Inhibitor of PI3K/Akt Signaling Pathway Small Molecule Promotes Motor Neuron Differentiation of Human Endometrial Stem Cells Cultured on Electrospun Biocomposite Polycaprolactone/Collagen Scaffolds

Small molecules as useful chemical tools can affect cell differentiation and even change cell fate. It is demonstrated that LY294002, a small molecule inhibitor of phosphatidylinositol 3-kinase (PI3K)/Akt signal pathway, can inhibit proliferation and promote neuronal differentiation of mesenchymal stem cells (MSCs). The purpose of this study was to investigate the differentiation effect of Ly294002 small molecule on the human endometrial stem cells (hEnSCs) into motor neuron-like cells on polycaprolactone (PCL)/collagen scaffolds. hEnSCs were cultured in a neurogenic inductive medium containing 1 μM LY294002 on the surface of PCL/collagen electrospun fibrous scaffolds. Cell attachment and viability of cells on scaffolds were characterized by scanning electron microscope (SEM) and 3-(4,5-dimethylthiazoyl-2-yl)2,5-diphenyltetrazolium bromide (MTT) assay. The expression of neuron-specific markers was assayed by real-time PCR and immunocytochemistry analysis after 15 days post induction. Results showed that attachment and differentiation of hEnSCs into motor neuron-like cells on the scaffolds with Ly294002 small molecule were higher than that of the cells on tissue culture plates as control group. In conclusion, PCL/collagen electrospun scaffolds with Ly294002 have potential for being used in neural tissue engineering because of its bioactive and three-dimensional structure which enhances viability and differentiation of hEnSCs into neurons through inhibition of the PI3K/Akt pathway. Thus, manipulation of this pathway by small molecules can enhance neural differentiation.     

Association between microRNA-21, microRNA-150, and micro-RNA-451 expression and clinical outcome of patients with acute lymphoblastic leukemia

Background

Acute lymphoblastic leukemia (ALL) occurs owing to the defective maturation, increased proliferation, and lack of differentiation of lymphoid cells. Evaluation of the expression levels of microRNAs (miRNAs) could help in the prognosis and improve the clinical outcome of ALL patients. Given the role of miR-21, miR-150, and miR-451 as oncogenes and tumor suppressors in lymphocytes, this study explored the relation between the expression levels of these miRNAs and the clinical outcomes of ALL patients.

Methods

cDNA synthesis and RT-PCR were performed for peripheral blood samples from 41 patients with ALL, as well as for U937 and Jurkat cell lines to examine the expression of miR-451, miR-150, and miR-21 after miRNA purification. We also performed an epidemiological analysis in which Mann–Whitney and Chi-square tests were used to investigate the relationship between the expression of miRNAs and other clinical and laboratory data. Binary logistic regression models were used to estimate the odds ratio in univariate and multivariate analyses for clinical outcomes.

Results

miR-21 and miR-150 expression was found to be decreased, while miR-451 expression showed no difference compared to the control group. There was a significant relationship between miR-451 expression and hemoglobin (Hb) levels, as well as between miR-150 expression and clinical outcomes of ALL patients.

Conclusion

Increased expression of miR-451 decreased the Hb levels; reduced expression of miR-150 was associated with increased relapse rate in patients. Age, increased WBC, and decreased Hb levels were associated with increased relapse rates in ALL patients. Therefore, miR-150 could be used as a biomarker to determine the clinical outcome of ALL patients.

