Vanillic Acid Nanocomposite: Synthesis, Characterization Analysis, Antimicrobial, and Anticancer Potentials.

Recently, nanoparticles have received considerable attention owing to their efficiency in overcoming the limitations of traditional chemotherapeutic drugs. In our study, we synthesized a vanillic acid nanocomposite using both chitosan and silver nanoparticles, tested its efficacy against lung cancer cells, and analyzed its antimicrobial effects. We used several characterization techniques such as ultraviolet-visible spectroscopy (UV-Vis), field emission scanning electron microscopy (FESEM), energy-dispersive X-ray spectroscopy (EDAX), thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC) to determine the stability, morphological characteristics, and properties of the biosynthesized vanillic acid nanocomposites. Furthermore, the vanillic acid nanocomposites were tested for their antimicrobial effects against Escherichia coli and Staphylococcus aureus, and Candida albicans. The data showed that the nanocomposite effectively inhibited microbes, but its efficacy was less than that of the individual silver and chitosan nanoparticles. Moreover, the vanillic acid nanocomposite exhibited anticancer effects by increasing the expression of pro-apoptotic proteins (BAX, Casp3, Casp7, cyt C, and p53) and decreasing the gene expression of Bcl-2. Overall, vanillic acid nanocomposites possess promising potential against microbes, exhibit anticancer effects, and can be effectively used for treating diseases such as cancers and infectious diseases.

Time-series data prediction problem analysis through multilayered intuitionistic fuzzy sets.

For several years, time-series prediction seems to have been a popular research topic. Sales plans, ECG forecasts, meteorological circumstances, and even COVID-19 spreading projections are among its uses. These implementations have inspired several scientists to develop an optimum forecasting method; however, the modeling method varies as the implementation domain evolves. Telemetry data prediction is an important component of networking and information center control software. As a generalization of such a fuzzy system, the concept of an intuitionistic fuzzified set was created, which has proven to become a highly valuable tool in dealing with indeterminacy (hesitation) as in-network. Indeterminacy is frequently overlooked in applying fuzzified time-series prediction for no obvious cause. We introduce the concept of intuitionistic fuzzified time series within a current study to deal with non-determinism with time-series prediction. Also, it seems to be an intuitionistic fuzzified time-series prediction framework. Using time-series information, the suggested intuitionistic fuzzified time-series predicting approach employs intuitionistic fuzzified logical relationships. The suggested method's effectiveness is tested using two-time sequence data sets. By contrasting the predicted result with some other intuitionistic timing series predicting techniques utilizing root-mean-square inaccuracy and averaged predicting errors, the usefulness of the suggested intuitionistic fuzzified time-series predicting approach is demonstrated.

Emerging role of nutritional short-chain fatty acids (SCFAs) against cancer via modulation of hematopoiesis.

The understanding of gut microbiota has emerged as a significant frontier in development of strategies to maintain normal human body's homeostasis and preventing the disease development over the last decade. The composition of the gut microbiota influences the clinical benefit of immune checkpoints in patients with advanced cancer, but the mechanisms underlying this relationship are unclear. Cancer is among the leading causes of mortality worldwide. So far, there is no universal treatment for cancer and despite significant advances, a lot of improvement on cancer therapy is required. Owing to its role in preserving the host's health and maintaining cellular integrity, the human gut microbiome has recently drawn a lot of interest as a target for cancer treatment. Dietary fiber is fermented by the gut microbiota to generate short-chain fatty acids (SCFAs), such as acetate, butyrate, and propionate, which are physiologically active metabolites. SCFAs can modulate the pathophysiology of the tumor environment through various critical signaling pathways. In addition, SCFAs can bind to carcinogens and other toxic chemicals, thus facilitating their biotransformation and elimination through different excretory mechanisms. This review discusses the mechanisms of action of short-chain fatty acids in modulating hematopoiesis of various immune system cells and the resultant beneficial anti-cancer effects. It also provides future perspectives on cancer therapy.

Genetic Disruption of Guanylyl Cyclase/Natriuretic Peptide Receptor-A Triggers Differential Cardiac Fibrosis and Disorders in Male and Female Mutant Mice: Role of TGF-ß1/SMAD Signaling Pathway.

