Normalization of miRNA qPCR high-throughput data: a comparison of methods

Low-density quantitative real-time PCR (qPCR) arrays are often used to profile expression patterns of microRNAs in various biological milieus. To achieve accurate analysis of expression of miRNAs, non-biological sources of variation in data should be removed through precise normalization of data. We have systematically compared the performance of 19 normalization methods on different subsets of a real miRNA qPCR dataset that covers 40 human tissues. After robustly modeling the mean squared error (MSE) in normalized data, we demonstrate lower variability between replicates is achieved using various methods not applied to high-throughput miRNA qPCR data yet. Normalization methods that use splines or wavelets smoothing to estimate and remove Cq dependent non-linearity between pairs of samples best reduced the MSE of differences in Cq values of replicate samples. These methods also retained between-group variability in different subsets of the dataset.     

Functional analyses of recombinant mouse hepcidin-1 in cell culture and animal model

Hepcidin is a peptide hormone that plays an important role in iron metabolism. We have produced a recombinant mouse hepcidin-1 by using baculovirus expression system. Its expression yield was 25 μg/ml when cell culture media were supplemented with a protease inhibitor cocktail. The recombinant mouse hepcidin-1 and synthetic human hepcidin-25 had similar effects on reducing ferroportin expression in J774A cell line and in peritoneal macrophages. However, synthetic human hepcidin-25 was more efficient than recombinant mouse hepcidin-1 in reducing iron concentration in blood circulation (p < 0.01).     

Carbon Nanotube-Encapsulated Drug Penetration Through the Cell Membrane: An Investigation Based on Steered Molecular Dynamics Simulation

Understanding the penetration mechanisms of carbon nanotube (CNTs)-encapsulated drugs through the phospholipid bilayer cell membrane is an important issue for the development of intracellular drug delivery systems. In the present work, steered molecular dynamics (SMD) simulation was used to explore the possibility of penetration of a polar drug, paclitaxel (PTX), encapsulated inside the CNT, through a dipalmitoylphosphatidylcholine bilayer membrane. The interactions between PTX and CNT and between PTX and the confined water molecules inside the CNT had a significant effect on the penetration process of PTX. The results reveal that the presence of a PTX molecule increases the magnitude of the pulling force. The effect of pulling velocity on the penetration mechanism was also investigated by a series of SMD simulations, and it is shown that the pulling velocity had a significant effect on pulling force and the interaction between lipid bilayer and drug molecule.     

Dating the “red beds” of the Eastern Moroccan High Plateaus: Evidence from late Late Cretaceous charophytes and dinosaur eggshells

A new locality in the poorly known “red beds” of Tendrara (High Plateaus, Morocco) has yielded four charophytes species (Feistiella anluensis, Lamprothamnium stipitatum, Peckisphaera portezueloensis, Platychara caudata) and dinosaur eggshells (Pseudomegaloolithus atlasi). These red beds, which overly the Cenomanian-Turonian marine deposits, generally assigned to “Senonian” based on geometric position, are directly dated by these fossils: the charophytes species and dinosaur oospecies association indicates a Campano-Maastrichtian or Maastrichtian age for these calm floodplain deposits.     

Involvement of the glutamate receptor AtGLR3.3 in plant defense signaling and resistance to Hyaloperonospora arabidopsidis

Like their animal counterparts, plant glutamate receptor‐like (GLR) homologs are intimately associated with Ca²⁺ influx through plasma membrane and participate in various physiological processes. In pathogen‐associated molecular patterns (PAMP)‐/elicitor‐mediated resistance, Ca²⁺ fluxes are necessary for activating downstream signaling events related to plant defense. In this study, oligogalacturonides (OGs), which are endogenous elicitors derived from cell wall degradation, were used to investigate the role of Arabidopsis GLRs in defense signaling. Pharmacological investigations indicated that GLRs are partly involved in free cytosolic [Ca²⁺] ([Ca²⁺]_cyt) variations, nitric oxide (NO) production, reactive oxygen species (ROS) production and expression of defense‐related genes by OGs. In addition, wild‐type Col‐0 plants treated with the glutamate‐receptor antagonist 6,7‐dinitriquinoxaline‐2,3‐dione (DNQX) had a compromised resistance to Botrytis cinerea and Hyaloperonospora arabidopsidis. Moreover, we provide genetic evidence that AtGLR3.3 is a key component of resistance against H. arabidopsidis. In addition, some OGs‐triggered immune events such as defense gene expression, NO and ROS production are also to different extents dependent on AtGLR3.3. Taken together, these data provide evidence for the involvement of GLRs in elicitor/pathogen‐mediated plant defense signaling pathways in Arabidopsis thaliana.     

