Circulating miRNA-33: a potential biomarker in patients with coronary artery disease

Background: Circulating microRNAs (miRNA) are present in body fluids in stable, cell-free form. Likewise, these miRNAs can be identified in various stages of coronary artery disease (CAD) such as inflammation, endothelial dysfunction, proliferation and atherosclerosis among others. miRNA expression levels can be identified.

Aims and objectives: To determine the expression of circulating miRNAs (miR-126, miR-92, miR-33, miR-145 and miR-155) in CAD patients of Indian origin.

Material and methods: miRNA profiling analysis in blood plasma was performed by quantitative real-time-PCR (qRT-PCR) in 60 angiographically verified subjects including 30 CAD patients and 30 age- and gender-matched controls. Association between the expression of all five circulating miRNAs and clinical characteristics of patients with CAD were analysed using Medcalc statistics. The severity of CAD was assessed using SYNTAX score (SS).

Results: Expression of plasma miR-33 increased by 2.9 folds in CAD patients than in control group (p value ≥0.002) also it was found that miR-33 expression levels in mild cases (SS: ≤22) were significantly higher than CAD controls. There was a modest negative correlation between miR-33 and total cholesterol/high density lipoprotein ratio, triglycerides and very low density lipoprotein.

Conclusion: The study reports a significant association between increased levels of plasma miR-33 and CAD. Thus, plasma miR-33 appears to be a promising non-invasive biomarker, but requires further validation in a large cohort.     

Plasmodium vivax: genetic diversity of the apical membrane antigen-1 (AMA-1) in isolates from India

Malaria parasites exhibit sequence diversity for a number of stage specific antigens. Several studies have proved that apical membrane antigen-1 (AMA-1) is an effective target for eliciting a protective immune response in humans and other experimental animals. We have investigated the sequence variation in Plasmodium vivax AMA-1 (Pv AMA-1) from different Indian isolates. This is the first study of its kind for the nearly full length Pv AMA-1 from India. Our analysis reveals greater degree of genetic diversity in Pv AMA-1 than reported so far and identifies five novel haplotypes. This is significant to establish the antigenic repertoire of isolates in a malaria endemic country like India.     

UBE1L2, a novel E1 enzyme specific for ubiquitin

UBE1 is known as the human ubiquitin-activating enzyme (E1), which activates ubiquitin in an ATP-dependent manner. Here, we identified a novel human ubiquitin-activating enzyme referred to as UBE1L2, which also shows specificity for ubiquitin. The UBE1L2 sequence displays a 40% identity to UBE1 and also contains an ATP-binding domain and an active site cysteine conserved among E1 family proteins. UBE1L2 forms a covalent link with ubiquitin in vitro and in vivo, which is sensitive to reducing conditions. In an in vitro polyubiquitylation assay, recombinant UBE1L2 could activate ubiquitin and transfer it onto the ubiquitin-conjugating enzyme UbcH5b. Ubiquitin activated by UBE1L2 could be used for ubiquitylation of p53 by MDM2 and supported the autoubiquitylation of the E3 ubiquitin ligases HectH9 and E6-AP. The UBE1L2 mRNA is most abundantly expressed in the testis, suggesting an organ-specific regulation of ubiquitin activation.     

Temporal changes in the growth, saponin content and antioxidant defense in the adventitious roots of Panax ginseng subjected to nitric oxide elicitation

Nitric oxide (NO) is a diffusible, gaseous signaling molecule. In plants, NO influences growth and development, and it can also affect plant responses to various stresses. Because NO induces root differentiation and interacts with reactive oxygen species, we examined the temporal effect of NO elicitation on root growth, saponin accumulation and antioxidant defense responses in the adventitious roots of mountain ginseng (Panax ginseng). The observations revealed that NO is involved in root growth and saponin production. Elicitation with sodium nitroprusside (SNP) activated O₂ ⁻-generating NADPH oxidase (NOX) activity, which most probably subsequently enhanced growth of adventitious roots of mountain ginseng. A severe inhibition of NOX activity and decline in dry weight of SNP elicited adventitious roots in the presence of NOX inhibitor (diphenyl iodonium, DPI), which further supports involvement of NOX in root growth. Enhanced activities of antioxidant enzymes by SNP appear to be responsible for low H₂O₂, less lipid peroxidation, and modulation of ascorbate and non-protein thiol statuses in the adventitious roots of mountain ginseng. Dry mass, saponin content and NOX activity was related with NO content present in adventitious roots of mountain ginseng.     

