The effect of decitabine on the expression and methylation of the PPP1CA,BTG2, and PTEN in association with changes in miR-125b,miR-17, and miR-181b in NALM6 cell line

Precursor B-cell acute lymphoblastic leukemia (B-ALL) is the most prevalent pediatric cancer. DNA methylation and changes in the microRNAs (miRNAs) expression are known to be important causes of B-ALL. Decitabine as a DNA methyltransferase inhibitor agent is able to induce hypomethylation in several tumor suppressor genes. Much evidence has proven BTG2, PPP1CA, and PTEN act as tumor suppressor genes in many malignancies. In this case control study, the messenger RNA (mRNA) expression of PPP1CA, BTG2, and PTEN genes using quantitative real-time polymerase chain reaction (rRT-PCR) in Nalm6 cell line and five patients suffer from ALL with mean age 5.6 years were determined in compare with seven normal healthy donors age and sex matched. qRT-PCR analysis revealed that the expression levels of PPP1CA, BTG2, and PTEN genes were significantly decreased in Nalm6 ([FC] = 0.46, [FC] = 0.046, [FC] = 0.54) and according to the Methylation-specific PCR (MSP) analysis, these genes were hypermethylated in Nalm6. In next step, the effects of decitabine treatment on the methylation and expression of these genes in association with changes in miR-125b, miR-17, and miR-181b expression levels were evaluated in optimal concentration 2.5 µM of decitabine. Our data showed that decitabine is able to restore the expression levels of aforementioned genes and downregulate expression levels of oncomiRs; including miR-125b, miR-17, and miR-181b in Nalm6 cell line. Therefore, it seems that decitabine can be used as a potential drug for the first line treatment of patients with B-ALL, but further in vivo investigation is necessary.     

Contrbtion to the biology of Zagros tooth-carp, <Emphasis Type="Italic">Aphanius vladykovi</Emphasis> (Cyprinodontidae) in central Iran

We studied some aspects of the biology of the Zagros tooth-carp, Aphanius vladykovi, an endemic and poorly known species from Chahar-Mahall-va-Bakhtiari province in central Iran, by regular monthly collections and direct observation in aquaria over a full year. We collected individuals from Modar-Dokhtar spring of Gandoman region; we preserved some in formalin and transferred some alive to aquaria. The stomach contents consisted mostly of freshwater crustaceans, suggesting a carnivorous habit. The eggs had an average diameter of 1mm and the average absolute and relative fecundity was 415 and 110, respectively. The gonadosomatic ratio and ovary condition suggested that the reproductive season of the species was between late March and June with a peak in early April. The species is euryhaline and eurythermal and prefers neutral to basic waters. It is usually found in well-oxygenated waters, but is tolerant of hypoxia as well.     

Structural studies of SNARE complex and its interaction with complexin by molecular dynamics simulation

Since neurotransmitter releasing into the synaptic space delivers electrical signals from presynaptic neural cell to the postsynaptic cell, neurotransmitter secretion must be much orchestrated. Crowded intracellular vesicles involving neurotransmitters present a question of the how secretory vesicles fuse onto the plasma membrane in a fast synchronized fashion. Complexin is one of the most experimentally studied proteins that regulate assembly of fusogenic four‐helix SNARE complex to synchronized neurotransmitter secretion. We used MD simulation to investigate the interaction of complexin with the neural SNARE complex in detail. Our results show that the SNARE complex interacts with the complexin central helix by forming salt bridges and hydrogen bonds. Complexin also can interact with the Q‐SNARE complex instead of synaptobrevin to decrease the Q‐SNARE flexibility. The complexin alpha‐accessory helix and the C‐terminal region of synaptobrevin can interact with the same region of syntaxin. Although the alpha‐accessory helix aids the tight binding of the central helix to the SNARE complex, its proximity with synaptobrevin causes the destabilization of syntaxin and Sn1 helices. This study suggests that the alpha‐accessory helix of complexin can be an inhibiting factor for membrane fusion by competing with synaptobrevin for binding to the Q‐SNARE complex. © 2010 Wiley Periodicals, Inc. Biopolymers 93: 560–570, 2010. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com     

Thermally induced changes in the structure and activity of yeast hexokinase B

Yeast hexokinase has been poorly characterized in regard with its stability. In the present study, various spectroscopic techniques were employed to investigate thermal stability of the monomeric form of yeast hexokinase B (YHB). The enzyme underwent a conformational transition with a T(m) of about 41.9 degrees C. The structural transition proved to be significantly reversible below 55 degrees C and irreversible at higher temperatures. Thermoinactivation studies revealed that enzymatic activity diminished significantly at high temperatures, with greater loss of activity observed above 55 degrees C. Release of ammonia upon deamidation of YHB obeyed a similar temperature-dependence pattern. Dynamic light scattering and size exclusion-HPLC indicated formation of stable aggregates. Taking various findings on the influence of osmolytes and chaperone-like agents on YHB thermal denaturation together, it is proposed that the purely conformational transition of YHB is reversible, and irreversibility is due to aggregation, as a major cause. Deamidation of a critical Asn or Gln residue(s) may also play an important role.     