     

A method for monitoring of oxygen saturation changes in brain tissue using diffuse reflectance spectroscopy

Continuous measurement of local brain oxygen saturation (SO₂) can be used to monitor the status of brain trauma patients in the neurocritical care unit. Currently, micro‐oxygen‐electrodes are considered as the “gold standard” in measuring cerebral oxygen pressure (pO₂), which is closely related to SO₂ through the oxygen dissociation curve (ODC) of hemoglobin, but with the drawback of slow in response time. The present study suggests estimation of SO₂ in brain tissue using diffuse reflectance spectroscopy (DRS) for finding an analytical relation between measured spectra and the SO₂ for different blood concentrations. The P₃ diffusion approximation is used to generate a set of spectra simulating brain tissue for various levels of blood concentrations in order to estimate SO₂. The algorithm is evaluated on optical phantoms mimicking white brain matter (blood volume of 0.5–2%) where pO₂ and temperature is controlled and on clinical data collected during brain surgery. The suggested method is capable of estimating the blood fraction and oxygen saturation changes from the spectroscopic signal and the hemoglobin absorption profile.

     

Establishment and characterization of Caspian horse fibroblast cell bank in Iran

Caspian horse, a rare horse breed found in 1965 by Louise Firouz in northern Iran, is a small horse which is reported to be in danger of extinction in its original homeland. There seems to be a great need to prevent extinction of this valuable horse. In this study, 51 fibroblast cell lines from Caspian horse ear marginal tissue were successfully established by sampling 60 horses using primary explant technique. Cells were authenticated and growth curve was plotted. According to results obtained, population doubling time (PDT) was calculated 23 ± 0.5 h for all cell lines. Multiplex polymerase chain reaction (multiplex PCR) revealed that cell lines had no cross-contamination with other species. Bacteria, fungi, and mycoplasma contamination were checked using standard methods such as PCR, direct culture, and Hoechst staining. In addition to providing a valuable source for genomic, postgenomic, and somatic cloning researches, the established cell lines would preserve Caspian horse genetic resources. It will also create an accessible database for researchers.     

A novel chemiluminescence sensor for the determination of indomethacin based on sulfur and nitrogen co‐doped carbon quantum dot–KMnO4 reaction

We report on a simple and sensitive sulfur and nitrogen co‐doped carbon quantum dot (S,N‐CQD)‐based chemiluminescence (CL) sensor for the determination of indomethacin. S,N‐CQDs were prepared by a hydrothermal method and characterized by fluorescence spectra, Fourier transform infrared spectroscopy and transmission electron microscopy. To obtain the best CL system for determination of indomethacin, the reaction of S,N‐CQDs with some common oxidants was studied. Among the tested systems, the S,N‐CQD–KMnO₄ reaction showed the highest sensitivity for the detection of indomethacin. Under optimum conditions, the calibration plot was linear over a concentration range of 0.1–1.5 mg L^?1, with a limit of detection (3σ) of 65 μg L^?1. The method was applied to the determination of indomethacin in environmental and biological samples with satisfactory results.     

A multi-spectroscopic and molecular docking approach to investigate the interaction of antiviral drug oseltamivir with ct-DNA

The possible interaction between the antiviral drug oseltamivir and calf thymus DNA at physiological pH was studied by spectrophotometry, competitive spectrofluorimetry, differential pulse voltammogram (DPV), circular dichroism spectroscopy (CD), viscosity measurements, salt effect, and computational studies. Intercalation of oseltamivir between the base pairs of DNA was shown by a sharp increase in specific viscosity of DNA and a decrease of the peak current and a positive shift in differential pulse voltammogram. Competitive fluorescence experiments were performed using neutral red (NR) as a probe for the intercalation binding mode. The studies showed that oseltamivir is able to release the NR.     

Experimental and computational studies on the effects of valganciclovir as an antiviral drug on calf thymus DNA

DNA-binding properties of an antiviral drug, valganciclovir (valcyte) was studied by using emission, absorption, circular dichroism, viscosity, differential pulse voltammetry, fluorescence techniques, and computational studies. The drug bound to calf thymus DNA (ct-DNA) in a groove-binding mode. The calculated binding constant of UV-vis, K_a, is comparable to groove-binding drugs. Competitive fluorimetric studies with Hoechst 33258 showed that valcyte could displace the DNA-bound Hoechst 33258. The drug could not displace intercalated methylene blue from DNA double helix. Furthermore, the induced detectable changes in the CD spectrum of ct-DNA as well as changes in its viscosity confirm the groove-binding mode. In addition, an integrated molecular docking was employed to further investigate the binding interactions between valcyte and calf thymus DNA.     

