Site-Specific Keloid Fibroblasts Alter the Behaviour of Normal Skin and Normal Scar Fibroblasts through Paracrine Signalling

Keloid disease (KD) is an abnormal cutaneous fibroproliferative disorder of unknown aetiopathogenesis. Keloid fibroblasts (KF) are implicated as mediators of elevated extracellular matrix deposition. Aberrant secretory behaviour by KF relative to normal skin fibroblasts (NF) may influence the disease state. To date, no previous reports exist on the ability of site-specific KF to induce fibrotic-like phenotypic changes in NF or normal scar fibroblasts (NS) by paracrine mechanisms. Therefore, the aim of this study was to investigate the influence of conditioned media from site-specific KF on the cellular and molecular behaviour of both NF and NS enabled by paracrine mechanisms. Conditioned media was collected from cultured primary fibroblasts during a proliferative log phase of growth including: NF, NS, peri-lesional keloid fibroblasts (PKF) and intra-lesional keloid fibroblasts (IKF). Conditioned media was used to grow NF, NS, PKF and IKF cells over 240 hrs. Cellular behavior was monitored through real time cell analysis (RTCA), proliferation rates and migration in a scratch wound assay. Fibrosis-associated marker expression was determined at both protein and gene level. PKF conditioned media treatment of both NF and NS elicited enhanced cell proliferation, spreading and viability as measured in real time over 240 hrs versus control conditioned media. Following PKF and IKF media treatments up to 240 hrs, both NF and NS showed significantly elevated proliferation rates (p<0.03) and migration in a scratch wound assay (p<0.04). Concomitant up-regulation of collagen I, fibronectin, α-SMA, PAI-1, TGF-β and CTGF (p<0.03) protein expression were also observed. Corresponding qRT-PCR analysis supported these findings (P<0.03). In all cases, conditioned media from growing marginal PKF elicited the strongest effects. In conclusion, primary NF and NS cells treated with PKF or IKF conditioned media exhibit enhanced expression of fibrosis-associated molecular markers and increased cellular activity as a result of keloid fibroblast-derived paracrine factors.     

Genetic manipulation, a feasible tool to enhance unique characteristic of Chlorella vulgaris as a feedstock for biodiesel production

Developing a reliable technique to transform unicellular green algae, Chlorella vulgaris, could boost potentials of using microalgae feedstock in variety of applications such as biodiesel production. Volumetric lipid productivity (VLP) is a suitable variable for evaluating potential of algal species. In the present study, the highest VLP level was recorded for C. vulgaris (79.08 mg l^?1 day^?1) followed by 3 other strains studied; C. emersonii, C. protothecoides, and C. salina by 54.41, 45 and 18.22 mg l^?1day^?1, respectively. Having considered the high productivity of C. vulgaris, it was selected for the preliminary transformation experiment through micro-particle bombardment. Plasmid pBI 121, bearing the reporter gene under the control of CaMV 35S promoter and the kanamycin marker gene, was used in cells bombardment. Primary selection was done on a medium supplemented by 50 mg l^?1 kanamycin. After several passages, the survived cells were PCR-tested to confirm the stability of transformation and then were found to exhibit β-glucuronidase (GUS) activity in comparison with the control cells. Southern hybridization of npt II probe with genomic DNA revealed stable integration of the cassette in three different positions in the genome. The whole process was successfully implemented as a pre-step to transform the algal cells by genes involved in lipid production pathway which will be carried out in our future studies.     