Single feature polymorphism discovery in rice

The discovery of nucleotide diversity captured as single feature polymorphism (SFP) by using the expression array is a high-throughput and effective method in detecting genome-wide polymorphism. The efficacy of such method was tested in rice, and the results presented in the paper indicate high sensitivity in predicting SFP. The sensitivity of polymorphism detection was further demonstrated by the fact that no biasness was observed in detecting SFP with either single or multiple nucleotide polymorphisms. The high density SFP data that can be generated quite effectively by the current method has promise for high resolution genetic mapping studies, as physical location of features are well-defined on rice genome.     

An aptasensor for rapid and sensitive detection of estrogen receptor alpha in human breast cancer

The analysis of estrogen receptor (ER) expression in breast carcinomas plays a crucial role in determining the endocrine responsiveness of tumors for systemic adjuvant therapy. Conventionally, the ER levels in breast carcinomas had been detected using the dextran-coated charcoal assay and radioimmunoassay, which are now substituted with safer and economic antibody-based assays such as immunohistochemistry (IHC) and enzyme-linked immunosorbent assay (ELISA). Despite a gold (Au) standard method, the IHC has been criticized for factors such as tissue fixation, antibody selection, and threshold staining for result interpretation that could falsify test accuracy and reproducibility. The quest for alternative methods of ER quantification in tissue samples paved the way for aptamer-based diagnostics. Previously, we have isolated a DNA aptamer against human ER alpha (ERα) using an in vitro evolution system. In this study, we developed an electrochemical sensor using the 76-nucleotide DNA ERα- aptamer for rapid, precise, and cost-effective detection of ERα expression in human breast cancer patients. The aptasensor was constructed by covalently immobilizing the thiolated ERα- aptamer onto a screen-printed Au electrode. Construction of aptasensors was confirmed through atomic force microscopy and differential pulse voltammetry measurements. A detection limit of 0.001 ng/ml was calculated for full-length ERα (66.2 kDa) in a detection time of 10 min. Analysis of the cancerous breast tissue samples using the ELISA and aptasensor methods enabled distinctive classification of samples into the categories of ER −ve, weak ER +ve, and strong ER +ve samples. The current change of this aptasensor lies within 5% after a storage of 60 days at 4°C. Further studies on a reasonably large sample size are required to realize the clinical potential of the sensor.     

Postnatal exposure to di-(2-ethylhexyl)phthalate alters cardiac insulin signaling molecules and GLUT4Ser488 phosphorylation in male rat offspring

Di-(2-ethylhexyl)phthalate (DEHP), a distinctive endocrine-disrupting chemical, is widely used as a plasticizer in a variety of consumer products. It can easily cross the placenta and enter breast milk and then it is rapidly absorbed by offspring. Since it is generally accepted that individuals are more sensitive to chemical exposure during vital developmental periods, we investigated whether DEHP exposure during lactation affects cardiac insulin signaling and glucose homeostasis in the F₁ male rat offspring at postnatal day 22 (PND22). Lactating Wistar rats were administered with DEHP (1, 10, and 100 mg/kg/d) or olive oil from lactation day 1 to 21 by oral gavage. All the male pups were perfused and killed on PND22. On the day before the killing, they were kept for fasting overnight and blood was collected. The cardiac muscle was dissected out, washed in ice-cold physiological saline repeatedly and used for the assay of various parameters. DEHP-exposed offspring had significantly lower body weight than the control. DEHP-exposed offspring showed elevated blood glucose, decreased ¹⁴C-2-deoxyglucose uptake and ¹⁴C-glucose oxidation in cardiac muscle at PND22. The concentration of upstream insulin signaling molecules such as insulin receptor subunit β (InsRβ) and insulin receptor substrate 1 (IRS1) were downregulated in DEHP-exposed offspring. However, no significant alterations were observed in protein kinase B (Akt) and Akt substrate of 160 kDa (AS160). Surprisingly, phosphorylation of IRS1 ^Tyr632 and Akt ^Ser473 were diminished. Low levels of glucose transporter type 4 (GLUT4) protein and increased GLUT4 ^Ser488 phosphorylation which decreases its intrinsic activity and translocation towards plasma membrane were also recorded. Lactational DEHP exposure predisposes F ₁ male offspring to cardiac glucometabolic disorders at PND22, which may impair cardiac function.