DNA hypomethylation activates Cdk4/6 and Atr to induce DNA replication and cell cycle arrest to constrain liver outgrowth in zebrafish.

Coordinating epigenomic inheritance and cell cycle progression is essential for organogenesis. UHRF1 connects these functions during development by facilitating maintenance of DNA methylation and cell cycle progression. Here, we provide evidence resolving the paradoxical phenotype of uhrf1 mutant zebrafish embryos which have activation of pro-proliferative genes and increased number of hepatocytes in S-phase, but the liver fails to grow. We uncover decreased Cdkn2a/b and persistent Cdk4/6 activation as the mechanism driving uhrf1 mutant hepatocytes into S-phase. This induces replication stress, DNA damage and Atr activation. Palbociclib treatment of uhrf1 mutants prevented aberrant S-phase entry, reduced DNA damage, and rescued most cellular and developmental phenotypes, but it did not rescue DNA hypomethylation, transposon expression or the interferon response. Inhibiting Atr reduced DNA replication and increased liver size in uhrf1 mutants, suggesting that Atr activation leads to dormant origin firing and prevents hepatocyte proliferation. Cdkn2a/b was downregulated pro-proliferative genes were also induced in a Cdk4/6 dependent fashion in the liver of dnmt1 mutants, suggesting DNA hypomethylation as a mechanism of Cdk4/6 activation during development. This shows that the developmental defects caused by DNA hypomethylation are attributed to persistent Cdk4/6 activation, DNA replication stress, dormant origin firing and cell cycle inhibition.

Effect of land use land cover changes on hydrological response of Punpun River basin.

The present study assessed the hydrological response of land use land cover (LULC) change on the Punpun River basin. High-resolution gridded rainfall and temperature data from the years 1995 to 2020 have been used in the Soil and Water Assessment Tool (SWAT) in the Geographic Information System (GIS) to analyze the hydrological response of the Punpun River basin and water balance components. Hydrological Response Units (HRUs) have been created for the basin. Each HRU is based on a distinct combination of soil, slope, and land use. Five SWAT models have been prepared based on the LULC of every 5-year interval to simulate the basin's hydrological response. The period selected for calibration is 1995-2015 and for validation is 2016-2020 for the modeling of daily streamflow data. The observed and simulated streamflow was checked for performance indices of coefficient of determination (R2), Nash-Sutcliffe Efficiency (NSE), and percent bias (PBIAS) on daily time steps. The results were found to be good with R2 = 0.72, NSE = 0.68, and PBIAS = 23.2 for calibration and R2 = 0.93, NSE = 0.77, and PBIAS = 19.8 for validation. The study reveals that 7.01% of evapotranspiration (ET) was increased from 1995 to 2020 with increase in agricultural area of 21.86%. It was also found that built-up area, surface runoff, and water yield have been increased by 9.14, 14.43, and 17.40%, respectively. Further, the groundwater contribution of the basin was decreased.

Feasibility of combining functional near-infrared spectroscopy with electroencephalography to identify chronic stroke responders to cerebellar transcranial direct current stimulation-a computational modeling and portable neuroimaging methodological study.

Feasibility of portable neuroimaging of cerebellar transcranial direct current stimulation (ctDCS) effects on the cerebral cortex has not been investigated vis-à-vis cerebellar lobular electric field strength. We studied functional near-infrared spectroscopy (fNIRS) in conjunction with electroencephalography (EEG) to measure changes in the brain activation at the prefrontal cortex (PFC) and the sensorimotor cortex (SMC) following ctDCS as well as virtual reality-based balance training (VBaT) before and after ctDCS treatment in 12 hemiparetic chronic stroke survivors. We performed general linear modeling (GLM) that putatively associated the lobular electric field strength with the changes in the fNIRS-EEG measures at the ipsilesional and contra-lesional PFC and SMC. Here, fNIRS-EEG measures were found in the latent space from canonical correlation analysis (CCA) between the changes in total hemoglobin (tHb) concentrations (0.01-0.07Hz and 0.07-0.13Hz bands) and log10-transformed EEG bandpower within 1-45 Hz where significant (Wilks' lambda>0.95) canonical correlations were found only for the 0.07-0.13-Hz band. Also, the first principal component (97.5% variance accounted for) of the mean lobular electric field strength was a good predictor of the latent variables of oxy-hemoglobin (O2Hb) concentrations and log10-transformed EEG bandpower. GLM also provided insights into non-responders to ctDCS who also performed poorly in the VBaT due to ideomotor apraxia. Future studies should investigate fNIRS-EEG joint-imaging in a larger cohort to identify non-responders based on GLM fitting to the fNIRS-EEG data.

