PHYTOCHEMICAL SCREENING AND IN VITRO ANTIOXIDANT POTENTIALS OF EXTRACTS OF TEN MEDICINAL PLANTS USED FOR THE TREATMENT OF DIABETES MELLITUS IN SRI LANKA

Background: Over the last decade, extensive research work has focused on the potential health benefits of antioxidants while many medicinal plant extracts have been evaluated for their antioxidant profile. Medicinal plants selected for this study are widely used in traditional medicine for the treatment of diabetes mellitus in Sri Lanka some of which are recommended as dietary supplements to the existing therapies. The present study aimed at determining the total polyphenol contents and total antioxidant activities of aqueous extracts of 10 selected Sri Lankan medicinal plants by three in vitro methods; Materials and Methods: DPPH(2, 2-diphenyl-2-picrylhydrazyl), FRAP (ferric reducing power) and NO (nitric oxide) assays. The aqueous plant extracts were tested at the concentration of 0.05 g/mL. The total polyphenol content was determined according to the Folin-Ciocalteu method while the total antioxidant activity was evaluated by DPPH, FRAP and NO assays with L-ascorbic acid as reference compound. Results: The total polyphenol content of the plant extracts varied from 0.41 to 13.00 mg GAE (gallic acid equivalents) per gram dry weight. The antioxidant activities ranged in IC50 of 36.89-101.27 µg/mL, IC50 of 139.56-419.93 µg/mL, 0.12-8.98 µM for DPPH, NO, FRAP assays, respectively. A significant positive correlation was observed between total polyphenol content and antioxidant activities (P< 0.05). Conclusion: Phytochemical analysis revealed the presence of polyphenolic compounds, alkaloids and flavonoids in the plant materials which also possessed in vitro antioxidant potentials. Polyphenolic compounds contribute significantly to the total antioxidant capacities of medicinal plant extracts

A Whole-Chromosome Analysis of Meiotic Recombination in Drosophila melanogaster

Although traditional genetic assays have characterized the pattern of crossing over across the genome in Drosophila melanogaster, these assays could not precisely define the location of crossovers. Even less is known about the frequency and distribution of noncrossover gene conversion events. To assess the specific number and positions of both meiotic gene conversion and crossover events, we sequenced the genomes of male progeny from females heterozygous for 93,538 X chromosomal single-nucleotide and InDel polymorphisms. From the analysis of the 30 F1 hemizygous X chromosomes, we detected 15 crossover and 5 noncrossover gene conversion events. Taking into account the nonuniform distribution of polymorphism along the chromosome arm, we estimate that most oocytes experience 1 crossover event and 1.6 gene conversion events per X chromosome pair per meiosis. An extrapolation to the entire genome would predict approximately 5 crossover events and 8.6 conversion events per meiosis. Mean gene conversion tract lengths were estimated to be 476 base pairs, yielding a per nucleotide conversion rate of 0.86 × 10−5 per meiosis. Both of these values are consistent with estimates of conversion frequency and tract length obtained from studies of rosy, the only gene for which gene conversion has been studied extensively in Drosophila. Motif-enrichment analysis revealed a GTGGAAA motif that was enriched near crossovers but not near gene conversions. The low-complexity and frequent occurrence of this motif may in part explain why, in contrast to mammalian systems, no meiotic crossover hotspots have been found in Drosophila

Off-target piRNA gene silencing in Drosophila melanogaster rescued by a transposable element insertion

