Evaluation of pharmacological efficacy and safety of hydroxyurea in sickle cell disease: Study of a pediatric cohort from Chhattisgarh, India.

Sickle cell disease (SCD) is a disease of abnormal hemoglobin associated with severe clinical phenotype and recurrent complications. Hydroxyurea (HU) is one of the US-FDA approved and commonly used drug for the treatment of adult SCD patients with clinical -severity. However, its use in the pediatric groups remains atypical. Despite a high prevalence of the disease in the state Chhattisgarh, there is a lack of evidence supporting its use in pediatric patients. This study aimed to evaluate the pharmacological and clinical efficacy and safety of HU in a large pediatric cohort with SCD from Central India. The study cohort consisted of 164 SCD (138 Hb SS and 26 Hb S beta-thalassemia) children (≤14 years of age) on HU therapy, who were monitored for toxicity, hematological and clinical efficacy at baseline (Pre-HU) and after 24 months (Post-HU). The results highlight the beneficial effects of HU at a mean dose of 18.7 ± 7.0 mg/kg/day. A significant improvement was observed, not only in physical and clinical parameters but also in hematological parameters which include fetal hemoglobin (Hb F), total hemoglobin, hematocrit, mean corpuscular volume (MCV) and mean corpuscular hemoglobin (MCH) levels, when evaluated against the baseline. We did not observe any significant adverse effects during the treatment period. Similar results were obtained on independent analysis of Hb SS and Hb Sß patients. These findings strengthen the beneficial effect of hydroxyurea in pediatric population also without any serious adverse effects and builds up ground for expanding its use under regular monitoring.

Rapid and accurate nucleobase detection using FnCas9 and its application in COVID-19 diagnosis.

Rapid detection of DNA/RNA pathogenic sequences or variants through point-of-care diagnostics is valuable for accelerated clinical prognosis, as witnessed during the recent COVID-19 outbreak. Traditional methods relying on qPCR or sequencing are tough to implement with limited resources, necessitating the development of accurate and robust alternative strategies. Here, we report FnCas9 Editor Linked Uniform Detection Assay (FELUDA) that utilizes a direct Cas9 based enzymatic readout for detecting nucleobase and nucleotide sequences without trans-cleavage of reporter molecules. We also demonstrate that FELUDA is 100% accurate in detecting single nucleotide variants (SNVs), including heterozygous carriers, and present a simple web-tool JATAYU to aid end-users. FELUDA is semi-quantitative, can adapt to multiple signal detection platforms, and deploy for versatile applications such as molecular diagnosis during infectious disease outbreaks like COVID-19. Employing a lateral flow readout, FELUDA shows 100% sensitivity and 97% specificity across all ranges of viral loads in clinical samples within 1hr. In combination with RT-RPA and a smartphone application True Outcome Predicted via Strip Evaluation (TOPSE), we present a prototype for FELUDA for CoV-2 detection closer to home.

Ultrafine particulate matter impairs mitochondrial redox homeostasis and activates phosphatidylinositol 3-kinase mediated DNA damage responses in lymphocytes.

Particulate matter (PM), broadly defined as coarse (2.5-10 µm), fine (0.1-2.5 µm) and ultrafine particles (≤0.1 µm), is a major constituent of ambient air pollution. Recent studies have linked PM exposure (coarse and fine particles) with several human diseases including cancer. However, the molecular mechanisms underlying ultrafine PM exposure induced cellular and sub-cellular repercussions are ill-defined. Since mitochondria are one of the major targets of different environmental pollutants, we herein aimed to understand the molecular repercussion of ultrafine PM exposure on mitochondrial machinery in peripheral blood lymphocytes. Upon comparative analysis, a significantly higher DCF fluorescence was observed in ultrafine PM exposed cells that confirmed the strong pro-oxidant nature of these particles. In addition, the depleted activity of antioxidant enzymes, glutathione reductase and superoxide dismutase suggested the strong association of ultrafine PM with oxidative stress. These results further coincided with mitochondrial membrane depolarization, altered mitochondrial respiratory chain enzyme activity and decline in mtDNA copy number. Moreover, the higher accumulation of DNA damage response proteins (γH2AX, pATM, p-p53), suggested that exposure to ultrafine PM induces DNA damage and triggers phosphatidylinositol 3 kinase mediated response pathway. Further, the alterations in mitochondrial machinery and redox balance among ultrafine PM exposed cells were accompanied by a considerably elevated pro-inflammatory cytokine response. Interestingly, the lower apoptosis levels observed in ultrafine particle treated cells suggest the possibility that the marked alterations may lead to the impairment of mitochondrial-nuclear cross talk. Together, our results showed that ultrafine PM, because of their smaller size possesses significant ability to disturb mitochondrial redox homeostasis and activates phosphatidylinositol 3 kinase mediated DNA damage response pathway, an unknown molecular paradigm of ultrafine PM exposure. Our findings also indicate that maneuvering through the mitochondrial function might be a viable, indirect method to modulate lymphocyte homeostasis in air pollution associated immune disorders.

Amelioration of oxidative and inflammatory changes by Swertia chirayita leaves in experimental arthritis.

The present study was aimed to determine the therapeutic effects of Swertia chirayita leaves against oxidative and inflammatory injuries in Freund's complete adjuvant (FCA) induced arthritic rats. The extract was evaluated for its phytoconstituents and various invitro antioxidant properties followed by its in vivo effects. The hydroethanolic extract of S. chirayita leaves (SCE) was orally administered (200 mg/kg body weight, per day, p.o.) and the effect on the liver lipid peroxidation (LPO), antioxidant status, protein carbonyl formation along with the histopathology of liver were evaluated after induction of adjuvant arthritis. The markers of inflammation and arthritis, such as tumor necrosis factor-α (TNF-α), interleukin 1α (IL-1α), inhibition of paw edema, along with the histological and radiographic changes in the arthritic ankle joint were studied with and without SCE administration. The result showed the presence of major phytoconstituents, such as phenolic, flavonoid and terpenoid content in SCE. HPLC analysis revealed the presence of swertiamarin and amarogentin in high concentration. The extract also showed in vitro antioxidant potential which has positive correlation with the phytoconstituents. The result of in vivo study showed elevated malondialdehyde (MDA) and carbonyl content indicative of LPO and protein oxidation, respectively, with compromised intracellular antioxidant defense system in arthritic rats, which were significantly normalized after SCE treatment. The increase in serum proinflammatory cytokines (TNF- α and IL-1α) and paw edema of arthritic rats was significantly suppressed by SCE. Histology and radiographic analysis of arthritic ankle joints indicated abnormal changes. Marked reduction in inflammation and arthritic changes were observed after treatment with SCE. The present investigation suggests that hydroethanolic extract of S. chirayita leaves exhibit potential immunomodulatory effects, which may possibly be due to boosting the intracellular antioxidant defense.