PCOD), positive on Chlamydia & gonorrhea, already on anti-tubercular therapy (ATT) excluded. Materials and Methods: Blood samples were collected in EDTA tubes from cases and controls stored at -20ºC. Genomic DNA extraction was carried out by salting-out method. Genotyping of VDR gene (ApaI&TaqI) polymorphism was performed by using single amplification refractory mutation system (ARMS) PCR technique. PCR products were analyzed by electrophoresis on 2% agarose gel. Statistical analysis was done by SPSS16.3 software & computing odds ratio (OR) with 95% CI. Results: Increased risk of female genital tuberculosis was observed in AA genotype (OR =1.1419-6.212 95% CI, P*<0.036) and A allele (OR =1.255-3.518, 95% CI, P* < 0.006) in FGTB as compared to controls. Moreover A allele was found more frequent in FGTB patients. No significant difference was observed in TaqI gene polymorphism of VDR gene. Conclusion: The ApaI polymorphism is significantly associated with etiology of FGTB and plays an important role as a genetic risk factor in FGTB women.

Keywords: ARMS, ATT, EPTB, FGTB, VDR

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Authors: Demiraw Bikila, Tadesse Lejisa, Yosef Tolcha, Chala Bashea, Mehari Meles Tigist Getahun Genet Ashebir, Wossene Habtu, Feyissa Challa, Ousman Mohammed, Melkitu Kassaw, Adisu Kebede, Letebrhan G. Egzeabher, Endalkachew Befekadu, Mistire Wolde, Aster Tsegaye

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Background: Even though several factors affect reference intervals (RIs), the company-derived values are currently in use in many laboratories worldwide. However, little or no data is available regarding serum protein RIs, mainly in resource-limited setting countries like Ethiopia. Objective: To establish a reference interval for serum protein electrophoresis of apparently healthy adults in Addis Ababa, Ethiopia. Method: A cross-sectional study was conducted on a total of 297 apparently healthy adults from April-October 2019 in four selected sub-cities (Akaki, Kirkos, Arada, Yeka) of Addis Ababa, Ethiopia. Laboratory analysis of collected samples was performed using Capillarys 2 Flex Piercing analyzer, while statistical analysis was done using SPSS version 23 and med-cal software. Mann-Whitney test was used to check Partitions. Non-parametric method of reference range establishment was performed as per CLSI guideline EP28A3C. Result: The established RIs were: Albumin 53.83-64.59%, 52.24-63.55%; Alpha-1 globulin 3.04-5.40%, 3.44-5.60%; Alpha-2 globulin 8.0-12.67%, 8.44-12.87%; and Beta-1 globulin 5.01-7.38%, 5.14-7.86%. Moreover, Albumin to globulin ratio was 1.16-1.8, 1.09-1.74 for males and females, respectively. The combined RIs for Beta-2 globulin and Gamma globulin were 2.54-4.90% and 12.40-21.66%, respectively. Conclusion: The established reference interval for serum protein fractions revealed gender-specific differences except for Beta-2 globulin and Gamma globulin.

Keywords: serum protein electrophoresis, reference interval, Addis Ababa, Ethiopia

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Authors: Abdullah Alaklabi, Ibrahim A. Arif, Sameera O. Bafeel, Ahmad H. Alfarhan, Anis Ahamed, Jacob Thomas, Mohammad A. Bakir

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Diospyros mespiliformis (Hochst. ex A.DC.; Ebenaceae) is a large deciduous medicinal plant. This plant species is currently listed as endangered in Saudi Arabia. Molecular identification of this plant species based on short sequence regions (571 and 664 bp) of plastid rbcL (ribulose-1, 5-biphosphate carboxylase) gene was investigated in this study. The endangered plant specimens were collected from Al-Baha, Saudi Arabia (GPS coordinate: 19.8543987, 41.3059349). Phylogenetic tree inferred from the rbcL gene sequences showed that this species is very closely related with D. brandisiana. The close relationship was also observed among D. bejaudii, D. Philippinensis and D. releyi (≥99.7% sequence homology). The partial rbcL gene sequence region (571 bp) that was amplified by rbcL primer-pair rbcLaF-rbcLaR failed to discriminate D. mespiliformis from the closely related plant species, D. brandisiana. In contrast, primer-pair rbcL1F-rbcL724R yielded longer amplicon, discriminated the species from D. brandisiana and demonstrated nucleotide variations in 3 different sites (645G>T; 663A>C; 710C>G). Although D. mespiliformis (EU980712) and D. brandisiana (EU980656) are very closely related species (99.4%); however, studied specimen showed 100% sequence homology with D. mespiliformis and 99.6% with D. brandisiana. The present findings showed that rbcL short sequence region (664 bp) of plastid rbcL gene, amplified by primer-pair rbcL1F-rbcL724R, can be used for authenticating samples of D. mespiliforformis and may provide help in authentic identification and management process of this medicinally valuable endangered plant species.

Keywords: Diospyros mespiliformis, endangered plant, identification partial rbcL

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Authors: Waqas Shafqat Chattha, Muhammad Iqbal, Amir Shakeel

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Transcriptional factors are proteins that play a vital role in regulating the transcription of target genes in different biological processes and are being widely studied in different plant species. In the current era of genomics, plant genomes sequencing has directed to the genome-wide identification, analyses and categorization of diverse transcription factor families and hence provide key insights into their structural as well as functional diversity. The DNA-binding with One Finger (DOF) proteins belongs to C2-C2-type zinc finger protein family. DOF proteins are plant-specific transcription factors implicated in diverse functions including seed maturation and germination, phytohormone signalling, light-mediated gene regulation, cotton-fiber elongation and responses of the plant to biotic as well as abiotic stresses. In this context, a genome-wide in-silico analysis of DOF TF family in diploid cotton species i.e. Gossypium raimondii has enabled us to identify 55 non-redundant genes encoding DOF proteins renamed as GrDofs (Gossypium raimondii Dof). Gene distribution studies have shown that all of the GrDof genes are unevenly distributed across 12 out of 13 G. raimondii chromosomes. The gene structure analysis illustrated that 34 out of 55 GrDof genes are intron-less while remaining 21 genes have a single intron. Protein sequence-based phylogenetic analysis of putative 55 GrDOFs has divided these proteins into 5 major groups with various paralogous gene pairs. Molecular evolutionary studies aided with the conserved domain as well as gene structure analysis suggested that segmental duplications were the principal contributors for the expansion of Dof genes in G. raimondii.