Comparison of Fracture Resistance of Two Resin-based Sealers to Root Canal Walls: An In Vitro Study.

AIM AND OBJECTIVE: In the present study, the prepared roots obturated by gutta-percha/AH plus and Resilon/Epiphany were tested and compared for fracture resistance. The study also does a scanning electron microscope (SEM) evaluation of the adaptability of these obturating materials to root canal walls. MATERIALS AND METHODS: One hundred extracted mandibular premolars were decoronated and the dimensions of the roots were standardized. Each root was prepared to a size of #25 with 6% taper. Roots were gauged after preparation and those requiring more preparation were discarded. Seventy-seven prepared roots were finally selected for the study. The samples were then divided into three groups. Group I with 25 specimens was control group in which no obturation was performed, group II with 26 specimens was obturated by gutta-percha/AH plus sealer, and group III with 26 specimens was filled by Resilon/Epiphany. The method for obturation was cold lateral condensation. The samples were then stored at 100% humidity for 2 weeks. One random sample from groups II and III was subjected to SEM analysis. Groups I, II, and III were then subjected to vertical loading in Instron machine. One-way analysis of variance (ANOVA) test and Tukey's multiple comparison test were used for statistical analysis. RESULTS: Group III exhibited the maximum fracture resistance as compared to groups I and II. The least mean fracture resistance of 370.05 N was seen in group II and the maximum mean fracture resistance of 481.05 kN was observed in group III. One-way ANOVA and Tukey's multiple comparison test between groups I, II, and III, group III showed a highly significant resistance to fracture as compared to groups I and II (p < 0.0001). Scanning electron microscope microphotographs showed a better adaptation of Resilon/Epiphany as compared to gutta-percha/AH plus to the root canal. CONCLUSION: The Resilon/Epiphany on obturation of root canals creates a monoblock by penetrating inside the dentinal irregularities, which strengthens the root and provides fracture resistance. This fracture resistance was significantly higher in the present study as compared to groups I and II. CLINICAL SIGNIFICANCE: In the present study, Resilon/Epiphany when used to obturate the prepared canals showed a promising result both in terms of fracture resistance and adaptability to root canal walls. This paves a way for the use of this combination of obturating material not only to strengthen the compromised root strength in clinical scenario but also providing an increased sealing ability which will contribute to the success of root canal treatment.

Energy-dispersive X-ray Spectroscopy Analysis of Erbium, Chromium:Yttrium-scandium-gallium-garnet-treated Enamel Surfaces: An In Vitro Study.

AIM AND OBJECTIVE: The present study evaluated the effects of erbium, chromium:yttrium-scandium-gallium-garnet (Er,Cr:YSGG) laser on the weight percentage of mineral content of enamel when etched at three different power settings. MATERIALS AND METHODS: Total 20 extracted molar teeth were taken as samples. Enamel slabs were prepared by sectioning the crown from the buccal and lingual aspect with a double-sided diamond disk at slow speed. The 40 specimens were divided into four groups, i.e., control, 1 W, 2 W, and 3 W of 10 specimens each and then irradiation by Er,Cr:YSGG was done. The elements evaluated were calcium (Ca), potassium (K), magnesium (Mg), sodium (Na), and phosphorus (P). The mean weights of these minerals and the Ca:P ratio in each slab were measured by energy-dispersive X-ray spectroscopy analysis (EDAX). One-way analysis of variance (ANOVA) followed by the Turkey's test was performed with the help of critical difference (CD) or least significant difference (LSD) at 5 and 1% level of significance. RESULTS: There was no significant differences among the four groups for the five minerals and for the calcium:phosphorous ratio (p > 0.05). Photomicrographs by scanning electron microscopy observations revealed that the surfaces exposed to a 3 watt irradiation showed more roughness than those of the 1 watt and 2 watt groups. CONCLUSION: The Er,Cr:YSGG irradiation of enamel at 1 W, 2 W, and 3 W had no significant effect on the mean percentage weights of Ca, K, Mg, Na, and P or the Ca/P ratio in any group. CLINICAL SIGNIFICANCE: Constriction with conviction is the new motto of restorative dentistry. Er,Cr:YSGG not only fulfills the aim but also is proving to alter the surface properties by recrystallization and change in composition making the prepared surface caries resistant.

