Journal, genetic engineering & biotechnology最新文献

Background: Carbohydrates are known as the main natural products of life activities.

Results: Streptomyces rochie strain OF1 isolated from a mangrove tree produced exopolysaccharide S5 (EPSS5) (14.2 gl^-1) containing uronic acid 21.98% sulfate content of 11.65 mg/ml, and a viscosity of 1.35 mm²/s. while total hexose amine content was 24.72%. The high performance liquid chromatography (HPLC) analysis of mono sugars revealed that EPS was composed of manouronic acid, glucuronic acid, xylose, and fructose at a molar ratio of 1.0:0.5:1.0:2.0, respectively. It showed that the whole antioxidant activity was 92.06%. It showed antibacterial activity against Staphylococcus aureus, and E. coli, MRSA and Klebsiella pneumoniae. But, EPSS5 displayed low antifungal activity against Candida albicans. While no antifungal activity has been detected against Aspergillus niger. EPSS5 has antibiofilm action that is noticeable toward S. aureus with an inhibition ratio of biofilm up to 50%. Effect of EPS on serum levels of TNF-α and COX2 by 2 fold and 1.9 fold of EPS reduced serum levels of Tumor necrosis factor-α (TNF-α) by 38%, 12%, 49%, and Cyclooxygenase-2 (COX2) by 61%, 34%, and 62%, respectively. By affected of EPSS5 on arthritis in rats stimulated by carrageenan.

Conclusions: Administration of EPS ameliorated carrageen-induced elevation in inflammatory mediators; TNF-α/COX and suppressed the expressions of metalloproteinase 9 (MMP9) by 68%, 86%, and 75% correspondingly in comparison to the group of carrageenans. Then again, therapy involving a high dose only reduced MMP9 level by 57%, compared to free drug suggesting that EPSS5 is a good inhibitor of the MMP9, as it brought MMP9 back to normal levels via the signaling pathway.

Background: Breadfruit (Artocarpus spp.) is the main genus of Moraceae with multipurpose benefits, both ecologically and economically important, e.g., food ingredients, building materials, traditional medicine, and natural insecticides. However, most endemic Artocarpus have been threatened due to natural disasters and habitat degradation. The objective of our study was to determine the genetic diversity and relationships of endemic Artocarpus from South Borneo, Indonesia, using an internal transcribed spacer (ITS) region and leaf morphology.

Results: Morphologically, endemic Artocarpus endemic to South Borneo, Indonesia, has a different leaf shape, i.e., narrow-obovate to broad-elliptic, from simple to deeply dissected. Following the ITS region, this germplasm has a moderate level of nucleotide diversity (0.069). The phylogenetic analysis revealed Artocarpus into four (4) main clades, where the nearest is shown by the 'Puyian' (Artocarpus rigidus) and 'Binturung' (Artocarpus odoratissimus) at a coefficient divergence of 0.027, whereas the furthest by 'Kulur' (A. camansi) and 'Tiwadak' (A. integer) at a coefficient of 0.132.

Conclusion: In brief, although an endemic Artocarpus of South Borneo, Indonesia, has a moderate level of nucleotide diversity, this germplasm also shows a unique phylogenetic relationship. Thus, this information is urgent in supporting the future Artocarpus breeding and preservation programs, mainly to save this germplasm from being threatened.

Background and aims: NAFLD is one of the fast-growing health problems that affects up to 25% of people worldwide. Numerous miRNAs have been clarified as important regulators of liver pathophysiology, including NAFLD. Thus, we investigated the expression of the MiRNA-34a and MiRNA-192 as diagnostic markers for NAFLD.

Patients and methods: Blood samples were collected from NAFLD cases and healthy controls. The expression profile of both studied miRNAs was detected via real-time PCR analysis.

