Implementation of flow cytometry testing on rare matrix samples: Special considerations and best practices when the sample is unique or difficult to obtain.

The publication of Clinical and Laboratory Standards Institute's guideline H62 has provided the flow cytometry community with much-needed guidance on development and validation of flow cytometric assays (CLSI, 2021). It has also paved the way for additional exploration of certain topics requiring additional guidance. Flow cytometric analysis of rare matrices, or unique and/or less frequently encountered specimen types, is one such topic and is the focus of this manuscript. This document is the result of a collaboration subject matter experts from a diverse range of backgrounds and seeks to provide best practice consensus guidance regarding these types of specimens. Herein, we define rare matrix samples in the setting of flow cytometric analysis, address validation implications and challenges with these samples, and describe important considerations of using these samples in both clinical and research settings.

T-bet deficiency and Hic1 induction override TGF-ß-dependency in the formation of CD103+ intestine-resident memory CD8+ T cells.

Transforming growth factor ß (TGF-ß) represents a well-established signal required for tissue-resident memory T cell (TRM) formation at intestinal surfaces, regulating the expression of a large collection of genes coordinately promoting intestinal TRM differentiation. The functional contribution from each TGF-ß-controlled transcription factor is not entirely known. Here, we find that TGF-ß-induced T-bet downregulation and Hic1 induction represent two critical events during intestinal TRM differentiation. Importantly, T-bet deficiency significantly rescues intestinal TRM formation in the absence of the TGF-ß receptor. Hic1 induction further strengthens TRM maturation in the absence of TGF-ß and T-bet. Our results reveal that provision of certain TGF-ß-induced molecular events can partially replace TGF-ß signaling to promote the establishment of intestinal TRMs, which allows the functional dissection of TGF-ß-induced transcriptional targets and molecular mechanisms for TRM differentiation.

Moringin, an isothiocyanate modulates multiple cellular signalling molecules in breast cancer cells.

Prohibitin (PHB) is a pleiotropic molecule with a variety of known functions and subcellular locations. PHB's function in breast cancer is poorly understood. Herein, we report that PHB is expressed in cancer types of diverse origin including breast cancer. The cancer patients with changes in PHB were reported to have significantly reduced 'overall survival' in comparison to the cases without alterations in PHB. The expression of PHB was increased by H2O2 and also by Moringin (MG), which is an isothiocyanate derived from the seeds of Moringa oleifera. MG interacted with PHB, DRP1, and SLP2 and inhibited the growth of MCF-7 and MDAMB-231 cells. The isothiocyanate triggered apoptosis in breast cancer cells as revealed by AO/PI assay, phosphatidylserine externalization, cell cycle analysis and DAPI staining. MG induced proapoptotic proteins expression such as cytochrome c, p53, and cleaved caspase-7. Further, cell survival proteins such as survivin, Bcl-2, and Bcl-xL were suppressed. A depolarization of membrane potential suggested that the apoptosis was triggered through mitochondria. The isothiocyanate suppressed the cancer cell migration and interacted with NF-κB subunits. MG suppressed p65 nuclear translocation induced by TNF-α. The reactive oxygen species generation was also induced by the isothiocyanate in breast cancer cells. MG also modulated the expression of lncRNAs. Collectively, the functions of PHB in breast cancer growth is evident from this study. The activities of MG against breast cancer might result from its ability to modulate multiple cancer-related targets.

Comparison of image quality and diagnostic efficacy of routine clinical lumbar spine imaging at 0.55T and 1.5/3T.