Lumbar canal stenosis: analyzing the role of stabilization and the futility of decompression as treatment.

OBJECTIVEThe authors report their current experience with their previously published novel form of treatment in 70 cases of lumbar canal stenosis. The treatment consisted of only fixation of the spinal segments by the transarticular screw fixation technique. No bone, ligament, osteophyte, or disc resection was done for spinal canal and neural foraminal decompression. The proposed treatment is based on the concept that vertical instability that results in telescoping of the facets on physical activity forms the nodal point of pathogenesis of lumbar canal stenosis.METHODSDuring the period June 2014 to May 2018, 70 patients presenting with the classically described symptoms of lumbar canal stenosis were treated surgically by only fixation of involved spinal segments. Apart from clinical and radiological guides, instability was diagnosed on the basis of physical observation of the status of articulation by direct manipulation of bones of the region. The operation involved transarticular insertion of 2 or 3 screws for each articulation. The Oswestry Disability Index and visual analog scale were used to assess the patients before and after surgery and at follow-up. Additionally, a personalized patient satisfaction score was used to assess the outcome of surgery.RESULTSClinical symptomatic recovery was observed in all patients in the immediate postoperative period. During the average follow-up period, 100% of patients had varying degrees of symptomatic relief. The patient satisfaction score suggested that all patients were very satisfied with the surgical procedure. The transarticular fixation technique provided strong spinal segment fixation and a reliable ground for bone arthrodesis. No patient needed any additional modality of treatment or reoperation for recurrence of symptoms.CONCLUSIONSSpinal instability is the nodal point of pathogenesis of spinal degeneration-related lumbar canal stenosis. Only fixation of the involved spinal segments is necessary-decompression by bone or soft-tissue resection is not necessary.

Rescue of neural crest-derived phenotypes in a zebrafish CHARGE model by Sox10 downregulation.

CHD7 mutations are implicated in a majority of cases of the congenital disorder, CHARGE syndrome. CHARGE, an autosomal dominant syndrome, is known to affect multiple tissues including eye, heart, ear, craniofacial nerves and skeleton and genital organs. Using a morpholino-antisense-oligonucleotide-based zebrafish model for CHARGE syndrome, we uncover a complex spectrum of abnormalities in the neural crest and the crest-derived cell types. We report for the first time, defects in myelinating Schwann cells, enteric neurons and pigment cells in a CHARGE model. We also observe defects in the specification of peripheral neurons and the craniofacial skeleton as previously reported. Chd7 morphants have impaired migration of neural crest cells and deregulation of sox10 expression from the early stages. Knocking down Sox10 in the zebrafish CHARGE model rescued the defects in Schwann cells and craniofacial cartilage. Our zebrafish CHARGE model thus reveals important regulatory roles for Chd7 at multiple points of neural crest development viz., migration, fate choice and differentiation and we suggest that sox10 deregulation is an important driver of the neural crest-derived aspects of Chd7 dependent CHARGE syndrome.

Chemical and genetic rescue of an ep300 knockdown model for Rubinstein Taybi Syndrome in zebrafish.