Results: The present study showed that both studied miRNAs were upregulated in NAFLD patients compared to controls. Interestingly, miRNA-34a and MiRNA-192 are upregulated in NAFLD patients with early fibrosis compared to controls [with a fold change of 4.02 ± 11.49 (P = 0.05) and 18.43 ± 47.8 (P = 0.017), respectively]. However, miRNA-34a is downregulated in NAFLD patients with advanced fibrosis compared to controls, with fold expression of 0.65 ± 1.17 (P = 0.831). The area under the receiver operating characteristics (AUROC) for miRNA-34a and miRNA-192 were 0.790 and 0.643, respectively; furthermore, the sensitivities and specificities were 76.7%, 100% for miRNA-34a and 63.3%, and 93.3% for miRNA-192 (P < 0.05). Additionally, MiRNA34a was positively correlated with hypertension and fasting blood sugar, and it also was negatively correlated with hemoglobin level and total leucocyte count (P < 0.05).

Conclusion: The results obtained indicated that both studied miRNAs could potentially be used as diagnostic biomarkers for the early stage of liver fibrosis in NAFLD cases. Also, miRNA-34a was positively correlated with metabolic disorders associated with NAFLD such as hypertension and diabetes. However, their expression showed no association with advanced fibrosis. Thus, larger cohorts are necessitated to certify the utility of serum MiRNA-34a and MiRNA-192 in monitoring the deterioration of NAFLD.

Background: There is always a need for a safe and efficient vaccine platform, especially when facing a pandemic such as COVID-19. Most of the SARS-CoV-2-based vaccines are based on the full spike protein, which is presented as a trimerized protein, and many viral vector vaccines express the spike protein into the host cells and do not display it on virus surfaces. However, the spike receptor-binding domain (RBD)-based vaccines are efficient and are currently under investigation and clinical trials.

Methodology: In this study, we are testing the efficacy of the RBD displayed on a baculovirus as a mean to formulate a safe and stable carrier to induce the immune system against SARS-CoV-2. Therefore, two pseudotyped baculoviruses were constructed to display the RBD, AcRBD-sfGFP-64, and AcRBD-sfGFP-V, using two different displaying strategies based on gp64 and VSV-G envelope glycoproteins, from Autographa californica multiple nucleopolyhedrovirus (AcMNPV) and vesicular stomatitis virus (VSV), respectively. BALB/C mice were immunized with the pseudotyped baculoviruses in a dose-optimized manner. Dot blot and Western blot were used to screen and validate the polyclonal antibodies' specificity to the SARS-CoV-2 RBD. A plaque reduction neutralization test (PRNT) was used to measure the sera neutralization capacity against a SARS-CoV-2 wild-type isolate from Egypt. ELISA was used to quantify certain cytokines for the assessment of the immune response.

Result: The outcome of our investigation showed that the monomeric RBD proteins were properly displayed on baculovirus and efficiently triggered the mouse immune system. The produced sera efficiently neutralized about 50% of SARS-CoV-2 in more than 100-fold serum dilution. The immunized mice showed a significant increase (p<0.01) in the levels of IL-2 and IFN-γ and a significant decrease (p<0.01) and (p<0.001) in the levels of IL-4 and IL-10, respectively, which suggest that AcRBD-sfGFP-64 and AcRBD-sfGFP-V induce Th1 cellular immune response.

Conclusion: The produced recombinant viruses can induce the immune response without adjuvant, which needs dose optimization and further stability tests. Neutralizing antibodies were induced without affecting the health of immunized mice. Th1 response can be attainable through the system, which is of great benefit in SARS CoV-2 infection and the system can be tested for future applications including vaccine development and polyclonal antibody production.

Background: Mating elicits significant changes in gene expression and leads to subsequent physiological and behavioural modifications in insects. The reproductive success of both sexes is contributed immensely by the male accessory gland (MAG) proteins that are transferred along with sperms to the female reproductive tract during mating where they facilitate several processes that modify the post-mating behaviour. The mating-responsive genes in the MAGs have been identified and reported in many insects but have not been well-characterized in the important agricultural pest Spodoptera litura. Here, we present RNA sequencing analysis to identify mating-responsive genes from the accessory glands of virgin males and males interrupted during mating.