PURPOSE: To compare image quality, assess inter-reader variability, and evaluate the diagnostic efficacy of routine clinical lumbar spine sequences at 0.55T compared with those collected at 1.5/3T to assess common spine pathology. METHODS: 665 image series across 70 studies, collected at 0.55T and 1.5/3T, were assessed by two neuroradiology fellows for overall imaging quality (OIQ), artifacts, and accurate visualization of anatomical features (intervertebral discs, neural foramina, spinal cord, bone marrow, and conus / cauda equina nerve roots) using a 4-point Likert scale (1 = non-diagnostic to 4 = excellent). For the 0.55T scans, the most appropriate diagnosis(es) from a picklist of common spine pathologies was selected. The mean ± SD of all scores for all features for each sequence and reader at 0.55T and 1.5/3T were calculated. Paired t-tests (p ≤ 0.05) were used to compare ratings between field strengths. The inter-reader agreement was calculated using linear-weighted Cohen's Kappa coefficient (p ≤ 0.05). Unpaired VCG analysis for OIQ was additionally employed to represent differences between 0.55T and 1.5/3T (95 % CI). RESULTS: All sequences at 0.55T were rated as acceptable (≥2) for diagnostic use by both readers despite significantly lower scores for some compared to those at 1.5/3T. While there was low inter-reader agreement on individual scores, the agreement on the diagnosis was high, demonstrating the potential of this system for detecting routine spine pathology. CONCLUSIONS: Clinical lumbar spine imaging at 0.55T produces diagnostic-quality images demonstrating the feasibility of its use in diagnosing spinal pathology, including osteomyelitis/discitis, post-surgical changes with complications, and metastatic disease.

Multiple Sclerosis Part 2: Advanced Imaging and Emerging Techniques.

Multiple advanced imaging methods for multiple sclerosis (MS) have been in investigation to identify new imaging biomarkers for early disease detection, predicting disease prognosis, and clinical trial endpoints. Multiple techniques probing different aspects of tissue microstructure (ie, advanced diffusion imaging, magnetization transfer, myelin water imaging, magnetic resonance spectroscopy, glymphatic imaging, and perfusion) support the notion that MS is a global disease with microstructural changes evident in normal-appearing white and gray matter. These global changes are likely better predictors of disability compared with lesion load alone. Emerging techniques in glymphatic and molecular imaging may improve understanding of pathophysiology and emerging treatments.

External jasmonic acid isoleucine mediates amplification of plant elicitor peptide receptor (PEPR) and jasmonate-based immune signalling.

Jasmonic acid-isoleucine (JA-Ile) is a plant defence hormone whose cellular levels are elevated upon herbivory and regulate defence signalling. Despite their pivotal role, our understanding of the rapid cellular perception of bioactive JA-Ile is limited. This study identifies cell type-specific JA-Ile-induced Ca2+ signal and its role in self-amplification and plant elicitor peptide receptor (PEPR)-mediated signalling. Using the Ca2+ reporter, R-GECO1 in Arabidopsis, we have characterized a monophasic and sustained JA-Ile-dependent Ca2+ signature in leaf epidermal cells. The rapid Ca2+ signal is independent of positive feedback by the JA-Ile receptor, COI1 and the transporter, JAT1. Microarray analysis identified up-regulation of receptors, PEPR1 and PEPR2 upon JA-Ile treatment. The pepr1 pepr2 double mutant in R-GECO1 background exhibits impaired external JA-Ile induced Ca2+ cyt elevation and impacts the canonical JA-Ile responsive genes. JA responsive transcription factor, MYC2 binds to the G-Box motif of PEPR1 and PEPR2 promoter and activates their expression upon JA-Ile treatment and in myc2 mutant, this is reduced. External JA-Ile amplifies AtPep-PEPR pathway by increasing the AtPep precursor, PROPEP expression. Our work shows a previously unknown non-canonical PEPR-JA-Ile-Ca2+ -MYC2 signalling module through which plants sense JA-Ile rapidly to amplify both AtPep-PEPR and jasmonate signalling in undamaged cells.

Differential cellular localization of DNA gyrase and topoisomerase IB in response to DNA damage in Deinococcus radiodurans.