Keywords: diploid cotton , G. raimondii, phylogenetic analysis, transcription factor

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Authors: John Saaka

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The increasing world population poses a threat to food security. To meet current and future food demands, sweetpotato stand a good chance because of its recent food security roles. Concerted efforts are needed for both regional and local level varietal development. Heterosis exploiting breeding scheme (HEBS) is one of the options used to improve yield in some crop species and could be a good approach for sweetpotato improvement in Ghana by establishing heterotic gene pools within a population. To achieve this, 22 parental lines were collected from different sources and put in a full diallel arrangement. A total of 149 families, 20 individual cuttings per family, were taken to the field, including ‘checks’ and parental lines for experimentation in a 1m X 0.3m planting order according to the Westcott design. Results from this study led to the characterization of the selected parents into three main heterotic gene pools based on their suitability for use as male, female or both, respectively. This study serves as a baseline for further characterization of the rest of the germplasm in the Ghanaian sweetpotato breeding program.

Keywords: sweetpotato, heterosis, germplasm, food security

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Authors: John E. Muñoz, Jose F. Lopez, David S. Lopez

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Attention Deficit Hyperactivity Disorder (ADHD) is a disorder that affects 1 out of 5 Colombian children, converting into a real public health problem in the country. Conventional treatments such as medication and neuropsychological therapy have been proved to be insufficient in order to decrease high incidence levels of ADHD in the principal Colombian cities. This work demonstrates a design and development of a videogame that uses a brain computer interface not only to serve as an input device but also as a tool to monitor neurophysiologic signal. The video game named “The Harvest Challenge” puts a cultural scene of a Colombian coffee grower in its context, where a player can use his/her avatar in three mini games created in order to reinforce four fundamental aspects: i) waiting ability, ii) planning ability, iii) ability to follow instructions and iv) ability to achieve objectives. The details of this collaborative designing process of the multimedia tool according to the exact clinic necessities and the description of interaction proposals are presented through the mental stages of attention and relaxation. The final videogame is presented as a tool for sustained attention training in children with ADHD using as an action mechanism the neuromodulation of Beta and Theta waves through an electrode located in the central part of the front lobe of the brain. The processing of an electroencephalographic signal is produced automatically inside the videogame allowing to generate a report of the theta/beta ratio evolution - a biological marker, which has been demonstrated to be a sufficient measure to discriminate of children with deficit and without.

Keywords: BCI, neuromodulation, ADHD, videogame, neurofeedback, theta/beta ratio

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Authors: M. Omer Faruk, A. M. A. M. Zonaed Siddiki, M. Fazal Karim, Md. Masuduzzaman, S. Chowdhury, Md. Shafiqul Islam, M. Alamgir Hossain

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The dog tapeworms Echinococcus granulosus develop hydatid cysts in various organs in human and domestic animals worldwide including Bangladesh. The aim of this study was to identify and characterize the genotype of E. granulosus isolated from cattle using 12S rRNA and Cytochrome oxidase 1 (COX 1) genes. A total of 43 hydatid cyst samples were collected from 390 examined cattle samples derived from slaughterhouses. Among them, three cysts were fertile. Genomic DNA was extracted from germinal membrane and/or protoscoleces followed by PCR amplification of mitochondrial 12S rRNA and Cytochrome oxidase 1 gene fragments. The sequence data revealed existence of G1 (64.28%) and possible G3 (21.43%) genotypes for the first time in Bangladesh. The study indicates that common sheep strain G1 is the dominant subtype of E. granulosus in Chittagong region of Bangladesh. This will increase our understanding of the epidemiology of hydatidosis in the southern part of the country and will be useful to plan suitable control measures in the long run.

Keywords: Echinococcus granulosus, Cox1, 12S rRNA, molecular characterization, Bangladesh

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Authors: Thanusha D. Abeywickrama, Inoka C. Perera, Genji Kurisu

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Tuberculosis, considered being as the ninth leading cause of death worldwide, cause from a single infectious agent M. tuberculosis and the drug resistance nature of this bacterium is a continuing threat to the world. Therefore TB preventing treatment is expanding, where this study designed to analyze the regulatory mechanism of GntR transcriptional regulator gene Rv0792c, which lie between several genes codes for some hypothetical proteins, a monooxygenase and an oxidoreductase. The gene encoding Rv0792c was cloned into pET28a and expressed protein was purified to near homogeneity by Nickel affinity chromatography. It was previously reported that the protein binds within the intergenic region (BS region) between Rv0792c gene and monooxygenase (Rv0793). This resulted in binding of three protein molecules with the BS region suggesting tight control of monooxygenase as well as its own gene. Since monooxygenase plays a key role in metabolism, this gene may have a global regulatory role. The natural ligand for this regulator is still under investigation. In relation to the Rv0792 protein structure, a Circular Dichroism (CD) spectrum was carried out to determine its secondary structure elements. Percentage-wise, 17.4% Helix, 21.8% Antiparallel, 5.1% Parallel, 12.3% turn and 43.5% other were revealed from CD spectrum data under room temperature. Differential Scanning Calorimetry (DSC) was conducted to assess the thermal stability of Rv0792, which the melting temperature of protein is 57.2 ± 0.6 °C. The graph of heat capacity (Cp) versus temperature for the best fit was obtained for non-two-state model, which concludes the folding of Rv0792 protein occurs through stable intermediates. Peak area (∆HCal ) and Peak shape (∆HVant ) was calculated from the graph and ∆HCal / ∆HVant was close to 0.5, suggesting dimeric nature of the protein.