Mesenchymal stem cell transfusion for desensitization of positive lymphocyte cross‐match before kidney transplantation: outcome of 3 cases

Objectives

Donor specific antibodies (DSA) and a positive cross‐match are contraindications for kidney transplantation. Trials of allograft transplantation across the HLA barrier have employed desensitization strategies, including the use of plasmapheresis, intravenous immunoglobulins, anti‐B‐cell monoclonal antibodies and splenectomy, associated with high‐intensity immunosuppressive regimens. Our case 1 report suffered from repeatedly positive lymphocyte cross match after 1st renal transplantation. Graft nephrectomy could not correct the state of sensitization. Splenectomy was done in a trial to get rid of the antibody producing clone. Furthermore plasmapheresis with low dose IVIG could not as well revert the state of sensitization for the patient.

Material and methods

About 50 millions donor specific MSCs were injected to the patient.

Results

MSCs transfusion proved to be the only procedure which could achieve successful desensitization before performing the second transplantation owing to their immunosuppressive properties.

Conclusion

This case indicates that DS‐MSCs is a potential option for anti‐HLA desensitization. In cases 2 and 3 IV DS‐MSCs transfusion was selected from the start as a successful line of treatment for pre renal transplantation desensitization to save other unnecessary lines of treatment that were tried in case 1.

     

The proportion of tetrahydrobiopterin deficiency and PAH gene deficiency variants among cases with hyperphenyalaninemia in Western Iran

BACKGROUND:

Defects either in phenylalanine hydroxylase (PheOH) or in the production and recycling of its cofactor (tetrahydrobiopterin [BH4]) are the causes of primary hyperphenylalaninemia (HPA). The aim of our study was to investigate the current status of different variants of HPA Kurdish patients in Kermanshah province, Iran.

MATERIALS AND METHODS:

From 33 cases enrolled in our study, 32 were identified as HPA patients. Reassessing of pre-treatment phenylalanine concentrations and the analysis of urinary pterins was done by high-performance liquid chromatography method.

RESULTS:

A total of 30 patients showed PAH deficiency and two patients were diagnosed with BH4 deficiency (BH4/HPA ratio = 6.25%). Both of these two BH4-deficient patients were assigned to severe variant of dihydropteridine reductase (DHPR) deficiency. More than 75% of patients with PAH deficiency classified as classic phenylketonuria (PKU) according their levels of pre-treatment phenylalanine concentrations.

CONCLUSION:

Based on the performed study, we think that the frequency of milder forms of PKU is higher than those was estimated before and/or our findings here. Furthermore, the frequency of DHPR deficiency seems to be relatively high in our province. Since the clinical symptoms of DHPR deficiency are confusingly similar to that of classic PKU and its prognosis are much worse than classical PKU and cannot be solely treated with the PKU regime, our pilot study support that it is crucial to set up screening for BH4 deficiency, along with PAH deficiency, among all HPA patients diagnosed with HPA.     

Gold-deferrioxamine nanometric interface for selective recognition of Fe(III) using square wave voltammetry and electrochemical impedance spectroscopy methods

Deferrioxamine, a bacterial hydroxamic siderophore having high binding affinity for Fe(III), is used in its immobilized form, as self-assembled monolayer on Au, for accumulation and recognition of Fe(III) from the solution phase. The accumulated Fe(III) is detected via both active mode based on faradaic reduction current of Fe(III), and inactive mode based on impedimetric effect of accumulated Fe(III) against redox reaction of a suitable probe. Appropriate electrochemical techniques, square wave voltammetry and electrochemical impedance spectroscopy, are used for the transduction of analytical signals obtained by this sensor. Then, the parameters influencing the sensor response are optimized. In the best conditions, a linear response, from 1.0×10(-10) to 1.0×10(-7)M Fe(III) in logarithmic scale with a detection limit of 2.0×10(-11)M, and mean relative standard deviation of 1.7% for n=4 is observed. The results show that the sensor can be used for determination of Fe(III) in the presence of various inorganic ions and biological species. Validity of the method and applicability of the sensor are successfully tested by determination of Fe(III) in various real samples including plant tissue (corn leaves), industrial alloy (Ferrotitanium), and pharmaceutical samples (Venofer(?) ampoule, Ironorm(?) capsule, and V.M. Protein(?) powder).     

Redescription and remarks on the species Minniza persica (Pseudoscorpiones: Olpiidae) from Iran

Minniza persica, which has been described briefly by Beier in 1951 on the basis of specimens from Hormozgan and Mazandaran provinces of Iran, was recently collected again from Hormozgan and Fars provinces and is described and illustrated here. The subspecies M. persica deminuta Beier is regarded as synonymous with the nominate subspecies.