Long bone mesenchymal stem cells (Lb-MSCs): clinically reliable cells for osteo-diseases

Mesenchymal stem cells (MSCs) have been designated as the most reliable cells in clinics to treat osteo-diseases because of their versatile nature. MSCs, isolated from long bone (Lb-MSCs) are rarely reported and named as RIA-MSCs because of the reamer–irrigator–aspirator (RIA) device. The potential of these cells in the treatment of non-union bone fractures made them the ideal candidates to be studied for clinical practices. In this work, effect of cryopreservation on the proliferation and differentiation capabilities of long bone MSCs (Lb-MSCs) has been studied. For this purpose, Lb-MSCs were isolated via RIA device and characterized using flow cytometry and differentiation assays. Cells were cryopreserved for 3, 6 and 12 months and thereafter were characterized using differentiation assays and genetic markers specific for osteogenic, chondrogenic, and adipogenic potential quantitatively by qRT-PCR. Lb-MSCs were found expressing MSC characteristic markers defining their identity. The population doubling time (PDT) was about 2.5 ± 0.5 days and colonies appeared after 7–10 days. Differentiation potential and gene expression of 3, 6 and 12 months cryopreserved Lb-MSCs were unaltered. The results show that cryopreservation did not have an effect on the differentiation potential of human Lb-MSCs. Therefore, our work offers Lb-MSCs as clinically cells for treating osteo-diseases.     

Mutation in the beta3 subunit of Guanine nucleotide-binding protein (GNB3) gene is not associated with Type II diabetes mellitus risk: a case–control study of a North Indian population

Context: Genetics play a major role in development and pathophysiology of Type 2 diabetes mellitus (T2DM).

Objective: To asses the association of Guanine nucleotide-binding protein (GNB3) (C825T) gene's polymorphism with T2DM.

Materials and methods: A case–control study including 400 North Indians was performed using Polymerase Chain Reaction–Restriction Fragment Length Polymorphism (PCR-RFLP) approach to analyze genetic polymorphism.

Results: No significant difference was observed in genotype and allele frequencies of GNB3 gene on comparing cases with controls.

Discussion: Our study is in agreement with studies on Polish, Japanese, Hispanic-American and Danish populations who observed no significant association between GNB3 (C825T) polymorphism and T2DM.

Conclusion: GNB3 (C825T) polymorphism is not associated with T2DM.     

Antioxidant defence system and physiological responses of Iranian crested wheatgrass (<Emphasis Type="Italic">Agropyron cristatum</Emphasis> L.) to drought and salinity stress

Crested wheatgrass (Agropyron cristatum L.) is a cool-season perennial grass, which has demonstrated its potential for use as turfgrass. However, limited information is available on its drought and salinity tolerance. The main purpose of this study was to investigate the changes in the antioxidant defence system and physiological traits of six Iranian crested wheatgrass genotypes under drought and salinity stresses. The experimental design comprised a split plot with water treatments (control well-watered, salinity stress and water stress) as main plots and genotypes as subplots. This study demonstrated the variations in drought and salinity tolerance among crested wheatgrass genotypes. ‘ACSKI’, ‘ACAMI’ and ‘ACDTI’ generally performed better than other genotypes under drought and salinity conditions, mainly by maintaining higher activities of antioxidant enzymes like superoxide dismutase, peroxidase, catalase, ascorbate peroxidase and non-enzyme antioxidants like glutathione, higher proline and total non-structural carbohydrates content. The increased, decreased, and unchanged activities of antioxidant enzymes in the crested wheatgrass genotypes indicates a different forms of metabolism of antioxidant enzymes in response to drought and salinity stress. In general, drought and salinity stress increases the malondialdehyde (MDA) content and hydrogen peroxide (H₂O₂) content; however, ‘ACSKI’, ‘ACAMI’ and ‘ACDTI’ genotypes could tolerate an increase in MDA content and H₂O₂ content; therefore, lower levels of MDA content and H₂O₂ content were observed. The results showed that increasing levels of diamine oxidase and polyamine oxidase have been associated with increasing drought and salinity tolerance. Based on the biochemical and physiological parameters that were evaluated, we concluded that the genotype ‘ACSKI’ was superior in terms of drought and salinity tolerance. This superiority was mainly a result of better enzymatic and non-enzymatic antioxidant defence system and better osmotic adjustment under stress conditions.     