Relationship between susceptibility to neonicotinoids and population dynamics of cotton aphid,Aphis gossypii Glover (Hemiptera: Aphididae)

Efficiency of imidacloprid and thiametoxam on population growth parameters of Aphis gossypii Glover (Hemiptera: Aphididae) was assessed using three bioassay methods including; residual, starvation and FAO dip methods. Aphids were assayed under laboratory conditions at 23 ± 2°C, photoperiod of 16:8 (L:D) and 70% relative humidity. Aphids were transferred to sprayed leaves and Petri dishes in residual and starvation methods, respectively. There was no feeding for the aphids in starvation method. In FAO dip method, insects were dipped for 5 s in pesticide solutions and then transferred to fresh leaves. Results revealed that LC₅₀ values calculated with the starvation and residual methods were respectively 15 and 11% more than FAO dip method (for imidacloprid) and 22 and 18% (for thiametoxam). The LC₅₀ value in starvation method was 3% more than the residual method. Calculated LC₅₀ in starvation and residual methods with imidacloprid respectively caused 160 and 34% increase in intrinsic rates of increase (r _m) and net reproductive rate (R ₀) in comparison with FAO dip method. Generation time (T) and doubling time (DT) were respectively 59 and 62% less than those in FAO dip method. In contrast, thiametoxam (LC₅₀ concentration) in starvation and residual method lead to 9 and 67% increase in r _m and R ₀ parameters compared to FAO dip method. However T and DT were respectively 65 and 92% less than mentioned parameters in FAO dip method. There was not any significant difference between life table parameters calculated by residual and starvation bioassay methods.     

Assessment of genetic diversity among and within Carthamus species using sequence-related amplified polymorphism (SRAP) markers

Due to precise evaluation of genetic diversity of Carthamus species, sixty-two genotypes consisting fifty-two from five wild (C. oxyacanthus M. Bieb, C. lanatus L., C. dentatus Vahl, C. boissieri Halácsy, C. glaucus M.B.) and ten from cultivated species (C. tinctorius L.) were selected for evaluation of the genetic diversity in Carthamus species. A total of 238 (81.2 %) polymorphic bands were detected by 12 SRAP primer combinations with an average of 22 bands per combination. Me4-Em1 and Me5-Em2 primer combinations were known as the most informative SRAP markers based on the PIC values (0.34) where they distinguished all studied Carthamus species. Cluster analysis classified all accessions into five main groups among which clusters containing cultivated individuals were distinctly separated from those containing wilds. The most and the least genetic variation based on analysis of molecular variance, were detected within (76.90 %) and among (22.84 %) groups, respectively. The obtained results suggested that C. dentatus, C. glaucus and C. boissieri species may be classified in one section including C. dentatus in one and C. glaucus and C. boissieri in another subsection. The results also revealed high genetic similarity between C. oxyacanthus and C. tinctorius despite their different morphological characteristics.     

Comparison of genetic variation and differentiation among annual Cicer species using start codon targeted (SCoT) polymorphism, DAMD-PCR, and ISSR markers

Three molecular markers, including start codon targeted (SCoT) polymorphism, directed amplification of minisatellite-region DNA polymerase chain reaction (DAMD-PCR), and inter simple sequence repeat (ISSR) markers, were compared in terms of their informativeness and efficiency for analysis of genetic relationships among 38 accessions of eight annual Cicer species. The results were as follows: (1) the highest level of detected polymorphism was observed for all three marker types; (2) the rate of diversity for the three marker techniques was approximately equal, and the correlation coefficients of similarity were statistically significant for all three marker systems; (3) the three molecular markers showed relatively similar phylogenetic grouping for examined species. Diversity analysis showed that Cicer reticulatum is the closest wild species to the cultivated chickpea, and this finding supports the hypothesis that C.?reticulatum is the most probable progenitor of the cultivated species. C.?bijugum, C.?judaicum, and C.?pinnatifidum were clustered together, and in other clusters C.?yamashitae and C.?cuneatum were grouped close together. To our knowledge, this is the first detailed comparison of performance among two targeted DNA region molecular markers (SCoT and DAMD-PCR) and the ISSR technique on a set of samples of Cicer. The results provide guidance for future efficient use of these molecular methods in genetic analysis of Cicer.     

