Single-molecule analysis of DNA base-stacking energetics using patterned DNA nanostructures.

The DNA double helix structure is stabilized by base-pairing and base-stacking interactions. However, a comprehensive understanding of dinucleotide base-stacking energetics is lacking. Here we combined multiplexed DNA-based point accumulation in nanoscale topography (DNA-PAINT) imaging with designer DNA nanostructures and measured the free energy of dinucleotide base stacking at the single-molecule level. Multiplexed imaging enabled us to extract the binding kinetics of an imager strand with and without additional dinucleotide stacking interactions. The DNA-PAINT data showed that a single additional dinucleotide base stacking results in up to 250-fold stabilization for the DNA duplex nanostructure. We found that the dinucleotide base-stacking energies vary from -0.95 ± 0.12 kcal mol-1 to -3.22 ± 0.04 kcal mol-1 for C|T and A|C base-stackings, respectively. We demonstrate the application of base-stacking energetics in designing DNA-PAINT probes for multiplexed super-resolution imaging, and efficient assembly of higher-order DNA nanostructures. Our results will aid in designing functional DNA nanostructures, and DNA and RNA aptamers, and facilitate better predictions of the local DNA structure.

Labeling approaches for DNA-PAINT super-resolution imaging.

Super-resolution imaging is becoming a commonly employed tool to visualize biological targets in unprecedented detail. DNA-PAINT is one of the single-molecule localization microscopy-based super-resolution imaging modalities allowing the ultra-high-resolution imaging with superior multiplexing capabilities. We discuss the importance of patterned DNA nanostructures in demonstrating the capabilities of DNA-PAINT and the design of various combinations of imager-docking strand pairs for imaging. Central to the implementation of DNA-PAINT imaging in a biological context is the generation of docking strand-conjugated binders against the target molecules. Several researchers have developed a variety of labelling probes for improving resolution while also providing multiplexing capabilities for the broader application of DNA-PAINT. This review provides a comprehensive summary of the repertoire of labelling probes used for DNA-PAINT in cells and the strategies implemented to chemically modify them with a docking strand.

Comparison of Standard Conservative Treatment and Early Initiation of Renal Replacement Therapy in Pregnancy-related Acute Kidney Injury: A Single-center Prospective Study.

INTRODUCTION AND OBJECTIVE: Pregnancy-related acute kidney injury (PRAKI) is one of the most important cause of maternal and fetal morbidity and mortality. Some of the reasons behind PRAKI may be due to sepsis, postpartum hemorrhage, preeclampsia, thrombotic microangiopathies (TMA), and acute fatty liver of pregnancy (AFLP). The timing of initiation of renal replacement therapy (RRT) for better patient outcome is still debatable. We conducted this study to establish whether earlier initiation of RRT in PRAKI improves fetomaternal survival compared to a more conservative approach. MATERIALS AND METHODS: This is a prospective study, where patients were screened at a multispecialty tertiary care hospital in north India for 1 year. The patients were divided into two groups: early RRT and standard AKI treatment. The demographic profile and clinical characteristics of the patients in terms of age, parity access to antenatal care, fetal outcome, urine output, hematological and biochemical profiles, RRT, duration of hospitalization, recovery of renal function, and patients survival were recorded. RESULTS: This prospective study conducted included 13 patients in the early RRT group and 23 patients in the standard group. Considering the fetal outcome, the number of fetal deaths in early RRT group were 14.29%, whereas for the standard group it was 85.71%. In all, 75% of early RRT group and 25% of standard group had normal term delivery. Probability test applied showed they were statistically significant. One maternal mortality was there in the standard group. The indices of maternal outcome in PRAKI patients were found to be statistically insignificant. CONCLUSION: The planning of RRT whether early or late does not make a difference in maternal mortality or morbidity. The fetal outcome is significantly better for patients with early RRT than conservative treatment in PRAKI. HOW TO CITE THIS ARTICLE: Banerjee A, Mehrotra G. Comparison of Standard Conservative Treatment and Early Initiation of Renal Replacement Therapy in Pregnancy-related Acute Kidney Injury: A Single-center Prospective Study. Indian J Crit Care Med 2020;24(8):688-694.

Comparison of Lung Ultrasound-based Weaning Indices with Rapid Shallow Breathing Index: Are They Helpful?

BACKGROUND AND AIMS: The diaphragm is considered the main respiratory muscle and difficulty in weaning can occur because of impaired diaphragmatic function. Hence, monitoring diaphragmatic function is important. The aim of this study is to assess the ability of various lung ultrasound (US) indices and the rapid shallow breathing index (RSBI) to predict the outcome of the weaning process and compare them with RSBI. MATERIALS AND METHODS: This was a prospective study conducted on patients admitted to critical care unit at a tertiary care hospital in north India from February 2017 to June 2017. Patients were put to spontaneous breathing trial (SBT) when they met the weaning criteria. Initial US was done immediately after putting the patient on SBT to check anatomy of the diaphragm and rule out patients according to exclusion criteria. This was followed by complete lung US (LUS) after 20 min of SBT. RESULTS: The RSBI performed better than all other parameters, with an area under the curve (AUC) of 0.996. The sensitivity and specificity is 100%. Only comparable to RSBI is the speed of diaphragmic contraction (DC) which has AUC of 0.93. All other parameters had an AUC <0.8. Moreover, the DC and LUS score are strongly positively correlated with RSBI, whereas diaphragmic excursion and diaphragmic thickness fraction (DTF %) are weakly correlated. CONCLUSION: In Intensive Care Unit, RSBI is the best clinical tool for weaning, and DC is found to be the best parameter for weaning among the US-based weaning parameters. It can even be a substitute for RSBI, in today's world of real-time monitoring methods.

Role of Correction Factor in Minimizing Errors While Calculating Electrolyte Values between Blood-gas Analyzer and Laboratory Autoanalyzer: A Comparative Study.

AIMS: Electrolytes are charged elements that play important functions in the body. They are measured by both arterial blood-gas (ABG) analyzers and autoanalyzers (AA). In this study, we tried to find out the correction factor for sodium and potassium to establish the concordance between ABG and AA values. MATERIALS AND METHODS: We prospectively studied 100 samples of patients, and for validation of the result, we applied our result on 30 patients later. 1.5 ml of blood collected in the 2.0 ml syringe preflushed with heparin and analyzed using blood-gas analyzer (ABG). Another sample was sent, to central laboratory, where serum Na+ and K+ concentrations were analyzed. Means, standard deviations, and coefficients of variation with Karl Pearson's correlation coefficients were found out. Deming regression analysis was performed and Bland-Altman plots were also constructed. RESULTS: The mean sodium and potassium were 130.27 ± 7.85 mmol/L and 3.542 ± 0.76 mmol/L using ABG and 139.28 ± 7.89 mmol/L and 4.196 ± 0.72 mmol/L using AA. Concordance between ABG and AA is done by adding the correction factor: for sodium, correction factor is 9.01, standard error = 1.113, class interval = 6.815-11.205; and for potassium (K+), correction factor is 0.654, standard error = 0.1047, class interval = 0.4475-0.8605. CONCLUSION: The instrument type and calibration methods differ in different hospitals, so it is important that each center conducts an in-hospital study to know the correction factor before installation of an ABG, and the factor should be used accordingly to minimize all errors.