DNA repair in human promyelocytic cell line, HL-60.

The human promyelocytic cell line, HL-60, shows large changes in endogenous poly(ADP-ribose) and in nuclear ADP-ribosyl transferase activity (ADPRT) during its induced myelocytic differentiation. DNA strand-breaks are an essential activator for this enzyme; and transient DNA strand breaks occur during the myelocytic differentiation of HL-60 cells. We have tested the hypothesis that these post-mitotic, terminally differentiating cells are less efficient in DNA repair, and specifically in DNA strand rejoining, than their proliferating precursor cells. We have found that this hypothesis is not tenable. We observe that there is no detectable reduction in the efficiency of DNA excision repair after exposure to either dimethyl sulphate or gamma-irradiation in HL-60 cells induced to differentiate by dimethyl sulphoxide. Moreover, the efficient excision repair of either dimethyl sulphate or gamma-irradiation induced lesions, both in the differentiated and undifferentiated HL-60 cells, is blocked by the inhibition of ADPRT activity.     

The next step of neurogenesis in the context of Alzheimer’s disease

Molecular Biology Reports - Among different pathological mechanisms, neuronal loss and neurogenesis impairment in the hippocampus play important roles in cognitive decline in Alzheimer’s...     

Conceptual Method of Temperature Sensation in Bionic Hand by Extraordinary Perceptual Phenomenon

Lack of temperature sensation of myoelectric prosthetic hand limits the daily activities of amputees.To this end,a non-invasive temperature sensation method is proposed to train amputees to sense temperature with psychophysical sensory substitution.In this study,22 healthy participants took part besides 5 amputee participants.The duration time of the study was 31 days with five test steps according to the Leitner technique.An adjustable temperature mug and a Peltier were used to change the temperature of the water/phantom digits to induce temperature to participants.Also,to isolate the surround-ings and show colors,a Virtual Reality(VR)glass was employed.The statistical results conducted are based on the response of participants with questionnaire method.Using Chi-square tests,it is concluded that participants answer the experiment significantly correctly using the Leitner technique(P value＜0.05).Also,by applying the"Repeated Measures ANOVA",it is noticed that the time of numbness felt by participants had significant(P value＜0.001)difference.Participants could remember lowest and highest temperatures significantly better than other temperatures(P value＜0.001);furthermore,the well-trained amputee participant practically using the prosthesis with 72.58％could identify object's temperature with only once time experimenting the color temperature.     

Heat-Induced Conformational Unfolding of Fatty Acid-Free and Fatted Camel Serum Albumin: A Comparative Study

Albumin is a multifunctional non-glycosylated, negatively charged plasma protein, with extraordinary ligand-binding and transport properties, antioxidant functions, and enzymatic activities. Physiologically, albumin transports free fatty acids in plasma and contributes in maintaining colloid osmotic pressure. Recent progresses in using albumin as a versatile protein carrier for drug targeting and for improving the pharmacokinetic profile of peptide or protein-based drugs, increased the attempts for improving albumin stability. Studying the thermal stability of camel albumin may provide us not only new clues for designing recombinant albumins, but also molecular insights on camel physiology. This study aims to determine the thermal stability of camel albumin. Fatted camel serum albumin (FCSA) was purified from blood via combination of Cohn’s method and anion-exchange chromatography. Activated charcoal treatment was used to obtain defatted camel serum albumin (CSA). Fluorescence spectroscopy and differential scanning calorimetry (DSC) were used to study thermal denaturation of this protein. The set of fluorescence spectra were deconvoluted using the convex constraint analysis method (CCA). The results from deconvolution of fluorescence spectroscopy and DSC showed three and two components for CSA and FCSA, respectively. The bimodal DSC transition can be attributed to a crevice between domains I and II and formation of two independent thermodynamic domains. The crevice formation can be prevented by fatty acid binding between domains I and II. The calculated values of ?H _v/?H _cal, approximately 0.4 for CSA and near 1 for FCSA, confirmed the presence of at least one intermediate in thermal unfolding of CSA and the absence of the intermediate for FCSA. The obtained midpoint transition temperature (T _m) of FCSA was about 20 °C higher than that of CSA. Such enormous stabilizing effect may be attributed to the fact that fatty acid serves as glue which preserves different domains beside each other and prevents formation of the mentioned intermediate.     