Keywords: CD spectrum, DSC analysis, GntR transcriptional regulator, protein structure

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Authors: Kadeejah Alsolami, Zainab Alrefay, Husaam Awad

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Obesity and vitamin D deficiency have both been related to cardiovascular disease. The present work aimed to investigate the possible protective effect of vitamin D on cardiac apoptosis in a rat model of dietary-induced obesity. Methods: 30 male Wistar rats included in this study. They were allocated into 4 groups: Control (n=5), animal were fed standard diet for 3 months: Control + vitamin D (VD) (n=5),animals were fed a standard diet with 400IU VD/kg for 3 months: hypercaloric diets group (n=10), animals were fed a high fat diet for 3 months: hypercaloric diet with VD group (n=10), animals were fed a high fat diet with 400IU VD/kg for 3 months. At the beginning of the experiment, the weight and length were measured to assess body mass index (BMI) and repeated every 45 days. Food intake and body weight were monitored throughout the study period. Then rats were sacrificed and heart tissues collected for Quantitative Real-time polymerase chain reaction (qRT-PCR). qRT-PCR used to detect different genetic markers of apoptosis (anti-apoptotic gene (BCL2), a pro-apoptotic gene(BAX), pro-apoptotic genes (FAS, FAS-L), tumour necrosis factor (TNF), mitogen-activated protein kinases (MAPK). Results: FAS and FAS-L gene expression were significantly upregulated in rats fed with high fat diet. And FAS-L gene expression was significantly upregulated in all groups on comparison with control. Whereas Bax gene expression was significantly downregulated in rats fed with high-fat diet supplied with vitamin D. TNF was significantly upregulated in rats fed with high-fat diet treated with vitamin D. MAPK was significantly upregulated in rats fed with high fat diet group, and in rats fed with high-fat diet supplied with vitamin D. Conclusion: The cardiac apoptotic pathways were more activated in rats fed with high-fat than lean rats. And vitamin D protect the heart from the cardiac mitochondrial-dependent apoptotic pathway.

Keywords: apoptosis, heart, obesity, Vitamin D

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Authors: Mariedel Autriz, Angel Lambio, Renato Vega, Severino Capitan, Rita Laude

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The study was conducted to associate genetic polymorphism of the leptin gene T3469C with reproductive performance in purebred sows. DNA were isolated from hair follicles of 29 Landrace and 24 Large White sows. Amplification of the leptin gene was done followed by Hinf1digestion to determine the base at the T3469C site. Electrophoresis of the digestion products revealed that there were 25 Landrace and 15 Large White sows with the TT genotype while there were 3 Landrace and 6 Large White TC. There was 1 CC for Landrace and 3 for Large White. Significant genotype associations were observed for total litter size born and total born alive. Significant breed differences, on the other hand, was observed for gestation length and average birth weight. Significant breed by genotype interaction was observed in litter size total born and litter size born alive.

Keywords: genetic polymorphism, leptin, swine, T3469C

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Authors: Malik SS, Masood N, Mubarik S, Khadim TM

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Introduction: Xeroderma pigmentosum group G (XPG) gene plays a crucial role in the correction of UV-induced DNA damage through nucleotide excision repair pathway. Single nucleotide polymorphisms in XPG gene have been reported to be associated with different cancers. Current case-control study was designed to evaluate the relationship between one of the most frequently found XPG (rs1047768 T>C) polymorphism and breast cancer risk. Methodology: A total of 200 individuals were screened for this polymorphism including 100 pathologically confirmed breast cancer cases and age-matched 100 controls. Genotyping was carried out using Tetra amplification-refractory mutation system (ARMS) PCR and results were confirmed by gel electrophoresis. Results: Conditional logistic regression analysis showed significant association between TC genotype (OR: 8.9, CI: 2.0 – 38.7) and increased breast cancer risk. Although homozygous CC genotype was more frequent in patients as compared to controls, but it was statistically non-significant (OR: 3.9, CI: 0.4 – 35.7). Conclusion: In conclusion, XPG (rs1047768 T>C) polymorphism may contribute towards increased risk of breast cancer but other polymorphisms may also be evaluated to elucidate their role in breast cancer.

Keywords: XPG, breast cancer, NER, ARMS-PCR

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Authors: Jasbir Singh, Devender Kumar, Davender Redhu, Surender Kumar, Vandana Bhardwaj

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Single Nucleotide polymorphism (SNP) of proteosomal gene complex is involved in the pathogenesis of hepatitis B Virus (HBV) infection. Some of such proteosomal gene complex are large multifunctional proteins (LMP) and antigen associated transporters that help in antigen presentation. Both are involved in intracellular processing and presentation of viral antigens in association with Major Histocompatability Complex (MHC) Class I molecules. A total of hundred each of hepatitis B virus infected and control samples from northern India were studied. Genomic DNA was extracted from all studied samples and PCR-RFLP method was used for genotyping at different positions of LMP genes. Genotypes at a given position were inferred from the pattern of bands and genotype frequencies and haplotype frequencies were also calculated. Homozygous SNP {A>C} was observed at codon 145 of LMP7 gene and having a protective role against HBV as there was statistically significant high distribution of this SNP among controls than cases. Heterozygous SNP {A>C} was observed at codon 145 of LMP7 gene and made individuals more susceptible to HBV infection as there was statistically significant high distribution of this SNP among cases than control. SNP {T>C} was observed at codon 60 of LMP2 gene but statistically significant differences were not observed among controls and cases. For codon 145 of LMP7 and codon 60 of LMP2 genes, four haplotypes were constructed. Haplotype I (LMP2 ‘C’ and LMP7 ‘A’) made individuals carrying it more susceptible to HBV infection as there was statistically significant high distribution of this haplotype among cases than control. Haplotype II (LMP2 ‘C’ and LMP7 ‘C’) made individuals carrying it more immune to HBV infection as there was statistically significant high distribution of this haplotype among control than cases. Thus it can be concluded that homozygous SNP {A>C} at codon 145 of LMP7 and Haplotype II (LMP2 ‘C’ and LMP7 ‘C’) has a protective role against HBV infection whereas heterozygous SNP {A>C} at codon 145 of LMP7 and Haplotype I (LMP2 ‘C’ and LMP7 ‘A’) made individuals more susceptible to HBV infection.