Results: Overall, 91,744 unigenes were generated after clustering the assembled transcript sequences of both samples, while the total number of transcripts annotated was 48,708 based on sequence homology against the non-redundant (NR) database. Comparative transcriptomics analysis revealed 16,969 genes that were differentially expressed between the two groups, including 9814 up-regulated and 7155 down-regulated genes. Among the top 80 genes that were selected for heat map analysis, several prominent genes including odorant binding protein, cytochrome P450, heat shock proteins, juvenile hormone binding protein, carboxypeptidases and serine protease were differentially expressed.

Conclusions: The identified genes are known or predicted to promote several processes that modify the female post-mating behaviour. Future studies with the individual MAG protein or in combination will be required to recognize the precise mechanisms by which these proteins alter female physiology and reproductive behaviour. Thus, our study provides essential data to address fundamental questions about reproduction within and among insects and also paves way for further exploration of the functions of these proteins in female insects.

Background: In today's society, cancer has become a big concern. The most common cancers in women are breast cancer (BC), endometrial cancer (EC), ovarian cancer (OC), and cervical cancer (CC). CC is a type of cervix cancer that is the fourth most common cancer in women and the fourth major cause of death.

Results: This research uses a network approach to discover genetic connections, functional enrichment, pathways analysis, microRNAs transcription factors (miRNA-TF) co-regulatory network, gene-disease associations, and therapeutic targets for CC. Three datasets from the NCBI's GEO collection were considered for this investigation. Then, using a comparison approach between the datasets, 315 common DEGs were discovered. The PPI network was built using a variety of combinatorial statistical approaches and bioinformatics tools, and the PPI network was then utilized to identify hub genes and critical modules.

Conclusion: Furthermore, we discovered that CC has specific similar links with the progression of different tumors using Gene Ontology terminology and pathway analysis. Transcription factors-gene linkages, gene-disease correlations, and the miRNA-TF co-regulatory network were revealed to have functional enrichments. We believe the candidate drugs identified in this study could be effective for advanced CC treatment.

Background: Cisplatin resistance is one of the major contributors to the poor survival rate among head and neck cancer (HNC) patients. Focusing on the protein-protein interaction rather than a single protein could provide a better understanding of drug resistance. Thus, this study aimed to identify hub genes in a complex network of cisplatin resistance associated genes in HNC chemotherapy via a series of bioinformatic tools.

Methods: The genes involved in cisplatin resistance were retrieved from the NCBI gene database using "head and neck cancer" and "cisplatin resistance" as key words. The human genes retrieved were analyzed for their interactions and enriched using the STRING database. The interaction between KEGG pathways and genes was visualized in Cytoscape 3.7.2. Further, the hub gene was identified using the Cytohubba plugin of Cytoscape and validated using UALCAN and Human Protein Atlas database. Validated genes were investigated for the drug-gene interaction using the DGIbd database.

Results: Out of 137 genes obtained using key words, 133 were associated with cisplatin resistance in the human species. A total of 150 KEGG pathways, 82 cellular components, 123 molecular functions, and 1752 biological processes were modulated on enrichment analysis. Out of 37 hub genes, CCND1, AXL, CDKN2A, TERT, and EXH2 genes were found to have significant (p < 0.05) mRNA expression and effect on overall survival whereas protein expression was found to be positive for all the significant genes except TERT. Thus, they can be targeted with palbociclib, methotrexate, bortezomib and fluorouracil, sorafenib, dasatinib, carboplatin, paclitaxel, gemcitabine, imatinib, doxorubicin, and vorinostat.

Conclusion: As the pathogenesis of head and neck cancer is complex, targeting hub genes and associated pathways involved in cisplatin resistance could bring a milestone change in the drug discovery and management of drug resistance which might uplift overall survival among HNC patients.

Background: Cadmium is a non-essential, third largest heavy metal contaminant with long retention time that poses environmental hazards. It emanating majorly from industrial processes and phosphate fertilizers. Cadmium is effortlessly assimilated by plants and leads to yield loss. Henceforth, identification of mechanisms to attenuate the heavy metal toxicity in crops is beneficial for enhanced yields.