The global targeted disruption of the natriuretic peptide receptor-A (NPRA) gene (Npr1) in mice provokes hypertension and cardiovascular dysfunction. The objective of this study was to determine the mechanisms regulating the development of cardiac fibrosis and dysfunction in Npr1 mutant mice. Npr1 knockout (Npr1-/-, 0-copy), heterozygous (Npr1+/-, 1-copy), and wild-type (Npr1+/+, 2-copy) mice were treated with the transforming growth factor (TGF)-ß1 receptor (TGF-ß1R) antagonist GW788388 (2 µg/g body weight/day; ip) for 28 days. Hearts were isolated and used for real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR), Western blot, and immunohistochemical analyses. The Npr1-/- (0-copy) mice showed a 6-fold induction of cardiac fibrosis and dysfunction with markedly induced expressions of collagen-1α (3.8-fold), monocyte chemoattractant protein (3.7-fold), connective tissue growth factor (CTGF, 5.3-fold), α-smooth muscle actin (α-SMA, 6.1-fold), TGF-ßRI (4.3-fold), TGF-ßRII (4.7-fold), and phosphorylated small mothers against decapentaplegic (pSMAD) proteins, including pSMAD-2 (3.2-fold) and pSMAD-3 (3.7-fold), compared with wild-type mice. The expressions of phosphorylated extracellular-regulated kinase ERK1/2 (pERK1/2), matrix metalloproteinases-2, -9, (MMP-2, -9), and proliferating cell nuclear antigen (PCNA) were also significantly upregulated in Npr1 0-copy mice. The treatment of mutant mice with GW788388 significantly blocked the expression of fibrotic markers, SMAD proteins, MMPs, and PCNA compared with the vehicle-treated control mice. The treatment with GW788388 significantly prevented cardiac dysfunctions in a sex-dependent manner in Npr1 0-copy and 1-copy mutant mice. The results suggest that the development of cardiac fibrosis and dysfunction in mutant mice is predominantly regulated through the TGF-ß1-mediated SMAD-dependent pathway.

S-Allylcysteine (SAC) Exerts Renoprotective Effects via Regulation of TGF- ß1/Smad3 Pathway Mediated Matrix Remodeling in Chronic Renal Failure.

BACKGROUND: S-Allylcysteine (SAC), an organosulfur phytochemical sourced from aged garlic extract, is well known for its varied biomedical applications, such as anti-oxidant, anti-inflammatory, and detoxification mechanisms. Despite this, the scientific findings on the defensive impact of SAC against kidney failure (KF) are still unclear. Therefore, in the current investigation, the animal model of KF was induced by adenine in Wistar rats, and the animals were divided into four groups as control, KF induction using adenine, SAC treated KF rats for an experimental duration of 8 weeks. METHODS: KF progression was assessed by various serum and tissue markers, and the results demonstrated that the renal functions' markers, KIM-1 (kidney injury molecule-1), cystatin, NGAL (neutrophil gelatinase-associated lipocalin), were found increased in adenine-treated rats compared to control. In addition, the inflammatory markers, matrix proteins, and fibrosis signatures explicated by RT-PCR, ELISA demonstrated a profound increase. On the other hand, rats received SAC mitigated KF considerably (p < 0.001) with restored cellular functions. Besides, SAC pre-treatment abrogated the cytokines and pro-inflammatory signals (COX-2 and PGE2) in a dose-dependent manner. CONCLUSION: Furthermore, the fibrosis signaling markers mediators, such as SMAD-2,-3 were increased with associated matrix proteins. Thus, the present study substantiated that SAC possesses a significant renoprotective effect that might have been demonstrated by the inhibition of the TGF-ß1/Smad3 signaling pathway.

Lipopolysaccharide Enhances Genotoxicity by Activating GADD45G and NF-κB in Human Corneal Epithelial Cells.