Keywords: Hepatitis B Virus, single nucleotide polymorphism, low molecular weight proteins, transporters associated with antigen presentation

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Authors: Ali Bayram, Burak Uz, Remzi Yiğiter

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The aim of the present study was to investigate the expression levels of homo sapiens nuclear factor of kappa light polypeptide gene enhancer in B-cells 1, transcript variant 1 (NFKB1*1) mRNA in the peripheral blood of patients with Parkinson to elucidate the role in the pathogenesis of Parkinson disease (PD). The study group comprised 50 patients with the diagnosis of PD who have applied to Gaziantep University Faculty of Medicine, and Department of Neurology. 50 healthy individuals without any neuro degenerative disease are included as controls. Ribonucleic acid (RNA) was obtained from blood samples of patient and control groups. Complementary deoxyribonucleic acid (cDNA) was obtained from RNA samples using reverse transcription polymerase chain reaction (RT-PCR) technique. The gene expression of NFKB1*1 in patient/control groups were observed to decrease significantly, and the differences between groups with the Mann-Whitney method within 95% confidence interval (p<0.05) were analyzed. This salient finding provide a clue for our hypothesis that reduced activity of NFKB1*1 gene might play a role, at least partly, in the pathophysiology of PD.

Keywords: Parkinson’s Disease, NFKB1, mRNA expression, RT-PCR

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Authors: Al-Mutary M., Alhimaidi A., Al-Ghadi M. Iwamoto D., Javed Ahmad. Abdulaziz A. Al-Khedhairy

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The aim of the present study was to evaluate the effect of ovine follicular fluid supplementation during IVM of sheep oocytes on the resumption of meiosis, glutathione (GSH) content and expression of Bax, Bcl-2, and HSPB1 genes. Sheep ovaries were collected from Riyadh slaughterhouse, KSA. Oocytes were aspirated from 3-6 mm follicles. Ovine oocytes were cultured in maturation medium with 0% (control), 10%, 20%, 40% of ovine follicular fluid for 24 h. Results indicated that the rate of oocyte maturation was significantly (P≤0.05) decreased in 40% OFF (36.87%) versus the control (61.3%), 10% OFF (63.95%) and 20% OFF (64.08%). Supplementation of 10% OFF to IVM medium induced an intra-oocyte GSH concentration significantly higher than that found in ovine oocytes cultured with 20% OFF and 40% OFF and similar to the GSH content in oocytes cultured without FF. Real time polymerase chain reaction analysis for gene expression revealed no differences in Bax, Bcl-2, HSPB1 genes between control and 10% OFF group, whereas they were strongly expressed in 20% OFF and 40% OFF (P < 0.05) when compared to the control and 10% OFF. In conclusion the addition of 10% OFF to the IVM culture of sheep oocytes is recommended to support cytoplasmic maturation and increase oocytes competence.

Keywords: IVM, oocyte maturation, gene expression, follicular fluid

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Authors: Yi-Hao Wong, Li-Li Chan, Chee-Onn Leong, Stephen Ambu, Joon-Wah Mak, Priyasashi Sahu

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Background: Acanthamoeba is a genus of amoebae which lives as a free-living in nature or as a human pathogen that causes severe brain and eye infections. Virulence potential of Acanthamoeba is not constant and can change with growth conditions. DNA methylation, an epigenetic process which adds methyl groups to DNA, is used by eukaryotic cells, including several human parasites to control their gene expression. We used qPCR, siRNA gene silencing, and RNA sequencing (RNA-Seq) to study DNA-methyltransferase gene family (DNMT) in order to indicate the possibility of its involvement in programming Acanthamoeba virulence potential. Methods: A virulence-attenuated Acanthamoeba isolate (designation: ATCC; original isolate: ATCC 50492) was subjected to mouse passages to restore its pathogenicity; a virulence-reactivated isolate (designation: AC/5) was generated. Several established factors associated with Acanthamoeba virulence phenotype were examined to confirm the succession of reactivation process. Differential gene expression of DNMT between ATCC and AC/5 isolates was performed by qPCR. Silencing on DNMT gene expression in AC/5 isolate was achieved by siRNA duplex. Total RNAs extracted from ATCC, AC/5, and siRNA-treated (designation: si-146) were subjected to RNA-Seq for comparative transcriptomic analysis in order to identify the genome-wide effect of DNMT in regulating Acanthamoeba gene expression. qPCR was performed to validate the RNA-Seq results. Results: Physiological and cytophatic assays demonstrated an increased in virulence potential of AC/5 isolate after mouse passages. DNMT gene expression was significantly higher in AC/5 compared to ATCC isolate (p ≤ 0.01) by qPCR. si-146 duplex reduced DNMT gene expression in AC/5 isolate by 30%. Comparative transcriptome analysis identified the differentially expressed genes, with 3768 genes in AC/5 vs ATCC isolate; 2102 genes in si-146 vs AC/5 isolate and 3422 genes in si-146 vs ATCC isolate, respectively (fold-change of ≥ 2 or ≤ 0.5, p-value adjusted (padj) < 0.05). Of these, 840 and 1262 genes were upregulated and downregulated, respectively, in si-146 vs AC/5 isolate. Eukaryotic orthologous group (KOG) assignments revealed a higher percentage of downregulated gene expression in si-146 compared to AC/5 isolate, were related to posttranslational modification, signal transduction and energy production. Gene Ontology (GO) terms for those downregulated genes shown were associated with transport activity, oxidation-reduction process, and metabolic process. Among these downregulated genes were putative genes encoded for heat shock proteins, transporters, ubiquitin-related proteins, proteins for vesicular trafficking (small GTPases), and oxidoreductases. Functional analysis of similar predicted proteins had been described in other parasitic protozoa for their survival and pathogenicity. Decreased expression of these genes in si146-treated isolate may account in part for Acanthamoeba reduced pathogenicity. qPCR on 6 selected genes upregulated in AC/5 compared to ATCC isolate corroborated the RNA sequencing findings, indicating a good concordance between these two analyses. Conclusion: To the best of our knowledge, this study represents the first genome-wide analysis of DNA methylation and its effects on gene expression in Acanthamoeba spp. The present data indicate that DNA methylation has substantial effect on global gene expression, allowing further dissection of the genome-wide effects of DNA-methyltransferase gene in regulating Acanthamoeba pathogenicity.