Topoisomerases are crucial enzymes in genome maintenance that modulate the topological changes during DNA metabolism. Deinococcus radiodurans, a Gram-positive bacterium is characterized by its resistance to many abiotic stresses including gamma radiation. Its multipartite genome encodes both type I and type II topoisomerases. Time-lapse studies using fluorescently tagged topoisomerase IB (drTopoIB-RFP) and DNA gyrase (GyrA-RFP) were performed to check the dynamics and localization with respect to DNA repair and cell division under normal and post-irradiation growth conditions. Results suggested that TopoIB and DNA gyrase are mostly found on nucleoid, highly dynamic, and show growth phase-dependent subcellular localization. The drTopoIB-RFP was also present at peripheral and septum regions but does not co-localize with the cell division protein, drFtsZ. On the other hand, DNA gyrase co-localizes with PprA a pleiotropic protein involved in radioresistance, on the nucleoid during the post-irradiation recovery (PIR). The topoIB mutant was found to be sensitive to hydroxyurea treatment, and showed more accumulation of single-stranded DNA during the PIR, compared to the wild type suggesting its role in DNA replication stress. Together, these results suggest differential localization of drTopoIB-RFP and GyrA-RFP in D. radiodurans and their interaction with PprA protein, emphasizing the functional significance and role in radioresistance.

Quality assessment of routine brain imaging at 0.55 T: initial experience in a clinical workflow.

The purpose of this study was to assess the quality of clinical brain imaging in healthy subjects and patients on an FDA-approved commercial 0.55 T MRI scanner, and to provide information about the feasibility of using this scanner in a clinical workflow. In this IRB-approved study, brain examinations on the scanner were prospectively performed in 10 healthy subjects (February-April 2022) and retrospectively derived from 44 patients (February-July 2022). Images collected using the following pulse sequences were available for assessment: axial DWI (diffusion-weighted imaging), apparent diffusion coefficient maps, 2D axial fluid-attenuated inversion recovery images, axial susceptibility-weighted images (both magnitude and phase), sagittal T1 -weighted (T1w) Sampling Perfection with Application Optimized Contrast images, sagittal T1w MPRAGE (magnetization prepared rapid gradient echo) with contrast enhancement, axial T1w turbo spin echo (TSE) with and without contrast enhancement, and axial T2 -weighted TSE. Two readers retrospectively and independently evaluated image quality and specific anatomical features in a blinded fashion on a four-point Likert scale, with a score of 1 being unacceptable and 4 being excellent, and determined the ability to answer the clinical question in patients. For each category of image sequences, the mean, standard deviation, and percentage of unacceptable quality images (<2) were calculated. Acceptable (rating ≥ 2) image quality was achieved at 0.55 T in all sequences for patients and 85% of the sequences for healthy subjects. Radiologists were able to answer the clinical question in all patients scanned. In total, 50% of the sequences used in patients and about 60% of the sequences used in healthy subjects exhibited good (rating ≥ 3) image quality. Based on these findings, we conclude that diagnostic quality clinical brain images can be successfully collected on this commercial 0.55 T scanner, indicating that the routine brain imaging protocol may be deployed on this system in the clinical workflow.

Quantitative approaches for including equity in risk and resilience infrastructure planning analyses.

Risk and resilience assessments for critical infrastructure focus on myriad objectives, from natural hazard evaluations to optimizing investments. Although research has started to characterize externalities associated with current or possible future states, incorporation of equity priorities at project inception is increasingly being recognized as critical for planning related activities. However, there is no standard methodology that guides development of equity-informed quantitative approaches for infrastructure planning activities. To address this gap, we introduce a logic model that can be tailored to capture nuances about specific geographies and community priorities, effectively incorporating them into different mathematical approaches for quantitative risk assessments. Specifically, the logic model uses a graded, iterative approach to clarify specific equity objectives as well as inform the development of equations being used to support analysis. We demonstrate the utility of this framework using case studies spanning aviation fuel, produced water, and microgrid electricity infrastructures. For each case study, the use of the logic model helps clarify the ways that local priorities and infrastructure needs are used to drive the types of data and quantitative methodologies used in the respective analyses. The explicit consideration of methodological limitations (e.g., data mismatches) and stakeholder engagements serves to increase the transparency of the associated findings as well as effectively integrate community nuances (e.g., ownership of assets) into infrastructure assessments. Such integration will become increasingly important to ensure that planning activities (which occur throughout the lifecycle of the infrastructure projects) lead to long-lasting solutions to meet both energy and sustainable development goals for communities.