Antibacterial activity of Punica granatum L. and Areca nut (P.A) combined extracts against some food born pathogenic bacteria

The antibacterial effects of combined extracts of Punica granatum L. and Areca nut (P.A) against resistant bacteria, a gram-positive bacterium, Staphylococcus aureus and three gram-negative bacteria, Escherichia coli, Salmonella, and Enterobacter aerogenes, in individual and biofilm forms was studied. Antibacterial activity was studied using disk diffusion method, microbroth dilution, and microtiter plate methods. Given the disc diffusion test (Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC)), the extracts had inhibitory effects on the individual forms of bacteria. However, the ethanolic extract had greater effectiveness than the methanolic extract. Generally, ethanol efficiently extracts flavonoids and their glycosides, catechol and tannins. This fact due to the ethanol polarity that is equal 0.654. The results indicated that the ability of extracts in inhibiting the formation of biofilms, destruction of biofilms, and prevention of metabolic activity of bacteria had a direct relationship with concentration and the highest inhibitory was seen on Staphylococcus aureus (98.98%), Staphylococcus aureus (94.98%), and Enterobacter aerogenes (88.55%). Based on the results, the P.A. combined extract can be used as an alternative combination with the ability to inhibit antibiotic-resistant bacteria in single and biofilm forms.     

Enhancement of CRISPR/Cas12a trans-cleavage activity using hairpin DNA reporters

The RNA programmed non-specific (trans) nuclease activity of CRISPR-Cas Type V and VI systems has opened a new era in the field of nucleic acid-based detection. Here, we report on the enhancement of trans-cleavage activity of Cas12a enzymes using hairpin DNA sequences as FRET-based reporters. We discover faster rate of trans-cleavage activity of Cas12a due to its improved affinity (K_m) for hairpin DNA structures, and provide mechanistic insights of our findings through Molecular Dynamics simulations. Using hairpin DNA probes we significantly enhance FRET-based signal transduction compared to the widely used linear single stranded DNA reporters. Our signal transduction enables faster detection of clinically relevant double stranded DNA targets with improved sensitivity and specificity either in the presence or in the absence of an upstream pre-amplification step.     

Posters. P11 Positive predictive values for high and low‐grade cytological abnormalities as surrogates for specificity and sensitivity

Introduction Positive predictive value (PPV), measuring the percentage of moderate dyskaryosis or worse confirmed as CIN2 or worse, is used as a measure of accuracy in cervical screening. However, it relates more to specificity than sensitivity because the denominator includes false positives rather than false negatives. Low values reflect over‐reporting of high‐grade dyskaryosis but high values may reflect under‐reporting. Sensitivity is impossible to measure from correlation of cytology with outcome because women with negative cytology are rarely referred for colposcopy. Rates of CIN3 resulting from referrals for low‐grade cytology may be used as a surrogate for sensitivity, as high values may reflect under‐reporting (ref). Study design Outcome of colposcopy referrals was monitored during a period of 4 years, using a fail‐safe database. Results PPV at Guy's & St Thomas rose from 54% in 1998/1999 to 69% in 2001/2002. The former was below the NHSCSP recommended range. During the same period of time CIN1 rates for moderate dyskaryosis fell from 37% to 24%, reflecting the main source of discrepancy. While specificity increased (as reflected by increasing PPV) sensitivity remained constant in that CIN3 rates for mild dyskaryosis and borderline remained below 6%: average rates in England have fallen over the last 3 years and were 7.4% in 2000/2001 (ref). CIN2 rates for mild dyskaryosis also remained constant at 11% to 12%. Conclusion Correlation of biopsy results with high‐ and low‐grade cytological abnormalities is a useful method of monitoring accuracy of cytology reporting, and can be used to measure over‐ and under‐reporting as surrogates for specificity and sensitivity.