Keywords: Acanthamoeba, DNA methylation, RNA sequencing, virulence

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Authors: Andrew Thayer, Courtney Rondeau, Paraskevi Papadopoulou

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The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) gene-editing technologies have generated considerable discussion about the applications and ethics of their use. However, no consistent guidelines for using CRISPR technologies have been developed -nor common legislation passed related to gene editing, especially as it is connected to genetically modified organisms (GMOs) in the European Union. The recent announcement that the first babies with CRISPR-edited genes were born, along with new studies exploring CRISPR’s applications in treating thalassemia, sickle-cell anemia, cancer, and certain forms of blindness, have demonstrated that the technology is developing faster than the policies needed to control it. Therefore, it can be seen that a reasonable and coherent regulatory framework for the use of CRISPR in human somatic and germline cells is necessary to ensure the ethical use of the technology in future years. The European Union serves as a unique region of interconnected countries without a standard set of regulations or legislation for CRISPR gene-editing. We posit that the EU would serve as a suitable model in comparing the legislations of its affiliated countries in order to understand the practicality and effectiveness of adopting majority-approved practices. Additionally, we present a proposed set of guidelines which could serve as a basis in developing a consistent regulatory framework for the EU countries to implement but also act as a good example for other countries to adhere to. Finally, an additional, multidimensional framework of smart solutions is proposed with which all stakeholders are engaged to become better-informed citizens.

Keywords: CRISPR, ethics, regulatory framework, European legislation

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Authors: Karla Cautivo, Thomas Riley, Daniel Knight

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Clostridium difficile infection (CDI) is the most common cause of infectious diarrhea in hospitalised humans. C. difficile colonises the gastrointestinal tract, causes disease in a variety of animal species and can persist as a spore in diverse environments. Genetic overlap between C. difficile strains from human, animal and environmental sources suggests CDI has a zoonotic or foodborne aetiology. In Australia, C. difficile PCR ribotype RT014 (MLST clade 1) and several ST11 (MLST clade 5) RTs are found commonly in livestock. The high prevalence and diversity of ST11 strains in Australian production animals indicates Australia might be the ancestral home for this lineage. This project describes for the first time the ecology of C. difficile in Australian native animals, providing insights into the prevalence, molecular epidemiology and evolution of C. difficile in this unique environment and a possible role in CDI in humans and animals in Australia. Faecal samples were collected from wild/captive reptiles (n=37), mammals (n=104) and birds (n=102) in Western Australia in 2020/21. Anaerobic enrichment culture was performed, and C. difficile isolates were characterised by PCR ribotyping and toxin gene profiling. Seventy isolates of C. difficile were recovered (prevalence of C. difficile in faecal samples 28%, n=68/243); 27 unique RTs were identified, 5 were novel. The prevalence of C. difficile was similar for reptiles and mammals, 46% (n=17/37) and 43%(n=45/104), respectively, but significantly lower in birds (7.8%, n=8/102; p<0.00001 for both reptiles and mammals). Of the 57 isolates available for typing, RT237 (clade 5) and RT002 (clade 2) were the most prevalent, 15.8% (n=9/57) and 14% (n=8/57), respectively. The high prevalence of C. difficile in reptiles and mammals, particularly clade 5 strains, supported by previous studies of C. difficile in Australian soils, suggest that Australia might be the ancestral home of MLST clade 5.

Keywords: Clostridium difficile, zoonosis, molecular epidemiology, ecology and evolution

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Authors: Diana G. Ramirez-Wong, Marius Ramirez, Regina Sanchez-Leija, Adriana Rugerio, R. Araceli Mauricio-Sanchez, Martin A. Hernandez-Landaverde, Arturo Carranza, John A. Pojman, Josue D. Mota-Morales, Gabriel Luna-Barcenas

Abstract:

Chitin films were prepared using alpha-chitin from shrimp shells as raw material and a simple method of precipitation-evaporation. Choline chloride: urea Deep Eutectic Solvent (DES) was used to disperse chitin and compared against hexafluoroisopropanol (HFIP). A careful analysis of the chemical and crystalline structure was followed along the synthesis of the films, revealing crystalline-phase transitions. The full conversion of alpha- to beta-, or alpha- to gamma-chitin structure were detected by XRD and NMR on the films. The synthesis of highly crystalline monophasic gamma-chitin films was achieved using a DES; whereas HFIP helps to promote the beta-phase. These results are encouraging to continue in the study of DES as good processing media to control the final properties of chitin based materials.

Keywords: chitin, deep eutectic solvent, polymorph, phase transformation

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Authors: Samina Jam Nazeer Ahmad, Samia Yasin, Ijaz Ahmad, Muhammad Tahir, Jam Nazeer Ahmad

Abstract:

Phytoplasmas are phloem-inhibited plant pathogenic bacteria that are transferred by insect vectors. Among biotic factors, Phytoplasma infection induces abnormality influencing the physiology as well as morphology of plants. In 16Sr-IX group phytoplasma-infected brassica compestris, flower abnormalities have been associated with changes in the expression of floral development genes. To determine whether methylation was involved in down-regulation of flower development, the process of DNA methylation and Demethylation was investigated as a possible mechanism for regulation of floral gene expression in phytoplasma infected Brassica transmitted by Orosious orientalis vector by using RT-PCR, MSRE-PCR, Southern blotting, Bisulfite Sequencing, etc. Transcriptional expression of methylated genes was found to be globally down-regulated in plants infected with phytoplasma, but not severely in those infested by insect vectors and variation in expression was found in genes involved in methylation. These results also showed that genes particularly orthologous to Arabidopsis APETALA3 involved in petal formation and flower development was down-regulated severely in phytoplasma-infected brassica and with the fact that phytoplasma and insect induce variation in developmental gene expression. The DNA methylation status of flower developmental gene in phytoplasma infected plants with 5-azacytidine restored gene expression strongly suggesting that DNA methylation was involved in down-regulation of floral development genes in phytoplasma infected brassica.