EP300 is a member of the EP300/CBP family of lysine acetyltransferases (KATs) with multiple roles in development and physiology. Loss of EP300/CBP activity in humans causes a very rare congenital disorder called Rubinstein Taybi Syndrome (RSTS). The zebrafish genome has two co-orthologs of lysine acetyltransferase EP300 (KAT3B) in zebrafish viz. ep300a and ep300b. Chemical inhibition of Ep300 with C646, a competitive inhibitor and morpholino-based genetic knockdown of ep300a and ep300b cause defects in embryonic development reminiscent of the human RSTS syndrome. Remarkably, overexpression of Ep300a KAT domain results in near complete rescue of the jaw development defects, a characteristic feature of RSTS in human suggesting the dispensability of the protein-interaction and DNA-binding domains for at least some developmental roles of Ep300. We also perform a chemical screen and identify two inhibitors of deacetylases, CHIC35 and HDACi III, that can partially rescue the RSTS-like phenotypes. Thus, modeling rare human genetic disorders in zebrafish allows for functional understanding of the genes involved and can also yield small molecule candidates towards therapeutic goals.

Identification of noreremophilane-based inhibitors of angiogenesis using zebrafish assays.

Noreremophilanes are a rare class of cis-hydrindanes produced by genus Ligularia herbaceous plants which are known to exhibit interesting biological activities. We synthesized cis-hydrindanes based on a naturally occurring noreremophilane scaffold using a Diels-Alder/aldol sequence and screened them for multiple biological activities using high-content zebrafish embryonic development assays. We discovered a noreremophilane that has strong anti-angiogenic effects on the developing zebrafish embryos as well as on tumor-induced angiogenesis in a zebrafish xenograft model. We synthesized several derivatives of this class of noreremophilanes and performed structure-activity relationship studies in zebrafish to identify more potent and less toxic analogs of the original structure.

Recovery from White cord Syndrome after Anterior Cervical Corpectomy and Fusion: A Case Report.

Introduction: A new neurologic deficit after spine surgery is always the biggest surgeon's nightmare. Worsening of neurology post-operatively in the absence of obvious per operative injury and with no extrinsic cause, the deficit is attributable to be caused by reperfusion injury of the spinal cord called as white cord syndrome (WCS). Hereby, we report 1-year follow-up of a case attributed as WCS after anterior cervical corpectomy with complete recovery. Case Report: A 64-year-old female patient presented with C5 - C6 tubercular lesion with extradural compression with ASIA C grade, treated with C5 - C6 corpectomy with harm cage reconstruction and tissue biopsy. Acute neurologic deterioration of both upper and lower extremities (ASIA A grade) was found 4 h after the operation upon extubation. Emergent imaging revealed no extrinsic causes. Methylprednisolone was initiated with rehabilitation therapies; her neurological status improved dramatically with complete neurological recovery at 1-year follow-up. Conclusion: New-onset neurologic deficit is always an unexpected complication. Early identification and correct treatments can avert incomplete spinal cord from permanent damage. Our experience in dealing with this patient and following up the case for nearly 1 year showed a good neurological recovery.

Portable Electromyogram-sensitive System for Assessment of Fear of Fall: Relevance of Gait Phases for Post-Stroke Patients.

Post-stroke patients often suffer from gait deficits along with Fear of Fall (FoF) that adversely affect their ambulation. The FoF has been reported to be negatively correlated with one's performance in daily life. Clinical scales, e.g., Falls Efficacy Scale are often used to assess one's FoF. Though powerful, it can suffer from subjectivity. Thus, it is important to have reliable assessment of FoF. Motivated by this, we used one's lower limb muscle activation during specific gait phases to assess one's FoF. For this, we developed a portable electromyogram-sensitive system that can synchronously measure one's muscle activation along with gait phases. We conducted an experimental study with post-stroke patients and age-matched healthy controls who walked under Dual-Task condition. We investigated the lower limb muscle (Gastrocnemius Lateralis (GM) and Tibialis Anterior (TA)) activation during Loading Response, Terminal Stance and Initial Swing phases of gait, associated with slips contributing to FoF. Results show that our system could quantify the disparity (∆) in muscle activation between the affected and unaffected sides of patients (∆ =  âˆ¼ 84% during Loading response for both GM and TA, ~32%for GM during Terminal Stance and TA during Initial Swing) which was considerably higher than that between dominant and non-dominant sides of healthy controls. This might infer reduced dynamic stability of patients leading to their FoF. Also, muscle activation could classify patients from healthy controls with 90% accuracy during Loading Response and Initial Swing phases with clinical relevance in diagnostics and monitoring rehabilitation outcomes. Clinical Relevance- This study indicated that our system has potential to be utilized as a tool for diagnostic and monitoring rehabilitation outcomes as it can quantify the residual muscle ability of the post-stroke patients during gait.