As the prevalence of microbial keratitis increases, it creates an environment conducive to genotoxicity response. A potential connection between growth arrest and DNA-damage-inducible 45 gamma (GADD45G) gene expression has not been proven in the corneal epithelial cells. The aim of this study was to determine whether lipopolysaccharide (LPS) enhances genotoxicity, DNA damage, and inflammatory responses in human corneal epithelial cells (HCECs) in vitro. In a set of parameters, cytotoxicity, reactive oxygen species, mitochondrial membrane potential, DNA damage, inflammatory response, and apoptosis were assessed. LPS (1, 5, and 10 µg/mL) treated HCECs were increased reactive oxygen species formation, mitochondrial membrane depolarization, and genotoxicity in a concentration-dependent manner. Similarly, NF-κB, PARP1, and TP53 were also overexpressed in the LPS treated HCECs. 24 hours after LPS induction, micronucleus scoring, and proapoptotic factors were also increased. Among them, the GADD45G, NF-κB, and γH2AX were overexpressed both on the mRNA and protein levels in LPS (10 µg/mL) treated HCECs. In our study, we show that the GADD45G signaling can trigger genotoxic instability in HCECs exposed to LPS. Therefore, understanding the factors contributing to infectious keratitis, such as GADD45G, NF-κB, and γH2AX signaling, may help to develop antigenotoxic and anti-inflammatory therapies for corneal dystrophy and epithelial cell remodeling.

Bioactive Compounds in Oxidative Stress-Mediated Diseases: Targeting the NRF2/ARE Signaling Pathway and Epigenetic Regulation.

Oxidative stress is a pathological condition occurring due to an imbalance between the oxidants and antioxidant defense systems in the body. Nuclear factor E2-related factor 2 (NRF2), encoded by the gene NFE2L2, is the master regulator of phase II antioxidant enzymes that protect against oxidative stress and inflammation. NRF2/ARE signaling has been considered as a promising target against oxidative stress-mediated diseases like diabetes, fibrosis, neurotoxicity, and cancer. The consumption of dietary phytochemicals acts as an effective modulator of NRF2/ARE in various acute and chronic diseases. In the present review, we discussed the role of NRF2 in diabetes, Alzheimer's disease (AD), Parkinson's disease (PD), cancer, and atherosclerosis. Additionally, we discussed the phytochemicals like curcumin, quercetin, resveratrol, epigallocatechin gallate, apigenin, sulforaphane, and ursolic acid that have effectively modified NRF2 signaling and prevented various diseases in both in vitro and in vivo models. Based on the literature, it is clear that dietary phytochemicals can prevent diseases by (1) blocking oxidative stress-inhibiting inflammatory mediators through inhibiting Keap1 or activating Nrf2 expression and its downstream targets in the nucleus, including HO-1, SOD, and CAT; (2) regulating NRF2 signaling by various kinases like GSK3beta, PI3/AKT, and MAPK; and (3) modifying epigenetic modulation, such as methylation, at the NRF2 promoter region; however, further investigation into other upstream signaling molecules like NRF2 and the effect of phytochemicals on them still need to be investigated in the near future.

Organopesticides and fertility: where does the link lead to?

Organopesticides (OPs) are a group of various synthetic chemicals prevalently used in agriculture and homestead plantations. OPs were originally developed to remove insects, weeds, and other pests from agricultural fields for improving crop yields. Modern pesticides including organochlorine pesticides, organophosphorus pesticides, and amido-formyl ester are closely related to our lives. Many people are exposed to various OPs during farming practice. OPs can cause adverse effects and provoke serious impacts on normal reproductive functions of humans, resulting in loss of fertility. The effects of OPs in the reproductive system include association with fluctuation in the levels of sex hormones, delayed menstrual cycle, ovarian dysfunction, alteration in ovary weight, changes of follicle growth, altered oocyte feasibility, and changed the quality of spermatogenesis. Current literature clearly states that exposure to various OPs can impair the fertility of women and cause a high risk of reproductive potential. However, investigations on OPs exposure to woman fertility remain scarce. This review highlights effects of exposure to OPs on the fertility of occupational women and mechanisms of action involved in such effects on the reproductive function of women along with their related impacts.

Radiation-induced H3K9 methylation on E-cadherin promoter mediated by ROS/Snail axis : Role of G9a signaling during lung epithelial-mesenchymal transition.