Transposable elements (TE) are selfish genetic elements that can cause harmful mutations. In Drosophila, it has been estimated that half of all spontaneous visible marker phenotypes are mutations caused by TE insertions. Several factors likely limit the accumulation of exponentially amplifying TEs within genomes. First, synergistic interactions between TEs that amplify their harm with increasing copy number are proposed to limit TE copy number. However, the nature of this synergy is poorly understood. Second, because of the harm posed by TEs, eukaryotes have evolved systems of small RNA-based genome defense to limit transposition. However, as in all immune systems, there is a cost of autoimmunity and small RNA-based systems that silence TEs can inadvertently silence genes flanking TE insertions. In a screen for essential meiotic genes in Drosophila melanogaster, a truncated Doc retrotransposon within a neighboring gene was found to trigger the germline silencing of ald, the Drosophila Mps1 homolog, a gene essential for proper chromosome segregation in meiosis. A subsequent screen for suppressors of this silencing identified a new insertion of a Hobo DNA transposon in the same neighboring gene. Here we describe how the original Doc insertion triggers flanking piRNA biogenesis and local gene silencing. We show that this local gene silencing occurs in cis and is dependent on deadlock, a component of the Rhino-Deadlock-Cutoff (RDC) complex, to trigger dual-strand piRNA biogenesis at TE insertions. We further show how the additional Hobo insertion leads to de-silencing by reducing flanking piRNA biogenesis triggered by the original Doc insertion. These results support a model of TE-mediated gene silencing by piRNA biogenesis in cis that depends on local determinants of transcription. This may explain complex patterns of off-target gene silencing triggered by TEs within populations and in the laboratory. It also provides a mechanism of sign epistasis among TE insertions, illuminates the complex nature of their interactions and supports a model in which off-target gene silencing shapes the evolution of the RDC complex

Evaluation of commercially available RNA amplification kits for RNA sequencing using very low input amounts of total RNA

This article includes supplemental data. Please visit http://www.fasebj.org to obtain this information.Multiple recent publications on RNA sequencing (RNA-seq) have demonstrated the power of next-generation sequencing technologies in whole-transcriptome analysis. Vendor-specific protocols used for RNA library construction often require at least 100 ng total RNA. However, under certain conditions, much less RNA is available for library construction. In these cases, effective transcriptome profiling requires amplification of subnanogram amounts of RNA. Several commercial RNA amplification kits are available for amplification prior to library construction for next-generation sequencing, but these kits have not been comprehensively field evaluated for accuracy and performance of RNA-seq for picogram amounts of RNA. To address this, 4 types of amplification kits were tested with 3 different concentrations, from 5 ng to 50 pg, of a commercially available RNA. Kits were tested at multiple sites to assess reproducibility and ease of use. The human total reference RNA used was spiked with a control pool of RNA molecules in order to further evaluate quantitative recovery of input material. Additional control data sets were generated from libraries constructed following polyA selection or ribosomal depletion using established kits and protocols. cDNA was collected from the different sites, and libraries were synthesized at a single site using established protocols. Sequencing runs were carried out on the Illumina platform. Numerous metrics were compared among the kits and dilutions used. Overall, no single kit appeared to meet all the challenges of small input material. However, it is encouraging that excellent data can be recovered with even the 50 pg input total RNA

scRNA-Seq reveals distinct stem cell populations that drive hair cell regeneration after loss of Fgf and Notch signaling.

Loss of sensory hair cells leads to deafness and balance deficiencies. In contrast to mammalian hair cells, zebrafish ear and lateral line hair cells regenerate from poorly characterized support cells. Equally ill-defined is the gene regulatory network underlying the progression of support cells to differentiated hair cells. scRNA-Seq of lateral line organs uncovered five different support cell types, including quiescent and activated stem cells. Ordering of support cells along a developmental trajectory identified self-renewing cells and genes required for hair cell differentiation. scRNA-Seq analyses of fgf3 mutants, in which hair cell regeneration is increased, demonstrates that Fgf and Notch signaling inhibit proliferation of support cells in parallel by inhibiting Wnt signaling. Our scRNA-Seq analyses set the foundation for mechanistic studies of sensory organ regeneration and is crucial for identifying factors to trigger hair cell production in mammals. The data is searchable and publicly accessible via a web-based interface

Gmelina arborea Roxb. (Family: Verbenaceae) Extract Upregulates the β-Cell Regeneration in STZ Induced Diabetic Rats