Results: Beneficial soil bacteria have been known to combat both biotic and abiotic stress, thereby promoting plant growth. Amongst them, Pseudomonas fluorescens has been shown to enhance abiotic stress resistance in umpteen crops for instance maize and groundnut. Here, we investigated the role of P. fluorescens in conferring cadmium stress resistance in Arabidopsis thaliana. In silico analysis of PCR2 gene and promoter revealed the role, in cadmium stress resistance of A. thaliana. Real-time expression analysis employing qRT-PCR ratified the upregulation of AtPCR2 transcript under cadmium stress up to 6 folds. Total leaf (50%), biomass (23%), chlorophyll content (chlorophyll-a and b 40%, and 36 %) silique number (50%), and other growth parameters significantly improved on bacterial treatment of the 2mM Cd-stressed plants.

Conclusion: Moreover, generated 35s-promoter driven AtPCR2 over-expressing transgenic lines that exhibited resistance to cadmium and other heavy metal stress. Taken together, a crucial interplay of P. fluorscens mediated enhanced expression of AtPCR2 significantly induced cadmium stress resistance in Arabidopsis plants.

Background: Thrombin is the most important enzyme in the hemostatic process by permitting rapid and localized coagulation in case of tissue damage. Camel thrombin is the natural and proper target enzyme for the previously purified camel tick salivary gland thrombin inhibitor.

Results: In this study, the camel thrombin was purified homogenously in a single affinity chromatographic step on the heparin-agarose affinity column with a specific activity of 3242 NIH units/mg proteins. On SDS-PAGE, the purified camel thrombin contained two forms, 37 kDa α-thrombin and 28 kDa β-thrombin, and the camel prothrombin was visualized as 72 kDa. The camel thrombin Km value was found out as 60 µM of N-(p-Tosyl)-Gly-Pro-Arg-p-nitroanilide acetate and displayed its optimum activity at pH 8.3. The PMSF was the most potent inhibitor of camel thrombin. Camel tick salivary gland thrombin inhibitor has two binding sites on camel thrombin and inhibited it competitively with Ki value of 0.45 µM.

Conclusions: The purified camel thrombin was found to be more susceptible toward the camel tick salivary gland thrombin inhibitor than bovine thrombin.

Background: Rg3-ginsenoside, a protopanaxadiol saponin, is a well-known adaptogen used for the prevention of cancer and inflammation. However, despite its distinct biological activity, the concentration of Rg3 in the total ginseng extract is insufficient for therapeutic applications. This study aims to convert PPD-class of major ginsenosides into a mixture of minor ginsenoside, to analyze its immune-regulatory role in macrophage cells.

Results: Using heat and organic acid treatment, three major ginsenosides, Rc, Rd, and Rb1, were converted into a mixture of minor ginsenosides, GRg3-mix [Rg3(S), Rg3(R), Rg5, and Rk1]. Purity and content analysis of the transformed compound were performed using thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC), compared with their standards. Preceding with the anti-inflammatory activity of GRg3-mix, lipopolysaccharide (LPS)-stimulated murine RAW264.7 macrophage cells were treated with various concentrations of GRg3-mix (6.25, 12.5, 25, and 50 μg/mL). The cell viability assay revealed that the level of cell proliferation was increased, while the nitric oxide (NO) assay showed that NO production decreased dose-dependently in activated RAW264.7 cells. The obtained results were compared to those of pure Rg3(S) ≥ 98% (6.25, 12.5, and 25 μg/mL). Preliminary analysis of the CCK-8 and NO assay demonstrated that GRg3-mix can be used as an anti-inflammatory mediator, but mRNA and protein expression levels were evaluated for further confirmation. The doses of GRg3-mix significantly suppressed the initially upregulated mRNA and protein expression of inflammation-related enzymes and cytokines, namely inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), nuclear transcription factor kappa B (NF-κB), tumor necrosis factor (TNF-α), and interleukins (IL-6 and IL1B), as measured by reverse transcription-polymerase chain reaction and western blotting.

Conclusions: Our pilot data confirmed that the mixture of minor ginsenosides, namely GRg3-mix, has high anti-inflammatory activity and has an easy production procedure.