Comparison of flowcytometry-based scoring system for the diagnosis of early T precursor-acute lymphoblastic leukemia.

BACKGROUND: Early T cell precursor-acute lymphoblastic leukemia (ETP-ALL) is a hematolymphoid malignancy where the blasts demonstrate T cell differentiation markers along with stem cell and myeloid antigen expression. The differential diagnosis of ETP-ALL from non-ETP ALL and mixed phenotype acute leukemia is often challenging due to its overlapping immunophenotypic picture with co-expression of myeloid antigens. In this study, we endeavored to describe the immune-phenotype profile of ETP-ALL in our patients and compared the utility of four different scoring systems for better discrimination of these entities. METHODS: This retrospective analysis included 31 ETP-ALL out of 860 acute leukemia cases consecutively diagnosed at the two tertiary care centers. Flowcytometry-based immunophenotype was reviewed for all the cases, and the utility of four flow-based objective scorings was assessed for the diagnosis of ETP-ALL. Receiver operating curves were drawn to compare the different flow-based scoring systems. RESULTS: The prevalence of ETP-ALL was 40% (n = 31/77 T-ALL) in our study group, comprised mainly of adults with a median age of 20 years. The five-marker scoring system had the maximum area under the curve, followed by the seven-marker scoring system. A cut-off of ≥2.5 was more specific (sensitivity: 91%; specificity: 100%), while a score of ≥1.5 was more sensitive but slightly less specific (sensitivity: 94%, specificity: 96%). CONCLUSION: The WHO criteria for the diagnosis of ETP-ALL should be followed across all laboratories to avoid confusion and for better treatment stratification. Flow-based scoring systems can be objectively employed for better detection of cases.

A Comparative Analysis of Multidimensional COVID-19 Poverty Determinants: An Observational Machine Learning Approach.

Poverty is a glaring issue in the twenty-first century, even after concerted efforts of organizations to eliminate the same. Predicting poverty using machine learning can offer practical models for facilitating the process of elimination of poverty. This paper uses Multidimensional Poverty Index Data from the Oxford Poverty and Human Development Initiative across the years 2019 and 2021 to make predictions of multidimensional poverty before and during the pandemic. Several poverty indicators under health, education and living standards are taken into consideration. The work implements several data analysis techniques like feature correlation and selection, and graphical visualizations to answer research questions about poverty. Various machine learning, such as Multiple Linear Regression, Decision Tree Regressor, Random Forest Regressor, XGBoost, AdaBoost, Gradient Boosting, Linear Support Vector Regressor (SVR), Ridge Regression, Lasso Regression, ElasticNet Regression, and K-Nearest Neighbor Regression algorithm, have been implemented to predict poverty across four datasets on a national and a subnational level. Regularization is used to increase the performance of the models, and cross-validation is used for estimation. Through a rigorous analysis and comparison of different models, this work identifies important poverty determinants and concludes that overall, Ridge Regression model performs the best with the highest R 2 score.

FtsZ phosphorylation brings about growth arrest upon DNA damage in Deinococcus radiodurans.

The polymerization/depolymerization dynamics of FtsZ play a pivotal role in cell division in the majority of the bacteria. Deinococcus radiodurans, a radiation-resistant bacterium, shows an arrest of growth in response to DNA damage with no change in the level of FtsZ. This bacterium does not deploy LexA/RecA type of DNA damage response and cell cycle regulation, and its genome does not encode SulA homologues of Escherichia coli, which attenuate FtsZ functions in response to DNA damage in other bacteria. A radiation-responsive Ser/Thr quinoprotein kinase (RqkA), characterized for its role in radiation resistance in this bacterium, could phosphorylate several cognate proteins, including FtsZ (drFtsZ) at Serine 235 (S235) and Serine 335 (S335) residues. Here, we reported the detailed characterization of S235 and S335 phosphorylation effects in the regulation of drFtsZ functions and demonstrated that the phospho-mimetic replacements of these residues in drFtsZ had grossly affected its functions that could result in cell cycle arrest in response to DNA damage in D. radiodurans. Interestingly, the phospho-ablative replacements were found to be nearly similar to drFtsZ, whereas the phospho-mimetic mutant lost the wild-type protein's signature characteristics, including its dynamics under normal conditions. The kinetics of post-bleaching recovery for drFtsZ and phospho-mimetic mutant were nearly similar at 2 h post-irradiation recovery but were found to be different under normal conditions. These results highlighted the role of S/T phosphorylation in the regulation of drFtsZ functions and cell cycle arrest in response to DNA damage, which is demonstrated for the first time, in any bacteria.