Keywords: genes expression, phytoplasma, DNA methylation, flower development

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Authors: Shalu Jain, Anju Bansal, Anup Kumar, Sunita Saxena

Abstract:

Objectives: The novel TMPRSS2:ERG gene fusion is a common somatic event in prostate cancer that in some studies is linked with a more aggressive disease phenotype. Thus, this study aims to determine whether clinical variables are associated with the presence of TMPRSS2:ERG-fusion gene transcript in Indian patients of prostate cancer. Methods: We evaluated the clinical variables with presence and absence of TMPRSS2:ERG gene fusion in prostate cancer and BPH association of clinical patients. Patients referred for prostate biopsy because of abnormal DRE or/and elevated sPSA were enrolled for this prospective clinical study. TMPRSS2:ERG mRNA copies in samples were quantified using a Taqman chemistry by real time PCR assay in prostate biopsy samples (N=42). The T2:ERG assay detects the gene fusion mRNA isoform TMPRSS2 exon1 to ERG exon4. Results: Histopathology report has confirmed 25 cases as prostate cancer adenocarcinoma (PCa) and 17 patients as benign prostate hyperplasia (BPH). Out of 25 PCa cases, 16 (64%) were T2: ERG fusion positive. All 17 BPH controls were fusion negative. The T2:ERG fusion transcript was exclusively specific for prostate cancer as no case of BPH was detected having T2:ERG fusion, showing 100% specificity. The positive predictive value of fusion marker for prostate cancer is thus 100% and the negative predictive value is 65.3%. The T2:ERG fusion marker is significantly associated with clinical variables like no. of positive cores in prostate biopsy, Gleason score, serum PSA, perineural invasion, perivascular invasion and periprostatic fat involvement. Conclusions: Prostate cancer is a heterogeneous disease that may be defined by molecular subtypes such as the TMPRSS2:ERG fusion. In the present prospective study, the T2:ERG quantitative assay demonstrated high specificity for predicting biopsy outcome; sensitivity was similar to the prevalence of T2:ERG gene fusions in prostate tumors. These data suggest that further improvement in diagnostic accuracy could be achieved using a nomogram that combines T2:ERG with other markers and risk factors for prostate cancer.

Keywords: prostate cancer, genetic rearrangement, TMPRSS2:ERG fusion, clinical variables

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Authors: Zerrin Orbak, Busra Demir

Abstract:

Osteopetrosis is a rare genetic disease characterized by impaired bone resorption and increased bone sclerosis. Clinical presentation is very different in osteopetrosis. It can be asymptomatic or can be seen with typical symptoms. Here, a case of osteopetrosis was presented when evaluated for nystagmus. She was 10 months old. Parents were second-degree relatives. On physical examination, pigeon chest deformity and horizontal nystagmus were observed. There was a failure of thrive but no fracture. The cardiovascular examination was normal. Cranial, vertebral and long bone roentgenograms revealed characteristic deformities of osteopetrosis and diffuse sclerosis. The diagnosis was confirmed by genetic testing. A Homozygous mutation was detected in the TNFRSF11A gene (c.508A>G p.(Arg170Gly)). RANKL is encoded by the tumor necrosis factor ligand superfamily member 11 (TNFSF11) gene, and the binding to its receptor RANK, encoded by the TNFRSF11A gene, determines the activation of the downstream pathway that drives osteoclast differentiation and activation (51). The complete absence of osteoclasts is the key feature of the osteoclast-poor form of osteopetrosis (46). Patients are characterized by the absence of TRAP-positive osteoclasts in bone biopsies. The osteoclast-poor subtype of osteopetrosis caused by mutations in TNFSF11 gene is ultra-rare in humans. Clinical presentation is usually severe, with onset in early infancy or in fetal life. But here, a case was presented with horizontal nystagmus. A case presented with horizontal nystagmus, which was evaluated by neurology and diagnosed incidentally, was shared.

Keywords: osteopetrosis, nystagmus, bone, osteoclast-poor

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Authors: Sumeyra Sevim, Gulsum Gizem Topal, Mercan Merve Tengilimoglu-Metin, Banu Sancak, Mevlude Kizil

Abstract:

Aflatoxin M1 (AFM1) represents mutagenic, carcinogenic, hepatotoxic and immunosuppressive properties, and shows adverse effect on human health. Recently the use of probiotics are focused on AFM1 detoxification because of the fact that probiotic strains have a binding ability to AFM1. Moreover, inulin is a prebiotic to improve the ability of probiotic bacteria. Therefore, the aim of the study is to investigate the effect of inulin on AFM1 binding ability of some probiotic bacteria. Yoghurt samples were manufactured by using skim milk powder artificially contaminated with AFM1 at concentration 100 pg/ml. Different samples were prepared for the study as: first sample consists of yoghurt starter bacteria (L. bulgaricus and S. thermophilus), the second sample consists of starter and L. plantarum, starter and B. bifidum ATCC were added to the third sample, starter and B. animalis ATCC 27672 were added to the forth sample, and the fifth sample is a binary culture consisted of starter and B. bifidum and B. animalis. Moreover, the same work groups were prepared with inulin (4%). The samples were incubated at 42°C for 4 hours, then stored for three different time interval (1,5 and 10 days). The toxin was measured by the ELISA. When inulin was added to work groups, there was significant change on AFM1 binding ability at least one sample in all groups except the one with L. plantarum (p<0.05). The highest levels of AFM1 binding ability (68.7%) in samples with inulin were found in the group which B. bifidum was added, whereas the lowest levels of AFM1 binding ability (44.4%) in samples with inulin was found in the fifth sample. The most impressive effect of inulin was found on B.bifidum. In this study, it was obtained that there was a significant effect of storage on AFM1 binding ability in the all groups with inulin except the one with L. plantarum (p<0.05). Consequently, results show that AFM1 detoxification by probiotics have a potential application to reduce toxin concentrations in yoghurt. Besides, inulin has different effects on AFM1 binding ability of each probiotic bacteria strain.