Gmelina arborea Roxb. (common name: Et-demata, Family: Verbenaceae) has been used traditionally in Sri Lanka as a remedy against diabetes mellitus. The objective of the present study was to evaluate antidiabetic mechanisms of the aqueous bark extract of G. arborea in streptozotocin induced (STZ) diabetic male Wistar rats. Aqueous bark extract of G. arborea (1.00 g/kg) and glibenclamide as the standard drug (0.50 mg/kg) were administered orally using a gavage to STZ diabetic rats (65 mg/kg, ip) for 30 days. The antidiabetic mechanisms of aqueous extract of G. arborea (1.00 g/kg) were determined at the end of the experiment. The fasting blood glucose concentration was significantly lowered and the serum insulin and C-peptide concentrations were increased by 57% and 39% in plant extract treated rats on day 30, respectively (p<0.05). The histopathology and immunohistochemistry results of the plant extract treated group showed a regenerative effect on β-cells of the pancreas in diabetic rats. In addition, serum lipid parameters were improved in G. arborea extract treated diabetic rats. The results revealed that the aqueous stem bark extract of G. arborea (1.00 g/kg) showed beneficial effects against diabetes mellitus through upregulating the β-cell regeneration and biosynthesis of insulin in diabetic rats

Antidiabetic Activity of Widely Used Medicinal Plants in the Sri Lankan Traditional Healthcare System: New Insight to Medicinal Flora in Sri Lanka

The use of medicinal plant extracts and their isolated bioactive compounds for the management of diabetes mellitus has been tremendously increased in recent decades. The present study aimed at providing in-depth information on medicinal flora that has been widely used in the Sri Lankan traditional healthcare system for the management of diabetes mellitus. The data of this review article were obtained from published articles from January 2000 to September 2020 in scientific databases of PubMed, Web of Science, and Google Scholar. In this review, a total number of 18 medicinal plants with the antidiabetic activity were expressed, and their isolated antidiabetic active compounds were highlighted as new drug leads. Results of the reported studies revealed that medicinal plants exert a potent antidiabetic activity via both in vitro and in vivo study settings. However, bioactive compounds and antidiabetic mechanism (s) of action of many of the reported medicinal plants have not been isolated/elucidated the structure in detail, to date. Reported antidiabetic medicinal plants with other properties such as antioxidant and antihyperlipidemic activities deliver new entities for the development of antidiabetic agents with multiple therapeutic targets. This is a comprehensive review on potential antidiabetic activities of the Sri Lankan medicinal plants that have been widely used in the traditional healthcare system. The information presented here would fill the gap between the use of them by traditional healers in the traditional medicine healthcare system in Sri Lanka and their potency for development of new drug entities in future

Value of simple clinical parameters to predict insulin resistance among newly diagnosed patients with type 2 diabetes in limited resource settings.

BackgroundInsulin resistance (IR) has been considered as a therapeutic target in the management of type 2 diabetes mellitus (T2DM). Readily available, simple and low cost measures to identify individuals with IR is of utmost importance for clinicians to plan optimal management strategies. Research on the associations between surrogate markers of IR and routine clinical and lipid parameters have not been carried out in Sri Lanka, a developing country with rising burden of T2DM with inadequate resources. Therefore, we aimed to study the utility of readily available clinical parameters such as age, body mass index (BMI), waist circumference (WC) and triglyceride to high density lipoprotein cholesterol ratio (TG/HDL-C) in the fasting lipid profile in predicting IR in a cohort of patients with newly diagnosed T2DM in Sri Lanka.Methods and findingsWe conducted a community based cross sectional study involving of 147 patients (age 30-60 years) with newly diagnosed T2DM in a suburban locality in Galle district, Sri Lanka. Data on age, BMI, WC, fasting plasma glucose (FPG) concentration, fasting insulin concentration and serum lipid profile were collected from each subject. The indirect IR indices namely homeostasis model assessment (HOMA), quantitative insulin sensitivity check index (QUICKI) and McAuley index (MCA) were estimated. Both clinical and biochemical parameters across the lowest and the highest fasting insulin quartiles were compared using independent sample t-test. Linear correlation analysis was performed to assess the correlation between selected clinical parameters and indirect IR indices. The area under the receiver operating characteristic (ROC) curve was obtained to calculate optimal cut-off values for the clinical markers to differentiate IR. BMI (pConclusionsThe results revealed that there was a significant positive correlation between BMI, WC and HOMA while a significant negative correlation with QUICKI and MCA among the cohort of patients with newly diagnosed T2DM. The cut-off values of BMI and WC as 24.91 kg/m2 and 81.5 cm respectively could be used as simple clinical parameters to identify IR in newly diagnosed patients with T2DM. Our results could be beneficial in rational decision making in the management of newly diagnosed patients with T2DM in limited resource settings