Lung cancer patients who have undergone radiotherapy developed severe complications such as pneumonitis and fibrosis. Upon irradiation, epithelial cells acquire mesenchymal phenotype via a process called epithelial to mesenchymal transition (EMT), which plays a vital role in organ fibrosis. Several mechanisms have been studied on EMT, however, the correlation between radiation-induced EMT and epigenetic changes are not well known. In the present study, we investigated the role of histone methyltransferase G9a on radiation-induced EMT signaling. There was an increase in total global histone methylation level in irradiated epithelial cells. Western blot analysis on irradiated cells showed an increased expression of H3K9me2/3. The pre-treatment of G9a inhibitor enhanced E-cadherin expression and decreased the mesenchymal markers like N-cadherin, vimentin in the radiated group. Surprisingly, radiation-induced ROS generation and pERK1/2 levels were also inhibited by G9a inhibitor BIX01294, which is showing its antioxidant potential. The ChIP-qPCR analysis on the E-cadherin promoter suggested that G9a and Snail might have formed complex to enrich suppressive marker H3K9me2/3. On the whole, our present study suggested that 1] ROS could modify H3K9 methylation via G9a and promote radiation-induced lung EMT in Beas2B and A549 cells 2] E-cadherin promoter enrichment with heterochromatin mark H3K9me2 expression upon irradiation could be modified by regulating G9a methyltransferase.

ß-Casomorphin: A complete health perspective.

ß-Casomorphin-7 (BCM-7) is a heptapeptide dietary molecule derived from the digestion of the ß-casein of dairy and dairy products. In this review, we have covered the extensive details about BCM and its derived peptides out of the gastrointestinal and enzymatic digestion of milk and milk products, its structure and properties, and its immunological aspects related to human health among infants and adults of both genders. We have left judgment about BCM's pros and cons to the reader by describing the details in a cyclopedic perspective. In addition, a section on the possible ways to detect BCMs from their sources using proteomics, genome-based techniques, such as PCR and aptamers, and other analytical techniques equip the reader to get an idea about the details of the diagnostics available and possible applications in future. Overall, this review will provide information to the end-users of milk and milk products to enable them to make their own decisions about BCMs.

Inhibition of histone deacetylases is the major pathway mediated by astaxanthin to antagonize LPS-induced inflammatory responses in mammary epithelial cells.

Mastitis is a major inflammatory response of the mammary gland due to various pathogenic invasions and is a serious disease that affects the production yield and health status of cows. Astaxanthin (AST), a xanthophyll carotenoid, is a secondary metabolite synthesized by microalgae and yeasts that has been reported to suppress various inflammatory responses. However, the protective effect of AST on lipopolysaccharide (LPS)-induced mammary epithelial cells has not yet been reported. The present study results indicated that AST treatment markedly attenuated the oxidative stress markers and nitric oxide (NO) while improving the anti-oxidant enzymes in LPS exposed cells. On the other hand, LPS-exposed cells showed nuclear translocation of nuclear factor-κB (NF-κB) with the activation of inflammatory cytokines such as monocyte chemoattractant protein-1, tumor necrosis factor-α, interferon-γ, and interleukin-6 (IL-6). In addition, mRNA expression analysis revealed that the histone deacetylase (HDAC) -1, -2, -3, -6, -7 and pentraxin 3 (PTX3) expressions were increased in the LPS group. Furthermore, the activity of HDAC was increased to 2-fold with a significant reduction in the histone acetyltransferase activity in cells exposed to LPS. However, AST was able to inhibit the nuclear translocation of NF-κB with attenuated HDAC activity. Intriguingly, HDAC inhibition studies demonstrated that the cytokines such as IL-4, IL-8, granulocyte-mcrophage colony stimulating factor, C-reactive protein, IL-17A, and IL-22 were significantly suppressed which were upregulated in LPS treatment; while AST was found acting by improving the anti-inflammatory cytokine IL-10, and thioredoxin reductase levels. Collectively, these findings provide novel insights into the role of HDACs in regulating cellular processes involved in the pathogenesis of LPS-induced mastitis as well as the potential use of AST as a therapeutic in treatment for controlling disease progression.