Keywords: aflatoxin M1, inulin, probiotics, storage

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Authors: M. A. Akulov, V. O. Zaharov, A. A. Tomskij

Abstract:

Introduction: Scapulohumeral periarthrosis, resulting as a reaction to mechanical injury of shoulder tendons and muscles, is associated with high incidence of temporal and permanent disability. There is a strong need for investigation of treatment of that patient group. Severe pain leads to limitation of movements range, which result in secondary alterations of joint capsule and ligamentous apparatus. Muscle tension and edema, swelling of fascial and fibrous structures result in nerve and vascular compression in intramuscular and osseo-muscular-fibrous spaces. Botulinum toxin injection leads to decrease of muscle tone, increase of movements range and associated pain alleviation. Study aim: Optimization of rehabilitation process in scapolohumeral periarthrosis using Xeomin. Patients and methods: 40 patients aged 37-56 years with scapulohumeral periarthrosis were evaluated. Patients were divided into two groups according to treatment regimen. The first (main) group included 21 patients, receiving intramuscular Xeomin 150-200 U in the area of brachio-scapular joint and trigger points (inducing motion range limitation and pain). Treatment procedures were combined with physical therapy and osteopathic procedures. The second (control) group included 19 patients, receiving conventional physical therapy and osteopathic procedures. The evaluation and efficacy comparison was carried out using McGill pain questionnaire, Clinical Global Impression scale (CGI), and patient-reported increase of brachio-scapular joint movement range and pain decrease at 1, 3 and 6 months of treatment. Results. The study demonstrated a significant improvement in the main group after one month of treatment, which persisted during months of treatment. At baseline, rank pain index on McGill pain questionnaire was 18,4±4,9 and 17,8±5,1 in the main and control group, respectively (p > 0,05). At 1 month of treatment we observed a significant decrease of pain syndrome (no pain or modest pain) and increase of movement range in angular degrees in the main group (р < 0,05). In the control group significant improvements were observed only on the 3 month of treatment (р < 0,05), but at 6 months of treatment the improvement in pain syndrome and motion range in brachio-scapular joint was significantly smaller, than in the main group. Rank pain index on McGill pain scale was 5,2±1,8 in the main group compared to 12,0±2,6 in the control group (р < 0,05). At 6 months of treatment patients in the first group reported a significant/highly significant improvement of general health on CGI, whereas in the second group most patients reported a minimal improvement. We observed a sustained and persistent improvement of motion range in brachio-scapular joint in the main group. Conclusion: Xeomin injections as a part of rehabilitation process in scapulohumeral periarthrosis lead to reduced time and increased quality of rehabilitation.

Keywords: botulinum toxin, rehabilitation, scapulohumeral periarthrosis

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Authors: Bindu I. Somarajan, Viney Gupta, Gagandeep Kaur Walia, Jasbir Kaur, Sunil Kumar, Shikha Gupta, Abadh K. Chaurasia, Dinesh Gupa, Abhinav Kaushik, Aditi Mehta, Vipin Gupta, Arundhati Sharma

Abstract:

Background: Juvenile onset primary open angle glaucoma (JOAG), affects individuals under the age of 40 years. Studies on a few families of JOAG, that led to the discovery of the Myocilin gene, reported the disease to have an autosomal dominant pattern of inheritance. However, sporadic forms of JOAG been seen to be more common in some populations. Most pathological homozygous mutations in the CYP1B1 gene associated with JOAG have been seen among sporadic cases. Given the higher prevalence of sporadic JOAG cases in our population, we aimed to look for common mutations E229K and R368H, the two most common variants in the CYP1B1 gene associated with glaucoma. Objective: To determine the frequency and evaluate genotype phenotype correlation of CYP1B1 E229K and R368H mutations in a cohort of 120 sporadic Juvenile open angle glaucoma patients.Methods: Unrelated JOAG patients whose first degree relatives had been examined and found to be unaffected were included in the study. The patients and their parents were screened for E229K and R368H mutations. The phenotypic characteristics were compared between probands with and with out these mutations by SPSS v16. Results: Out of 120 JOAG patients included in the study, the E229K mutation was seen in 9 probands (7.5%) and R368H in 7 (5.8%). The average age of onset of the disease (p=0.3) and the highest untreated IOP (p=0.4) among those carrying mutations was not significantly different from those who did not have these mutations. The proportion of probands with angle dysgenesis among those with E229K and R368H mutations was 70% (11 out of 16) in comparison to 65% (67 out of 104) of those who did not harbour these mutations (p=0.56). Similarly the probands with moderate to high myopia among those with E229K and R368H mutations was 20% (3 out of 16) in comparison to 18% (18 out of 104) of those who did not harbour these mutations(p=0.59). Conclusion: The frequency of E229K and R368H mutations of the CYP1B1 gene is low even among sporadic JOAG patients. Moreover there is no clinical correlation between the presence of these mutations and disease severity

Keywords: CYP1B1, gene, IOP, JOAG, mutation

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Authors: C. Kridtayopas, W. Danvilai, P. Sopannarath, A. Kayan, W. Loongyai

Abstract:

Both Betong chicken (KU Line) and Thai Native chickens were the high quality of the meat and low carcass fat compared to broiler chickens. The objective of this study was to determine the growth performance, carcass characteristic, meat quality and association of polymorphism in the ApoVLDL-II gene with fat accumulation in the female broiler, Thai Native and Betong (KU line) chickens at 4-14 weeks. The chickens were used and reared under the same environment and management (100 chicks per breed). The results showed that body weight (BW) of broiler chickens was significantly higher than Thai Native and Betong (KU line) chickens (P < 0.01) through all the experiment. At 4-8 weeks of age, feed conversion ratio (FCR) of broiler chickens was significantly better than Thai Native and Betong (KU line) chickens (P < 0.01), then increased at week 8-14. The percentage of breast, abdominal fat and subcutaneous fat of broiler chickens was significantly greater than Thai Native and Betong (KU line) chickens (P < 0.01). However, Thai Native chickens showed the highest percentage of liver (P < 0.01) when compared to other breeds. In addition, the percentage of wing of Thai Native and Betong (KU line) chickens were significantly (P < 0.01) higher than broiler chickens. Meat quality was also determined and found that, pH of breast meat left from slaughter 45 minutes (pH45) and 24 hours (pH24) of broiler was significantly higher than Thai Native and Betong (KU line) (P < 0.01) whereas the percentage of drip loss, thawing loss, cooking loss and shear force was not significantly different between breeds. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique was used to genotype the polymorphism in the ApoVLDL-II gene in the broiler, Thai Native and Betong (KU line) chickens. The results found that, the polymorphism in the ApoVLDL-II gene at VLDL6 loci was not associated with fat accumulation in those studied population.

