Spatial and temporal expression analysis of D-myo-inositol 3-phosphate synthase (MIPS) gene family in Glycine max

Phytic acid, the principal storage form of phosphorus in plant seeds accounts for up to 60 to 80% of the total seed phosphorus content in soybean. Its accumulation increases linearly throughout seed  development and it strongly chelates essential mineral cations and charged proteins decreasing their bioavailability. D-Myo-inositol 3-phosphate synthase (MIPS; EC 5.5.1.4), the evolutionarily conserved enzyme in plants, catalyzes the first and the rate limiting step in phytic acid biosynthetic pathway. Aiming at controlling the level of phytate, we monitored the differential expression profile of four, previously reported, members of the MIPS gene family in developing seeds and vegetative tissues of soybean by quantitative real-time PCR (qRT-PCR). Transcript levels were measured relative to the endogenous reference gene eEF-1á (eukaryotic elongation factor 1-alpha) using SYBER-Green. The qRT-PCR data analysis indicated that the expression of the four highly conserved MIPS genes is both temporally and spatially regulated, information much needed for reverse genetic applications. MIPS1 exhibited high transcript levels in the early developing cotyledons with the levels peaking at around 4 to 6 mm seed size stage. Despite of high level of nucleotide sequence conservation amongst the MIPS gene family members, MIPS2, MIPS3 and MIPS4 were poorly expressed in developing seed tissues, although their transcript levels were relatively high in the other organ tissues. MIPS1 was however moderately expressed in seedlings where MIPS2 showed relatively higher expression levels. Among the four isoforms, MIPS4 had the highest transcript levels in the leaf tissue. The data was clearly indicative of the fact that the four isoforms had diverged regulatory elements resulting in their differential expression. Of the four members of the MIPS gene family, MIPS1 is thus the major isoform that had high expression in the developing seed tissues and can be targeted using the dsRNA induced sequence specific RNA degradation mechanism for reduction of phytate levels without affecting the critical aspects of inositol metabolism in other tissues of the plant.Key words: Soybean, MIPS isoforms, differential expression, endogenous reference gene, qRT-PCR

Hodgkin Lymphoma in a Case of Chronic Myeloid Leukemia Treated with Tyrosine Kinase Inhibitors

Chronic myeloid leukemia (CML) is characterized by increased and unregulated proliferation of granulocytic lineage in the bone marrow and presence of these immature myeloid cells in the peripheral blood with presence of Philadelphia (Ph) chromosome. Tyrosine kinase inhibitors are the most important drugs in the CML therapy and provide long disease-free survival. Due to the increased survival of CML patients with continual administration of these drugs, the chance of development of secondary malignancies may increase. The most common secondary malignancies are prostate, colorectal and lung cancer, non-Hodgkin lymphoma, malignant melanoma, non-melanoma skin tumors and breast cancer. Herein, we are describing a rare case of Hodgkin lymphoma in a patient of CML after ten year of primary disease presentation. Hodgkin lymphoma in a known case of CML is very rare and further studies are also needed to know the pathogenic relationship between the two entities and to assess the risk of secondary Hodgkin lymphoma in CML patients treated with tyrosine kinase inhibitors. CML itself is a risk factor for development of solid cancers and hematologic malignancies. In addition, patients on chemotherapy are immune-compromised and may be at greater risk of neoplasm driven by infectious agents such as Epstein-Barr virus

Valorisation of black carrot pomace: microwave assisted extraction of bioactive phytoceuticals and antioxidant activity using Box–Behnken design

The present study compares three methods viz. microwave assisted extraction (MAE), ultrasonic-assisted extraction (UAE) and conventional solvent extraction (CSE) for extraction of phenolic compounds from black carrot pomace (BCP). BCP is the major by-product generated during processing and poses big disposal problem. Box–Behnken design using response surface methodology was employed to investigate and optimize the MAE of phenolics, antioxidant activity and colour density from BCP. The conditions for maximum recovery of polyphenolics were: microwave power (348.07 W), extraction time (9.8 min), solvent–solid ratio (19.3 mL/g) and ethanol concentration (19.8%). Under these conditions, the extract contained total phenolic content of 264.9 ± 10.02 mg gallic acid equivalents (GAE)/100 mL, antioxidant capacity (AOC) of 13.14 ± 1.05 lmol Trolox equivalents (TE)/mL and colour density of 68.63 ± 5.40 units. The total anthocyanin content at optimized condition was 753.40 ± 31.6 mg/L with low % polymeric colour of 7.40 ± 0.42. At optimized conditions, MAE yielded higher colour density (68.63 ± 5.40), polyphenolic content (264.9 ± 10.025 mg GAE/100 mL) and AOC (13.14 ± 1.05 lmol TE/mL) in a short time as compared to UAE and CSE. Overall results clearly indicate that MAE is the best suited method for extraction in comparison to UAE and CSE. The phenolic rich extract can be used as an effective functional ingredient in foods

Optimization of the use of cellulolytic enzyme preparation for the extraction of health promoting anthocyanins from black carrot using response surface methodology

Anthocyanin-rich extracts from black carrots are being considered as a candidate replacer for the red colour in processed foods. The present investigation optimizes the extraction of anthocyanins with high phenolic content and low degradation parameters from black carrots using a cellulolytic multi-enzyme preparation known as Viscozyme. The optimized conditions for Viscozyme using a Box-Behnken design (BBD) of response surface methodology (RSM) were as follow: temperature (50.2 ◦C), extraction time (58.4 min) and enzyme concentration (0.20%). The predicted value of anthocyanins content was 1380 mg/L, which was near to the optimized experimental value of 1375 mg/L. The extracted anthocyanins based on above mentioned conditions exhibited the lowest degradation parameters such as degradation index (DI) of (0.86), browning index of (BI) (1.31) and were characterized with cyanidin 3-sinapoylxylosylglucosylgalactoside as being the most abundant. The findings clearly reveal that Viscozyme-assisted extraction (VAE) is the best approach for extracting superior quality extracts from black carrots with high anthocyanin and other phenolic component concentrations

Targeting the MAPK7/MMP9 axis for metastasis in primary bone cancer

Metastasis is the leading cause of cancer related death. This multistage process involves contribution from both tumour cells and the tumour stroma to release metastatic cells into the circulation. Circulating tumour cells (CTCs) survive circulatory cytotoxicity, extravasate and colonise secondary sites effecting metastatic outcome. Reprogramming the transcriptomic landscape is a metastatic hallmark but detecting underlying master regulators that drive pathological gene expression is a key challenge, especially in childhood cancer. Here we used whole tumour plus single cell RNA sequencing in primary bone cancer and CTCs to perform weighted gene co-expression network analysis to systematically detect coordinated changes in metastatic transcript expression. This approach with comparisons applied to data collected from cell line models, clinical samples and xenograft mouse models revealed MAPK7/MMP9 signalling as a driver for primary bone cancer metastasis. RNAi knockdown of MAPK7 reduces proliferation, colony formation, migration, tumour growth, macrophage residency/polarisation and lung metastasis. Parallel to these observations were reduction of activated interleukins IL1B, IL6, IL8 plus mesenchymal markers VIM and VEGF in response to MAPK7 loss. Our results implicate a newly discovered, multidimensional MAPK7/MMP9 signalling hub in primary bone cancer metastasis that is clinically actionable