Smart Wearable Device for Quantification of Risk of Fall: Exploring Role of Gait Phases and Knee Bending Angle for Parkinson's Patients.

Gait disturbances with falls are common among patients with Parkinson's disease. Falls commonly occur from slips while walking on pathways with turns. Gait phases namely Loading Response and Terminal Stance are linked with forward and backward slips. Also, postural deformities (connected with knee joint angles) are debilitating symptoms of Parkinson's patients and are related with falls. Here, we have focused on exploring the contribution of Loading Response and Terminal Stance to risk of fall along with the relevance of postural deformity (e.g., knee bending) while an individual walked overground on pathways (with 0° and 180° turn) under dual task condition. For this, we have used a wearable device consisting of a pair of Sensored shoes and Knee Bending Angle Recorder Units. The device was used to compute Coefficient of Variation of knee bending angle during different gait phases as an indicator of one's risk of fall that corroborated with clinical measure. Clinical Relevance- A study with age and gender matched healthy and Parkinson's individuals indicated the importance of Loading Response and pathway turn while assessing risk of fall. This can serve as important pre-clinical input while designing intervention paradigms.

Antegrade Subperiosteal Temporalis Muscle Elevation and Posterior Retraction Technique Avoiding Muscle Incision for Pterional Craniotomy: A Technical Note.

Background: The current technique of pterional craniotomy involves temporalis muscle incision followed by retrograde elevation. Feasibility of antegrade temporalis muscle elevation without any direct incision over its bulk is evaluated. Objective: Incisionless "antegrade, subgaleal, subfascial, and subperiosteal elevation" of temporalis muscle preserves vascularity and muscle bulk. Posterior maneuvering of "bare" temporalis muscle bulk either above (out rolling) or under (in rolling) the scalp for pterional craniotomy is discussed. Material and Methods: Technique of antegrade, subfascial, subperiosteal elevation, and posterior rotation of temporalis muscle without incising in its bulk by "out rolling" or "in rolling" along the posterior aspect of the scalp incision was carried out in 15 cadavers and later in 50 surgical cases undergoing pterional craniotomy. Postoperatively, patients were evaluated for subgaleal collection and periorbital edema. Operated side cosmesis and temporalis muscle bulk was compared with nonoperated temporalis muscle at 6 months interval. Results: Antegrade subperiosteal dissection of temporalis muscle was possible in all cases. "In-rolling" or "out rolling" technique provided adequate surgical exposure during pterional craniotomy. Postoperative subgaleal collection and periorbital edema was prevented. Facial nerve paresis or temporalis muscle-related complications were avoided. Conclusion: Antegrade, subgaleal, subfascial, and subperiosteal dissection techniques of temporalis muscle elevation without any direct incision in its bulk enables neurovascular and muscle volume preservation. Posterior maneuvering of elevated temporalis muscle with "out rolling" or "in-rolling" technique is easy, quick, and provides adequate exposure during pterional craniotomy. Opening and closing of scalp layers without violating subgaleal space prevent postoperative subgaleal hematoma and periorbital edema.

Feasibility of Cerebellar Transcranial Direct Current Stimulation to Facilitate Goal-Directed Weight Shifting in Chronic Post-Stroke Hemiplegics.