Molecular characterization of <Emphasis Type="Italic">AIPL1</Emphasis> gene region in the Iranian population: application of novel informative haplotypes and detection of mutational founder effect

Leber’s congenital amaurosis (LCA) is considered as one of the main causes of congenital blindness. In view of the genetically heterogeneous nature of the disease, indirect diagnosis using linkage analysis has proven to be useful in molecular diagnosis procedure. Mutations in AIPL1 gene are one of the leading causes of LCA. In the present study, the application of three single nucleotide polymorphic (SNP) markers related to the gene, including rs7212734, rs11658369 and rs8066853 was evaluated for the first time in the Iranian population. The markers were genotyped using tetra-primer ARMS PCR in 154 unrelated healthy individuals. Haplotype frequency and other characteristics of the markers were examined by using the GENEPOP, PowerMarker and Cubic Exact Solution software. The data indicated the presence of six different haplotypes in the Iranian population. Among them, three haplotypes showed high informativeness with frequencies ≥0.05. Three unrelated Iranian LCA families were analyzed. The p.W278* mutation in exon 6 of AIPL1 was found in all the families using APEX microarray chips. SNP analysis for the families showed the conservation of T?A?A haplotype linked to p.W278* mutation. All families bearing the mutation were from the Isfahan province and a founder effect was suggested in this population. Prenatal diagnosis using the markers led to the successful prediction of the fetus genotypes in all the at risk pregnancies and showed that rs7212734, rs11658369 and rs8066853 can be considered as the three informative markers for linkage analysis in carrier detection and molecular diagnosis of LCA in the Iranian population.     

The delayed effect of mustard gas on housekeeping gene expression in lung biopsy of chemical injuries

ObjectiveSulfur mustard (SM) was used as a chemical weapon in Iraq-Iran war. Exposed people have major complications in important organs such as pulmonary system. Some studies have shown that SM could affect the expression of endogenous genes and non-housekeeping genes, time dependently. To understand the accurate molecular mechanism of the delayed effect of SM, the identification of the gene expression pattern in these patients is essential. Hence, we have evaluated mRNA expression of four common housekeeping genes (ACTIN, PGK1, β2m, GAPDH) in SM-exposed and non-exposed (control) formalin-fixed, paraffin-embedded (FFPE) human lung tissues.MethodParaffin block of lung biopsy of SM-exposed people (11 cases) and people without exposure to SM as control group (9 cases) have been selected. The mRNA expression of four endogenous control genes has been evaluated by qRT-PCR. The stability value of each gene was calculated by different methods.ResultIt was found that ACTIN mRNA has the highest expression (30.26±2.87) and PGK1 has the lowest standard deviation (SD) (30.885±2.215) between pooled groups. The best correlation was between ACTIN and PGK1 expressions. The M value has shown that ACTIN and then PGK1 are the most stable housekeeping genes among. The results obtained from the GeNorm and NormFinder have indicated that the pair ACTIN- PGK1 is the most suitable choice for endogenous control genes.ConclusionACTIN and PGK1 genes are stable in studied lung tissues and are the better than two other housekeeping genes. In addition, mustard gas does not affect their expression in long term.     

Deprotection of centromeric cohesin at meiosis II requires APC/C activity but not kinetochore tension

Genome haploidization involves sequential loss of cohesin from chromosome arms and centromeres during two meiotic divisions. At centromeres, cohesin''s Rec8 subunit is protected from separase cleavage at meiosis I and then deprotected to allow its cleavage at meiosis II. Protection of centromeric cohesin by shugoshin‐PP2A seems evolutionarily conserved. However, deprotection has been proposed to rely on spindle forces separating the Rec8 protector from cohesin at metaphase II in mammalian oocytes and on APC/C‐dependent destruction of the protector at anaphase II in yeast. Here, we have activated APC/C in the absence of sister kinetochore biorientation at meiosis II in yeast and mouse oocytes, and find that bipolar spindle forces are dispensable for sister centromere separation in both systems. Furthermore, we show that at least in yeast, protection of Rec8 by shugoshin and inhibition of separase by securin are both required for the stability of centromeric cohesin at metaphase II. Our data imply that related mechanisms preserve the integrity of dyad chromosomes during the short metaphase II of yeast and the prolonged metaphase II arrest of mammalian oocytes.     

Temperature and photoperiod affect stress resistance traits in Drosophila melanogaster

The long‐term survival of species and populations depends on their ability to adjust phenotypic values to environmental conditions. In particular, the capability of dealing with environmental stress to buffer detrimental effects on fitness is considered to be of pivotal importance. Resistance traits are readily modulated by a wide range of environmental factors. In the present study, Drosophila melanogaster Meigen is used to investigate plastic responses to temperature and photoperiod in stress resistance traits. The results reveal that stress resistance traits (cold, heat, starvation and desiccation resistance) are affected by the factors temperature and sex predominantly. Cooler temperatures compared with warmer temperatures increase cold tolerance, desiccation and starvation resistance, whereas they reduce heat tolerance. Except for heat resistance, females are more stress‐resistant than males. Stress resistance traits are also affected by photoperiod. Shorter photoperiods decrease cold tolerance, whereas longer photoperiods enhance desiccation resistance. Overall, thermal effects are pervasive throughout all measured resistance traits, whereas photoperiodic effects are of limited importance in the directly developing (i.e. nondiapausing) flies used here, suggesting that pronounced photoperiodic effects on stress resistance traits may be largely limited to, and triggered by, diapause‐inducing effects.