Sarcocystis and Its Complications in Camels (Camelus dromedarius) of Eastern Provinces of Iran

The prevalence of Sarcocystis spp. was investigated by gross and histopathological examinations in 250 camels (Camelus dromedarius) slaughtered from 2002 to 2005 in the Mashhad Slaughterhouse, eastern Iran. Samples were taken from the diaphragm, heart, tongue, esophagus and masseter muscles for histopathological studies. No macroscopic sarcocysts were found in the samples at gross inspection. Sarcocysts were detected in 209 of 250 (83.6%) examined camels at histopathological level. The infection rate of the esophagus, heart, masseter muscles, diaphragm, and tongue was 58.8%, 48.0%, 46.8%, 41.6%, and 28.0%, respectively. There was no significant difference in the rate of infection between male (85.8%) and female (81.0%) camels. The tissue response to vital cysts was minimal; however, reaction to the degenerating cysts was severe and caused tissue damages resulting in hyperemia, hemorrhages, mononuclear cell infiltration, necrotic changes, and fibrosis. The wild and domestic carnivores especially dogs may be the final hosts of Sarcocystis spp. in this area.     

Rapid fertility decline in Iran: analysis of intermediate variables

The remarkable decline in fertility in Iran, which saw the total fertility rate fall from 7 children per woman in 1986 to 2 in 2000, has received only limited analysis in the demographic literature. Using the 2000 Iran Demographic and Health Survey and Bongaarts' age-specific fertility model, this paper examines the role of the major proximate determinants of fertility in bringing about the rapid decrease in fertility in Iran. The analysis indicates that contraception had the largest effect on fertility, accounting for 61% of the reduction in fertility from its theoretical maximum. The fertility-inhibiting effect of marriage patterns accounted for an additional 31% reduction, and was most important among the young. Further analysis of contraceptive behaviour suggests that the current period fertility rate of 2.0 children per woman is an outcome of a synchronization of delaying and spacing of births among younger women with stopping of childbearing among women in the middle and late reproductive ages. The policy implications of the results are discussed.     

Adipose derived stem cells (ASCs) isolated from breast cancer tissue express IL-4, IL-10 and TGF-β1 and upregulate expression of regulatory molecules on T cells: do they protect breast cancer cells from the immune response?

Immunomodulatory function of bone marrow derived mesenchymal stem cells in cancer has recently been investigated. But the resident mesenchymal stem cells as whole in cancer and in the breast cancer tissue have not been studied well. In the present work we isolated adipose derived stem cells (ASCs) from breast cancer and normal breast tissues to investigate the expressions of IL-4, IL-10 and transforming growth factor (TGF)-β1 in ASCs and to see if ASCs isolated from patients can modulate the regulatory molecules on peripheral blood lymphocytes. Our results showed that IL-10 and TGF-β1 have significantly higher mRNA expressions in ASCs isolated from breast cancer patients than those from normal individuals (P value <0.05). The culture supernatant of ASCs isolated from breast cancer patients with pathological stage III induced upregulation of the mRNA expression levels of IL-4, TGF-β1, IL-10, CCR4 and CD25 in PBLs. In addition, the percentage of CD4+CD25^highFoxp3⁺ T regulatory cells was increased in vitro. When the same culture supernatant was added to ASCs isolated from normal subjects augmentation of the mRNA expressions of IL-4, IL-10, IL-8, MMP2, VEGF and SDF-1 in normal ASCs was also observed. These data collectively conclude that resident ASCs in breast cancer tissue may have crucial roles in breast tumor growth and progression by inducing regulatory molecules and promoting anti-inflammatory reaction within the tumor microenvironment. Further investigation is required to see if the immune suppression induced by ASCs is an independent property from tumor cells or ASCs gain their immunosuppressive potential from malignant cells.     

Effects of indoleamine 2, 3-dioxygenase (IDO) silencing on immunomodulatory function and cancer-promoting characteristic of adipose-derived mesenchymal stem cells (ASCs)

Indoleamine 2, 3-dioxygenase (IDO) catabolizes tryptophan, mediates immunomodulatory functions, and is released by stromal cells such as mesenchymal stem cells. The aims of this study were to investigate the effects of IDO silencing on immunosuppressive function of adipose-derived mesenchymal stem cells (ASCs), T cells phenotype, and the proliferation/migration of tumor cells. ASCs isolated from adipose tissues of healthy women were transfected with IDO-siRNA. Galectin-3, transforming growth factor-β1, hepatocyte growth factor, and interleukin-10 as immunomodulators were measured in ASCs using qRT-PCR. T cells phenotype, interferon-γ, and interleukin-17 expression were evaluated in peripheral blood lymphocytes (PBLs) cocultured with IDO silenced-ASCs by flow cytometry and qRT-PCR, respectively. Scratch assay was applied to assess the proliferation/migration of MDA-MB-231 cell line. Galectin-3 was upregulated (p ˂ 0.05) while hepatocyte growth factor was downregulated (p ˂ 0.05) in IDO-silenced ASCs compared to control groups. Regulatory T cells were inhibited in PBLs cocultured with IDO-silenced ASCs; also T helper2 was decreased in PBLs cocultured with IDO-silenced ASCs relative to the scramble group. IDO-silenced ASCs caused interferon-γ overexpression but interleukin-17 downregulation in PBLs. The proliferation/migration of MDA-MB-231 was suppressed after exposing to condition media of IDO-silenced ASCs compared with condition media of untransfected (p < 0.01) and scramble-transfected ASCs (p < 0.05). The results exhibited the weakened capacity of IDO-silenced ASCs for suppressing the immune cells and promoting the tumor cells' proliferation/migration. IDO suppression may be utilized as a strategy for cancer treatment. Simultaneous blocking of immunomodulators along with IDO inhibitors may show more effects on boosting the efficiency of immune-based cancer therapies.