Keywords: ApoVLDL-II gene, Betong (KU line) chickens, broiler chickens, carcass characteristic, growth performance, meat quality, Thai native chickens

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Authors: Vivian A. Panes, Raymond John S. Rebong, Miel Q. Diaz

Abstract:

Moringa oleifera Lam. is known mainly for its high nutritional value and medicinal properties contributing to its popular reputation as a 'miracle plant' in the tropical climates where it usually grows. The main objective of this study is to discover the genes and gene products involved in abiotic stress-induced activity that may impact the M. oleifera Lam. mature seeds as well as their corresponding functions. In this study, RNA-sequencing and de novo transcriptome assembly were performed using two assemblers, Trinity and Oases, which produced 177,417 and 120,818 contigs respectively. These transcripts were then subjected to various bioinformatics tools such as Blast2GO, UniProt, KEGG, and COG for gene annotation and the analysis of relevant metabolic pathways. Furthermore, FPKM analysis was performed to identify gene expression levels. The sequences were filtered according to the 'response to stress' GO term since this study dealt with stress response. Clustered Orthologous Groups (COG) showed that the highest frequencies of stress response gene functions were those of cytoskeleton which make up approximately 14% and 23% of stress-related sequences under Trinity and Oases respectively, recombination, repair and replication at 11% and 14% respectively, carbohydrate transport and metabolism at 23% and 9% respectively and defense mechanisms 16% and 12% respectively. KEGG pathway analysis determined the most abundant stress-response genes in the phenylpropanoid biosynthesis at counts of 187 and 166 pathways for Oases and Trinity respectively, purine metabolism at 123 and 230 pathways, and biosynthesis of antibiotics at 105 and 102. Unique and cumulative GO term counts revealed that majority of the stress response genes belonged to the category of cellular response to stress at cumulative counts of 1,487 to 2,187 for Oases and Trinity respectively, defense response at 754 and 1,255, and response to heat at 213 and 208, response to water deprivation at 229 and 228, and oxidative stress at 508 and 488. Lastly, FPKM was used to determine the levels of expression of each stress response gene. The most upregulated gene encodes for thiamine thiazole synthase chloroplastic-like enzyme which plays a significant role in DNA damage tolerance. Data analysis implies that M. oleifera stress response genes are directed towards the effects of climate change more than other stresses indicating the potential of M. oleifera for cultivation in harsh environments because it is resistant to climate change, pathogens, and foreign invaders.

Keywords: stress response, genes, Moringa oleifera, transcriptomics

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Authors: Felicia Segeth, Florian G. Klein, Lea Berger, Andreas Kolk, Per S. Holm

Abstract:

The entry of oncolytic viruses (OVs) into clinical application opens groundbreaking changes in current and future treatment regimens. However, despite their potent anti-cancer activity in vitro, clinical studies revealed limitations of OVs as monotherapy. The same applies to CDK 4/6 inhibitors (CDK4/6i) targeting cell cycle as well as bromodomain and extra-terminal domain inhibitors (BETi) targeting gene expression. In this study, the anti-tumoral effect of XVir-N-31, an YB-1 dependent oncolytic adenovirus, was evaluated in combination with Ribociclib, a CDK4/6i, and JQ1, a BETi. The head and neck squamous cell carcinoma (HNSCC) cell lines Fadu, SAS, and Cal-33 were used. DNA replication and gene expression of XVir-N-31 was measured by RT-qPCR, protein expression by western blotting, and cell lysis by SRB assays. Treatment with CDK4/6i and BETi increased viral gene expression, viral DNA replication, and viral particle formation. The data show that the combination of oncolytic adenovirus XVir-N-31 with CDK4/6i & BETi acts highly synergistic in cancer cell lysis. Furthermore, additional molecular analyses on this subject demonstrate that the positive transcription elongation factor P-TEFb plays a decisive role in this regard, indicating an influence of the combinational therapy on gene transcription control. The combination of CDK4/6i & BETi and XVir-N-31 is an attractive strategy to achieve substantial cancer cell killing and is highly suitable for clinical testing.

Keywords: adenovirus, BET, CDK4/6, HNSCC, P-TEFb, YB-1

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Authors: Marawan Abd Elbaset Mohamed, Hanan A. Ogaly, Rehab F. Abdel-Rahman, Ahmed-Farid O.A., Marwa S. Khattab, Reham M. Abd-Elsalam

Abstract:

Liver fibrosis is a severe worldwide health concern due to various chronic liver disorders. Hepatic encephalopathy (HE) is one of its most common complications affecting liver and brain cognitive function. Beta-Carotene (B-Car) is an organic, strongly colored red-orange pigment abundant in fungi, plants, and fruits. The study attempted to know B-Car neuroprotective potential against thioacetamide (TAA)-induced neurotoxicity and cognitive decline in HE in rats. Hepatic encephalopathy was induced by TAA (100 mg/kg, i.p.) three times per week for two weeks. B-Car was given orally (10 or 20 mg/kg) daily for two weeks after TAA injections. Organ body weight ratio, Serum transaminase activities, liver’s antioxidant parameters, ammonia, and liver histopathology were assessed. Also, the brain’s mitogen-activated protein kinase (MAPK), nuclear factor kappa B (NF-κB), antioxidant parameters, adenosine triphosphate (ATP), adenosine monophosphate (AMP), norepinephrine (NE), dopamine (DA), serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA) cAMP response element-binding protein (CREB) expression and B-cell lymphoma 2 (Bcl-2) expression were measured. The brain’s cognitive functions (Spontaneous locomotor activity, Rotarod performance test, Object recognition test) were assessed. B-Car prevented alteration of the brain’s cognitive function in a dose-dependent manner. The histopathological outcomes supported these biochemical evidences. Based on these results, it could be established that B-Car could be assigned to treat the brain’s neurotoxicity consequences of HE via downregualtion of MAPK/NF-κB signaling pathways.

Keywords: beta-carotene, liver injury, MAPK, NF-κB, rat, thioacetamide

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