Neurological disorder such as stroke can adversely affect one's weight-bearing symmetry leading to dysfunctional postural control. Recovery after stroke is facilitated through functionally-relevant neuroplastic modulation. Functionally-relevant cerebellum coordinates voluntary movements. Specifically, the dentate nuclei and lower limb representations (lobules VII-IX) of the cerebellum are involved in error-correction, crucial for postural control. It is postulated that cerebellar transcranial direct current stimulation (ctDCS) of the dentate nuclei and lobules VII-IX can modulate postural control in chronic stroke survivors. The objectives of this work were to (1) present a refined Virtual Reality (VR)-based balance training platform (VBaT) that can measure Center of Pressure (CoP) and (2) carry out a study to understand the implication of ctDCS stimulating the dentate nuclei (PhaseD) and lobules VII-IX (PhaseL) on the postural control of chronic stroke patients when they interacted with VBaT. Also, we investigated whether hemiplegic patients (with intact cerebellum) having Basal Ganglia (BG) infarction had any differential abilities to correct postural sway from those with no BG infarction (while shifting weight to the Affected side). Results of a single-session single-blind crossover study on randomized PhaseD and PhaseL stimulation (with an intermediate resting state bipolar bilateral ctDCS) on 12 chronic hemiplegic patients on separate days indicated differentiated findings (post stimulation) on CoP-related indices. We observed an incremental effect on one's postural control during PhaseD and inhibitory effect on the dentate nuclei during PhaseL. Clustering analysis showed that those with BG infarction demonstrated poor postural control and deficit in error correction ability irrespective of the ctDCS Phase.

Wearable Technology for Evaluation of Risk of Falls.

One's risk of fall can be quantified in terms of variability in one's gait, reflecting a loss of automatic rhythm of one's gait. In gait analysis, variability is commonly understood in terms of the fluctuation in the kinematic, kinetic, spatio-temporal, or physiological information. Here, we have focused on the estimation of knee joint angle (kinematic variable) synchronized with some of the kinetic and spatio-temporal gait parameters while an individual walked overground. Our system consisted of a pair of shoes with instrumented insoles and knee flexion/extension recorder unit having bend sensors. In addition, we have used the Coefficient of Variation for estimating the variability in the knee flexion/extension angle while walking overground as an indicator of the risk of fall. A study with healthy individuals (young and old) walking overground on pathways having 00 and 1800 turning angles indicated the feasibility of our wearable system to compute the variability in knee flexion/extension angle as an indicator of the risk of fall.

Computational investigation of potential inhibitors of novel coronavirus 2019 through structure-based virtual screening, molecular dynamics and density functional theory studies.

Despite the intensive research efforts towards antiviral drug against COVID-19, no potential drug or vaccines has not yet discovered. Initially, the binding site of COVID-19 main protease was predicted which located between regions 2 and 3. Structure-based virtual screening was performed through a hierarchal mode of elimination technique after generating a grid box. This led to the identification of five top hit molecules that were selected on the basis of docking score and visualization of non-bonding interactions. The docking results revealed that the hydrogen bonding and hydrophobic interactions are the major contributing factors in the stabilization of complexes. The docking scores were found between -7.524 and -6.711 kcal/mol indicating strong ligand-protein interactions. Amino acid residues Phe140, Leu141, Gly143, Asn142, Thr26, Glu166 and Thr190 (hydrogen bonding interactions) and Phe140, Cys145, Cys44, Met49, Leu167, Pro168, Met165, Val42, Leu27 and Ala191 (hydrophobic interactions) formed the binding pocket of COVID-19 main protease. From identified hits, ZINC13144609 and ZINC01581128 were selected for atomistic MD simulation and density functional theory calculations. MD simulation results confirm that the protein interacting with both hit molecules is stabilized in the chosen POPC lipid bilayer membrane. The presence of lowest unoccupied molecular orbital (LUMO) and highest occupied molecular orbital (HOMO) in the hydrophobic region of the hit molecules leads to favorable ligand-protein contacts. The calculated pharmacokinetic descriptors were found to be in their acceptable range and therefore confirming their drug-like properties. Hence, the present investigation can serve as the basis for designing and developing COVID-19 inhibitors. Communicated by Ramaswamy H. Sarma.

Impact of Cassia Bark Consumption on Glucose and Lipid Control in Type 2 Diabetes: An Updated Systematic Review and Meta-Analysis.