The Effects of Magnesium, l-Carnitine, and Concurrent Magnesium–l-Carnitine Supplementation in Migraine Prophylaxis

Given the conflicting results about the positive effects of magnesium and l-carnitine and as there is no report concerning concurrent supplementation of magnesium and l-carnitine on migraine prophylaxis, the effects of magnesium, l-carnitine, and concurrent magnesium?Cl-carnitine supplementation on migraine indicators were assessed. In this clinical trial, 133 migrainous patients were randomly assigned into three intervention groups: magnesium oxide (500?mg/day), l-carnitine (500?mg/day), and Mg?Cl-carnitine (500?mg/day magnesium and 500?mg/day?l-carnitine), and a control group. After 12?weeks of supplementation, the checklist of migraine indicators including migraine attacks/month, migraine days/month, and headache severity was completed, and serum concentrations of magnesium and l-carnitine were measured by atomic absorption spectrophotometry and enzymatic UV test, respectively. The results showed a significant reduction in all migraine indicators in all studied groups (p?<?0.05). The ANOVA results showed a significant reduction in migraine frequency across various supplemented and control groups (p?=?0.008). By separating the effects of magnesium supplementation from other confounding factors such as routine treatments using the repeated measures and nested model, it was clarified that magnesium supplementation had a significant effect on all migraine indicators. Oral supplementation with magnesium oxide and l-carnitine and concurrent supplementation of Mg?Cl-carnitine besides routine treatments could be effective in migraine prophylaxis; however, larger trials are needed to confirm these preliminary findings.     

Oxydemeton-methyl resistance, mechanisms, and associated fitness cost in green peach aphids (Hemiptera: Aphididae)

Susceptibility to oxydemeton-methyl and imidacloprid, and the inhibitory effects of oxydemeton-methyl and some organophosphate compounds on acetylcholinesterase (AChE) and carboxylesterase activity were studied in two populations (Karaj and Rasht) of green peach aphids, Myzus persicae (Sulzer). Results show that the Karaj population was resistant to oxydemeton-methyl but susceptible to imidacloprid. The esterase activity of the resistant and susceptible populations suggests that one of the resistance mechanisms to oxydemeton-methyl was esterase-based. The inhibition assay shows that the AChE of the Karaj population is less sensitive to oxydemeton-methyl and paraoxon derivatives. Regarding the paraoxon derivatives, the smaller paraoxon side chain is more potent against the modified AChE than against the AChE from the susceptible strain. Fertility life table parameters of green peach aphid populations resistant and susceptible to oxydemeton-methyl also were studied under laboratory conditions. The standard errors of the population growth parameters were calculated using the Jackknife method. Results showed that susceptible strain exhibits a significantly higher r(m) than the resistant strain, probably because the resistant strain had a higher generation time than the susceptible strain. These results suggested that the resistant Karaj strain may be less fit than the susceptible strain.     

Diversity of Sinorhizobium strains nodulating Medicago sativa from different Iranian regions

Alfalfa is believed to have originated in north-western Iran and has a long history of coexistence with its bacterial symbiont Sinorhizobium in soils of Iran. However, little is known about the diversity of Sinorhizobium strains nodulating Iranian alfalfa genotypes. In this study, Sinorhizobium populations were sampled from eight different Iranian sites using three cultivars of Medicago sativa as trap host plants. A total of 982 rhizobial strains were isolated and species were identified showing a large prevalence of Sinorhizobium meliloti over Sinorhizobium medicae. Analysis of salt tolerance demonstrated a great phenotypic diversity. The genetic diversity of the Sinorhizobium isolates was analysed using BOX-PCR and enterobacterial repetitive intergenic consensus (ERIC)-PCR. Patterns ofBOX-PCR fingerprinting were statistically analysed with AMOVA to evaluate the role of plant variety and site of origin in the genetic variance observed. Results indicated that most of the total molecular variance was attributable to divergence among strains isolated from different sites and cultivars (intrapopulation, strain-by-strain variance). Moreover, the analysis showed the presence of two geographic populations (west and northwest), indicating that the effect of the site of origin could be more relevant in shaping population genetic diversity than the effect of cultivar or individual plant.