Dissolved oxygen concentration regulates human hepatic organoid formation from pluripotent stem cells in a fully controlled bioreactor

Developing technologies for scalable production of human organoids has gained increased attention for “organoid medicine” and drug discovery. We developed a scalable and integrated differentiation process for generation of hepatic organoid from human pluripotent stem cells (hPSCs) in a fully controlled stirred tank bioreactor with 150 ml working volume by application of physiological oxygen concentrations in different liver tissue zones. We found that the 20–40% dissolved oxygen concentration [DO] (corresponded to 30–60 mmHg pO₂ within the liver tissue) significantly influences the process outcome via regulating the differentiation fate of hPSC aggregates by enhancing mesoderm induction. Regulation of the [DO] at 30% DO resulted in efficient generation of human fetal-like hepatic organoids that had a uniform size distribution and were comprised of red blood cells and functional hepatocytes, which exhibited improved liver-specific marker gene expressions, key liver metabolic functions, and, more important, higher inducible cytochrome P450 activity compared to the other trials. These hepatic organoids were successfully engrafted in an acute liver injury mouse model and produced albumin after implantation. These results demonstrated the significant impact of the dissolved oxygen concentration on hPSC hepatic differentiation fate and differentiation efficacy that should be considered ascritical translational aspect of established scalable liver organoid generation protocols for potential clinical and drug discovery applications.     

Human umbilical cord mesenchymal stem cell-derived extracellular vesicles: A novel therapeutic paradigm

Mesenchymal stem cells (MSCs) have been revealed to hold great potential for the development of new treatment approaches for various diseases. However, the clinical use of these cells is limited due to their tumorigenic effects. The therapeutic benefits of MSCs are largely dependent on paracrine factors including extracellular vesicles (EVs). EVs are nano-sized bilayer membrane structures containing lipids, microRNAs and proteins which play key roles in cell-to-cell communications. Because of their lower immunogenicity, tumorigenicity, and easier management, EVs have emerged as a new promising alternative to whole-cell therapy. Therefore, this paper reviews current preclinical studies on the use of EVs derived from human umbilical cord MSCs (hucMSCs) as a therapeutic approach in treatment of several diseases including neurological, cardiovascular, liver, kidney, and bone diseases as well as the cutaneous wound, inflammatory bowel disease, cancers, infertility, and other disorders.     

Construction and eukaryotic expression of recombinant large hepatitis delta antigen

Background:

Hepatitis delta virus (HDV) is a subviral human pathogen that exploits host RNA editing activity to produce two essential forms of the sole viral protein, hepatitis delta antigen (HDAg). Editing at the amber/W site of HDV antigenomic RNA leads to the production of the large form (L-HDAg), which is required for RNA packaging.

Methods:

In this study, PCR-based site-directed mutagenesis by the overlap extension method was used to create the point mutation converting the small-HDAg (S-HDAg) stop codon to a tryptophan codon through three stages.

Results:

Sequencing confirmed the desirable mutation and integrity of the L-HDAg open reading frame. The amplicon was ligated into pcDNA3.1 and transfected to Huh7 and HEK 293 cell lines. Western blot analysis using enhanced chemiluminescence confirmed L-HDAg expression. The recombinant L-HDAg localized within the nuclei of cells as determined by immunofluorescence and confocal microscopy.

Conclusion:

Because L-HDAg requires extensive post-translational modifications, the recombinant protein expressed in a mammalian system might be fully functional and applicable as a tool in HDV molecular studies, as well as in future vaccine research.Key Words: Hepatitis Delta Virus, L-HDAg, SOEing-PCR