Control of diabetes is a constant challenge and natural remedies are being searched along with modern medicine. The effectiveness of cinnamon in managing it lacks consensus. Besides this, earlier trials had a variant in the type of product they used, quantity, duration, the form of molecules, etc. So, we aimed to measure the impact of cassia ground bark powder consumption, 1-2 gm/day for 90 days, in lowering plasma glucose and lipids among those with type 2 diabetes. The authors searched the PubMed, Medline, Embase, CINAHL, Clinical Key, Ovid, and Scopus databases and the Cochrane Central Register (last search December 30, 2020) with the MeSH terms and keywords of cinnamon, cassia cinnamon, Chinese cinnamon, and type 2 diabetes mellitus to conclude the effects of cassia cinnamon on diabetes based on the evidence of human clinical trials that reported at least one of the following: glycosylated hemoglobin (HbA1C), fasting blood glucose, total cholesterol, triglycerides, low-density lipoprotein (LDL), and high-density lipoprotein (HDL). Weighted mean differences were calculated by using the random-effect model of RevMan software (The Cochrane Collaboration, London, UK), and the pooled analysis found an insignificant reduction of the outcome variable (p>0.05).

Systematic Evaluation of the Effects of Toxicant Exposure on Survival in Zebrafish Embryos and Larvae.

The use of whole animal models in toxicological studies is essential for understanding the physiological responses caused by chemical exposures. However, such studies can face reproducibility challenges due to unaccounted experimental parameters that can have a marked influence on toxicological outcomes. Zebrafish embryos and larvae are a popular vertebrate animal model for studying cellular, tissue, and organ responses to toxicant exposures. Despite the popularity of this system, standardized protocols that control for the influence of various experimental parameters and culture conditions on the toxicological response in these animals have not been widely adopted, making it difficult to compare findings from different laboratories. Here, we describe a detailed approach for designing and optimizing protocols to assess the impact of chemical exposures on the development and survival of zebrafish embryos and larvae. We first describe our standard procedure to determine two key toxicological thresholds, the maximum tolerable concentration (MTC) and the lethal concentration (LC50 , defined as that in which 50% of larvae die), in response to an exposure that persists from early development through larval maturation. We then describe two protocols to systematically test how key experimental parameters, including genetic background, culture media, animal density, volume, plate material, and developmental stage in which the embryos are exposed, alter the MTC and LC50 . Finally, we provide a step-by-step guide to assess the interaction between two chemicals using this model. These protocols will guide the standardization of toxicological studies using zebrafish and maximize reproducibility. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Zebrafish embryo collection and culture, and establishment of the MTC and LC50 Basic Protocol 2: Evaluation of the impact of culture conditions on toxicant responses of zebrafish embryo and larvae Basic Protocol 3: Identification of the developmental window of sensitivity to toxicant exposure Basic Protocol 4: Testing interaction between multiple toxicants.

Lateral flow assay-based detection of long non-coding RNAs: A point-of-care platform for cancer diagnosis.

Lateral flow assay (LFA) is a flexible, simple, low-costpoint-of-care platform for rapid detection of disease-specific biomarkers. Importantly, the ability of the assay to capture the circulating bio-molecules has gained significant attention, as it offers a potential minimal invasive system for early disease diagnosis and prognosis. In the present article, we review an innovative concept of LFA-based detection of circulating long non-coding RNAs (lncRNAs), one of the key regulators of fundamental biological processes. In addition, their disease-specific expression pattern and presence in biological fluids at differential levels make them excellent biomarker candidates for cancer detection. Our article also provides an update on the requirements for developing and improving such systems and discusses the key aspects of material selection, operational concepts, principles and conceptual design. We assume that the reviewed points will be helpful to improve the diagnostic applicability of LFA based lncRNA detection in cancer diagnosis.

Point-of-care diagnostics approaches for detection of lung cancer-associated circulating miRNAs.

Circulating cell-free miRNAs (ccf-miRs) have gained significant interest as biomarkers for lung cancer (LC) diagnosis. However, the clinical application of ccf-miRs is mainly limited by time, cost, and expertise-related problems of existing detection strategies. Recently, the development of different point-of-care (POC) approaches offers useful on-site platforms, because these technologies have important features such as portability, rapid turnaround time, minimal sample requirement, and cost-effectiveness. In this review, we discuss different POC approaches for detecting ccf-miRs and highlight the utility of incorporating nanomaterials for enhanced biorecognition and signal transduction, further improving their diagnostic applicability in LC settings.