Inhibitory Effect of Ursolic Acid on Ultraviolet B Radiation-Induced Oxidative Stress and Proinflammatory Response-Mediated Senescence in Human Skin Dermal Fibroblasts.

Ultraviolet radiation is an environmental carcinogenic agent that enhances inflammation and immunological reactions in the exposed human skin cells leading to oxidative photoaging of the epidermal and dermal segment. In the present study, we investigated the protective role of ursolic acid (UA) against ultraviolet B (UVB) radiation- induced photoaging an in vitro model of human skin dermal fibroblasts. UA-pretreated human skin dermal fibroblast (HDF) cells were exposed to UVB radiation to evaluated cell viability, reactive oxygen species (ROS), mitochondrial membrane potential, lipid peroxidation, antioxidant status, DNA damage, proinflammatory response, apoptotic induction, and matrix metalloproteinase (MMP) alteration. The UA pretreatment of HDFs mitigated the UVB irradiation-induced cytotoxicity, ROS generation, and mitochondrial membrane potential alteration and lipid peroxidation, depletion of antioxidant status, DNA damage, and apoptotic induction. UA pretreatment of HDFs also attenuated the UVB-induced expression of inflammatory (TNF-α and NF-κB) and apoptotic (p53, Bax, and caspase-3) and MMPs (MMP-2 and MMP-9) and enhanced the Bcl-2 protein levels in 20 µM UA treatment, when compared to concentrations. Hence, these results revealed that UA has the potential to mitigate UVB-induced extracellular damage by interfering with the ROS-mediated apoptotic induction and photoaging senescence and thus is a potential therapeutic agent to protect the skin against UVB-irradiation induced photooxidative damage.

MicroRNA Profiling of Highly Enriched Human Corneal Epithelial Stem Cells by Small RNA Sequencing.

The objective of the study was to elucidate the microRNA (miRNA) profile of an enriched human corneal epithelial stem cell (CESC) population in comparison to differentiated central corneal epithelial cells (CCECs) by small RNA sequencing. The CESCs were enriched by differential enzymatic treatment to isolate the basal limbal epithelial cells followed by laser capture microdissection of cells with nucleus to cytoplasm ratio ≥0.7, from donor tissues. Small RNA sequencing was carried out using Illumina NextSeq. 500 platform and the validation of differentially expressed miRNAs by quantitative real-time PCR (qPCR) and locked nucleic acid miRNA in-situ hybridization (LNA-ISH). The sequencing identified 62 miRNAs in CESCs and 611 in CCECs. Six miRNAs: hsa-miR-21-5p, 3168, 143-3p, 10a-5p, 150-5p and 1910-5p were found to be significantly upregulated in enriched CESCs, which was further confirmed by qPCR and LNA-ISH. The expression of hsa-miR-143-3p was exclusive to clusters of limbal basal epithelial cells. The targets of the upregulated miRNAs were predicted to be associated with signaling pathways -Wnt, PI3K-AKT, MAPK and pathways that regulate pluripotency of stem cells, cell migration, growth and proliferation. Further studies are essential to elucidate their functional role in maintenance of stemness. The findings of the study also hypothesize the inherent potential of hsa-miR-143-3p to serve as a biomarker for identifying CESCs.

Sensitive screen-printed electrodes with the colorimetric zone for simultaneous determination of mastitis and ketosis in bovine milk samples.

Bovine metabolic diseases are silent clinical conditions caused by a disturbance in normal cellular metabolic processes majorly due to the demand for nutrients, physiological stress and inflammation in lactating animals. Even though these are non-devastating diseases, conditions like mastitis and ketosis highly influence cow's lactation and fertility performance that leads to economic losses. Hence, in the present study, a combined detection including a sensitive paper-based colorimetric device for the rapid determination of somatic cells (SCC) and electrochemical detection of ß-hydroxybutyrate (BHB) in the milk samples has been developed. The working range was 1 × 105 to 2 × 106 for SCC and 0.05 to 2.5 mM BHB in which the normal and extreme cases of mastitis and ketosis values exist. The detection method is based on reaction using substrate 5-Bromo-indoxyl acetate (5BIA) and enzyme (BHB dehydrogenase) for the enzymes (esterase) and substrate (BHB) present in the milk samples during mastitis and ketosis respectively. An increase in blue color development was associated with an increase in the number of SCC; likewise, the increased voltage denoted the increase concentration of BHB in the samples. The repeatability was evaluated, and the % relative standard deviation (RSD) value <8.0 (n = 12). The limit of detection (LOD) was close to that achieved by a desktop spectrophotometer. The applicability of the developed kit demonstrated good sensitivity that can be deployed for other biological samples of interest. The proposed method provides a rapid, sensitive and promising tool for the regular monitoring of cows as a point of care tests (POC) at the field level.