A DM-ELM based classifier for EEG brain signal classification for epileptic seizure detection.

Epilepsy is one of the dreaded conditions that had taken billions of people under its cloud worldwide. Detecting the seizure at the correct time in an individual is something that medical practitioners focus in order to help people save their lives. Analysis of the Electroencephalogram (EEG) signal from the scalp area of the human brain can help in detecting the seizure beforehand. This paper presents a novel classification technique to classify EEG brain signals for epilepsy identification based on Discrete Wavelet Transform and Moth Flame Optimization-based Extreme Learning Machine (DM-ELM). ELM is a very popular machine learning method based on Neural Networks (NN) where the model is trained rigorously to get the minimized error rate and maximized accuracy. Here we have used several experimental evaluations to compare the performance of basic ELM and DM-ELM and it has been experimentally proved that DM-ELM outperforms basic ELM but with few time constraints.

Enhanced Tumor Accumulation of Multimodal Magneto-Plasmonic Nanoparticles via an Implanted Micromagnet-Assisted Delivery Strategy.

One of the major shortcomings of nano carriers-assisted cancer therapeutic strategies continues to be the inadequate tumor penetration and retention of systemically administered nanoformulations and its off-target toxicity. Stromal parameters-related heterogeneity in enhanced permeability and retention effect and physicochemical properties of the nanoformulations immensely contributes to their poor tumor extravasation. Herein, a novel tumor targeting strategy, where an intratumorally implanted micromagnet can significantly enhance accumulation of magneto-plasmonic nanoparticles (NPs) at the micromagnet-implanted tumor in bilateral colorectal tumor models while limiting their off-target accumulation, is demonstrated. To this end, novel multimodal gold/iron oxide NPs comprised of an array of multifunctional moieties with high therapeutic, sensing, and imaging potential are developed. It is also discovered that cancer cell targeted NPs in combination with static magnetic field can selectively induce cancer cell death. A multimodal caspase-3 nanosensor is also developed for real-time visualization of selective induction of apoptosis in cancer cells. In addition, the photothermal killing capability of these NPs in vitro is evaluated, and their potential for enhanced photothermal ablation in tissue samples is demonstrated. Building on current uses of implantable devices for therapeutic purposes, this study envisions the proposed micromagnet-assisted NPs delivery approach may be used to accelerate the clinical translation of various nanoformulations.

TGF-ß-dependent lymphoid tissue residency of stem-like T cells limits response to tumor vaccine.

TGF-ß signaling is necessary for CD8+ T cell differentiation into tissue resident memory T cells (TRM). Although higher frequency of CD8+ TRM cells in the tumor microenvironment is associated with better prognosis, TGF-ß-blockade typically improves rather than worsens outcomes. Here we show that in a mouse melanoma model, in the tumor-draining lymph nodes (TDLN) rather than in the tumors themselves, stem-like CD8+ T cells differentiate into TRMs in a TGF-ß and tumor antigen dependent manner. Following vaccination against a melanoma-specific epitope, most tumour-specific CD8+ T cells are maintained in a stem-like state, but a proportion of cells lost TRM status and differentiate into CX3CR1+ effector CD8+ T cells in the TDLN, which are subsequently migrating into the tumours. Disruption of TGF-ß signaling changes the dynamics of these developmental processes, with the net result of improving effector CD8+ T cell migration into the tumours. In summary, TDLN stem-like T cells transiently switch from a TGF-ß-dependent TRM differentiation program to an anti-tumor migratory effector development upon vaccination, which transition can be facilitated by targeted TGF-ß blockade.