Up-converting phosphor technology-based lateral flow assay for quantitative detection of ß-hydroxybutyrate in biological samples.

In the present study, a novel up-converting phosphor technology-based lateral flow (UPT-LF) assay was developed as a point of care testing method for the rapid and quantitative detection of ß-Hydroxybutyrate (BHB) in milk and serum samples. BHB is one of the most important serum molecular markers not only in ketosis but also for other metabolic disorders in bovines. The developed UPT-LF assay has high sensitivity compared with existing kits in the market for BHB quantification. The detection limit of UPT-BHB- LF assay, with a coefficient of variation (CV) < 10% was 0.01 mM, which is lower than the clinical diagnosis cutoff value of serum BHB (1 mM); while 0.1 mM is the standard level of BHB in milk samples. The relatively good stability and reproducibility of UPT-LF assay with CV<10% results for both serum and milk samples at various storage temperatures demonstrates the superiority of this method and therefore it could be used as a point of care method for the quantitative detection of BHB in the biological samples for veterinary applications. The claimed performance of this assay along with a shorter detection time makes it a viable point-of-care method for the quantitative detection of BHB in the biological samples of veterinary applications.

Exosomes: Current use and future applications.

Exosomes are endosomal-derived vesicles that play a critical role in cell-to-cell communication and are secreted in several biological fluids including serum, saliva, urine, ascites, and cerebro-spinal fluid amongst others. Exosomes are small (30-150 nm diameter) with a distinctive bilipid protein structure. They can carry and exchange various cargos between cells and are used as a non-invasive biomarker for several diseases. Exosomes are considered the best biomarkers for cancer diagnosis, owing to their unique characteristics. Here, we provide a review of the up-to-date applications of exosomes, derived from various sources, in the prognosis and diagnosis of several diseases including cancer, cardiovascular and regenerative diseases as well as, arthritis, neurological diseases, and diabetes mellitus. The role of exosomes and their applications in biomedical research and preclinical trials have also been briefly discussed.

Insights on the current status and advancement of diabetes mellitus type 2 and to avert complications: An overview.

Type 2 diabetes mellitus (T2DM) is an endocrine metabolic disorder, occurring worldwide due to aging, advancement in lifestyle by modernization. T2DM is characterized by higher levels of glucose in the blood due to unresponsive secretion of pancreatic insulin and insulin activity or altogether. T2DM is regarded as a powerful genetic susceptible disease that leads to high risk with insulin resistance and ß-cell dysfunction. To manage and overcome type 2 diabetes, physical activity, diet strategies, and other therapeutic medications along with usage of antiglycemic agents are developed and attempted appropriately. In the present review, attention has been focused on the understanding of T2DM outcomes, complications with possible management strategies, and pathophysiology of T2DM. Further, a detailed note on antiglycemic agents in use and other possible drugs of choice was discussed in the light of current preventive strategies are presented in this review.

'Biofilm Clippers'- enzyme formulation for bovine mastitic biofilm therapy.