FtsK, a DNA Motor Protein, Coordinates the Genome Segregation and Early Cell Division Processes in Deinococcus radiodurans.

Filament temperature-sensitive mutant K (FtsK)/SpoIIIE family proteins are DNA translocases known as the fastest DNA motor proteins that use ATP for their movement on DNA. Most of the studies in single chromosome-containing bacteria have established the role of FtsK in chromosome dimer resolution (CDR), connecting the bacterial chromosome segregation process with cell division. Only limited reports, however, are available on the interdependent regulation of genome segregation and cell division in multipartite genome harboring (MGH) bacteria. In this study, for the first time, we report the characterization of FtsK from the radioresistant MGH bacterium Deinococcus radiodurans R1 (drFtsK). drFtsK shows the activity characteristics of a typical FtsK/SpoIIIE/Tra family. It stimulates the site-specific recombination catalyzed by Escherichia coli tyrosine recombinases. drFtsK interacts with various cell division and genome segregation proteins of D. radiodurans. Microscopic examination of different domain deletion mutants of this protein reveals alterations in cellular membrane architecture and nucleoid morphology. In vivo localization studies of drFtsK-RFP show that it forms multiple foci on nucleoid as well as on the membrane with maximum density on the septum. drFtsK coordinates its movement with nucleoid separation. The alignment of its foci shifts from old to new septum indicating its cellular dynamics with the FtsZ ring during the cell division process. Nearly, similar positional dynamicity of FtsK was observed in cells recovering from gamma radiation exposure. These results suggest that FtsK forms a part of chromosome segregation, cell envelope, and cell division machinery in D. radiodurans. IMPORTANCE Deinococcus radiodurans show extraordinary resistance to gamma radiation. It is polyploid and harbors a multipartite genome comprised of 2 chromosomes and 2 plasmids, packaged in a doughnut-shaped toroidal nucleoid. Very little is known about how the tightly packed genome is accurately segregated and the next divisional plane is determined. Filament temperature-sensitive mutant K (FtsK), a multifunctional protein, helps in pumping the septum-trapped DNA in several bacteria. Here, we characterized FtsK of D. radiodurans R1 (drFtsK) for the first time and showed it to be an active protein. The absence of drFtsK causes many defects in morphology at both cellular and nucleoid levels. The compact packaging of the deinococcal genome and cell membrane formation is hindered in ftsK mutants. In vivo drFtsK is dynamic, forms foci on both nucleoid and septum, and coordinates with FtsZ for the next cell division. Thus, drFtsK role in maintaining the normal genome phenotype and cell division in D. radiodurans is suggested.

TGF-ß promotes stem-like T cells via enforcing their lymphoid tissue retention.

Stem-like CD8+ T cells sustain the antigen-specific CD8+ T cell response during chronic antigen exposure. However, the signals that control the maintenance and differentiation of these cells are largely unknown. Here, we demonstrated that TGF-ß was essential for the optimal maintenance of these cells and inhibited their differentiation into migratory effectors during chronic viral infection. Mechanistically, stem-like CD8+ T cells carried a unique expression pattern of α4 integrins (i.e., α4ß1hi and α4ß7lo) controlled by TGF-ß. In the absence of TGF-ß signaling, greatly enhanced expression of migration-related markers, including altered expression of α4 integrins, led to enhanced egress of stem-like CD8+ T cells into circulation accompanied by further differentiation into transitional states. Blocking α4 integrin significantly promoted their lymphoid tissue retention and therefore partially rescued the defective maintenance of Tcf-1+ subset in the absence of TGF-ß signaling. Thus, TGF-ß promotes the maintenance and inhibits the further differentiation of stem-like T cells at least partially via enforcing their lymphoid tissue residency.

Usefulness of Leucocyte Cell Population Data by Sysmex XN1000 Hematology Analyzer in Rapid Identification of Acute Leukemia.