Mastitis is one of the most important diseases that are threatening modern dairy farms. Biofilms of mastitic teat canal have serious clinical implications because of colonized pathogens having the ability to construct an extracellular polymeric substance (EPS) with increased tolerance to antimicrobials leads to difficulty in eradicating the infection. In this study, we investigated the synergistic biofilm disruptive effect of a combination of carbohydrate hydrolases targeting extracellular polysaccharides of biofilm matrix and we termed it as 'Biofilm Clippers (BC)'. Our findings demonstrate that the BC formulation exhibits intense biofilm-disrupting activity against Staphylococcus aureus biofilms. The results of the study showed that BC enables activity equivalent to physiologically achievable concentrations in disrupting biofilms of S. aureus in vitro. The synergistic anti-biofilm activities of BC on S. aureus biofilms demonstrated that the biofilm matrix is predominant of complex polysaccharides. Further, the confocal microscopic analysis demonstrates that the BC formulation is highly effective compared to the single treatment of either of the enzymes in disrupting the biofilm. To the best of our knowledge, this is the first report on the synergistic anti-biofilm activity of a class of enzyme formulation against mastitic biofilm mass. Even though a small study showed a promising effect on mastitic teat canal, further extensive investigation on a large number of bovines for mastitis therapeutic potential of this BC-derived product is now warranted.

Performance assessment of variant calling pipelines using human whole exome sequencing and simulated data.

BACKGROUND: Whole exome sequencing (WES) is a cost-effective method that identifies clinical variants but it demands accurate variant caller tools. Currently available tools have variable accuracy in predicting specific clinical variants. But it may be possible to find the best combination of aligner-variant caller tools for detecting accurate single nucleotide variants (SNVs) and small insertion and deletion (InDels) separately. Moreover, many important aspects of InDel detection are overlooked while comparing the performance of tools, particularly its base pair length. RESULTS: We assessed the performance of variant calling pipelines using the combinations of four variant callers and five aligners on human NA12878 and simulated exome data. We used high confidence variant calls from Genome in a Bottle (GiaB) consortium for validation, and GRCh37 and GRCh38 as the human reference genome. Based on the performance metrics, both BWA and Novoalign aligners performed better with DeepVariant and SAMtools callers for detecting SNVs, and with DeepVariant and GATK for InDels. Furthermore, we obtained similar results on human NA24385 and NA24631 exome data from GiaB. CONCLUSION: In this study, DeepVariant with BWA and Novoalign performed best for detecting accurate SNVs and InDels. The accuracy of variant calling was improved by merging the top performing pipelines. The results of our study provide useful recommendations for analysis of WES data in clinical genomics.

Genetic disruption of Npr1 depletes regulatory T cells and provokes high levels of proinflammatory cytokines and fibrosis in the kidneys of female mutant mice.

The present study was designed to determine the effects of gene knockout of guanylyl cyclase/natriuretic peptide receptor-A (GC-A/NPRA) on immunogenic responses affecting kidney function and blood pressure (BP) in Npr1 (coding for GC-A/NPRA)-null mutant mice. We used female Npr1 gene-disrupted (Npr1-/-, 0 copy), heterozygous (Npr1+/-, 1 copy), wild-type (Npr1+/+, 2 copy), and gene-duplicated (Npr1++/++, 4 copy) mice. Expression levels of Toll-like receptor (TLR)2/TLR4 mRNA were increased 4- to 5-fold in 1-copy mice and 6- to 10-fold in 0-copy mice; protein levels were increased 2.5- to 3-fold in 1-copy mice and 4- to 5-fold in 0-copy mice. Expression of proinflammatory cytokines and BP was significantly elevated in 1-copy and 0-copy mice compared with 2-copy and 4-copy mice. In addition, 0-copy and 1-copy mice exhibited drastic reductions in regulatory T cells (Tregs). After rapamycin treatment, Tregs were increased by 17% (P < 0.001) in 0-copy mice and 8% (P < 0.001) in 1-copy mice. Renal mRNA and protein levels of TLR2 and TLR4 were decreased by 70% in 0-copy mice and 50% in 1-copy mice. There were significantly higher levels of Tregs and very low levels of TLR2/TLR4 expression in 4-copy mice (P < 0.001). These findings indicate that the disruption of Npr1 in female mice triggers renal immunogenic pathways, which transactivate the expression of proinflammatory cytokines and renal fibrosis with elevated BP in mutant animals. The data suggest that rapamycin treatment attenuates proinflammatory cytokine expression, dramatically increases anti-inflammatory cytokines, and substantially reduces BP and renal fibrosis in mutant animals.