Leukocyte cell population data (CPD) generated by hematology auto analyzers are reported to be useful in screening of sepsis patients. However, there is a paucity of literature highlighting the utility of CPD in screening of acute leukemias (AL). Leucocyte CPD obtained by Sysmex XN1000 hematology analyzer from 210 cases of ALs [22 acute promyelocytic leukemia (APL), 79 non-APL acute myeloid leukemia (non-APL-AML) and 109 acute lymphoblastic leukemia (ALL)] were compared with 100 healthy and 52 reactive controls. Receiver operator curves were drawn to determine the cut-off values of individual parameters. The regression equations combining the best parameters were then formulated to calculate a cut-off value for discrimination among AL subgroups and controls. Acute leukemias showed significant differences (p < 0.05) in various CPD parameters compared to control subjects. A combination of best CPD parameters discriminated ALs from healthy controls (cut off; 0.443, sensitivity of 94% and specificity of 91%), ALs from reactive controls (cut off; 0.576, sensitivity; 97%, specificity; 92%), APL from non-APL-AML (cut off; 0.174, sensitivity of 91% and specificity of 67%), and AML from ALL (cut off; 1.338, sensitivity; 86.1%, specificity; 75%). The CPD from Sysmex XN 1000 analyzer could be a useful tool in screening and lineage characterization of acute leukemias; particularly at centers where high-end technical expertise is still not available. Supplementary Information: The online version contains supplementary material available at 10.1007/s12288-021-01488-9.

Piperlongumine, a piper alkaloid, enhances the efficacy of doxorubicin in breast cancer: involvement of glucose import, ROS, NF-κB and lncRNAs.

Piperlongumine (PL, piplartine) is an alkaloid derived from the Piper longum L. (long pepper) roots. Originally discovered in 1961, the biological activities of this molecule against some cancer types was reported during the last decade. Whether PL can synergize with doxorubicin and the underlying mechanism in breast cancer remains elusive. Herein, we report the activities of PL in numerous breast cancer cell lines. PL reduced the migration and colony formation by cancer cells. An enhancement in the sub-G1 population, reduction in the mitochondrial membrane potential, chromatin condensation, DNA laddering and suppression in the cell survival proteins was observed by the alkaloid. Further, PL induced ROS generation in breast cancer cells. While TNF-α induced p65 nuclear translocation, PL suppressed the translocation in cancer cells. The expression of lncRNAs such as MEG3, GAS5 and H19 were also modulated by the alkaloid. The molecular docking studies revealed that PL can interact with both p65 and p50 subunits. PL reduced the glucose import and altered the pH of the medium towards the alkaline side. PL also suppressed the expression of glucose and lactate transporter in breast cancer cells. In tumor bearing mouse model, PL was found to synergize with doxorubicin and reduced the size, volume and weight of the tumor. Overall, the effects of doxorubicin in cancer cells are enhanced by PL. The modulation of glucose import, NF-κB activation and lncRNAs expression may have contributory role for the activities of PL in breast cancer.

2D Materials-Based Aptamer Biosensors: Present Status and Way Forward.

Current advances in constructing functional nanomaterials and elegantly designed nanostructures have opened up new possibilities for the fabrication of viable field biosensors. Two-dimensional materials (2DMs) have fascinated much attention due to their chemical, optical, physicochemical, and electronic properties. They are ultrathin nanomaterials with unique properties such as high surface-to-volume ratio, surface charge, shape, high anisotropy, and adjustable chemical functionality. 2DMs such as graphene-based 2D materials, Silicate clays, layered double hydroxides (LDHs), MXenes, transition metal dichalcogenides (TMDs), and transition metal oxides (TMOs) offer intensified physicochemical and biological functionality and have proven to be very promising candidates for biological applications and technologies. 2DMs have a multivalent structure that can easily bind to single-stranded DNA/RNA (aptamers) through covalent, non-covalent, hydrogen bond, and π-stacking interactions, whereas aptamers have a small size, excellent chemical stability, and low immunogenicity with high affinity and specificity. This review discussed the potential of various 2D material-based aptasensor for diagnostic applications, e.g., protein detection, environmental monitoring, pathogens detection, etc.