Immunotherapy: Constructive Approach for Breast Cancer Treatment.

A novel and rapid therapeutic approach is the treatment of human breast cancer by enhancing the host's immune system. In initial findings, program death one (PD-1) and program cell death ligand one (PD-L1) showed positive results towards solid tumors, but tumor relapse and drug resistance are the major concerns. Breast cancer therapy has been transformed by the advent of immune checkpoint blockades (ICBs). Triple-negative breast cancers (TNBCs) have exhibited enduring responses to clinical usage of immune checkpoint inhibitors (ICBs) like atezolizumab and pembrolizumab. Nonetheless, a notable proportion of individuals with TNBC do not experience advantages from these treatments, and there is limited comprehension of the resistance mechanisms. Another approach to overcome resistance is cancer stem cells (CSCs), as these cells are crucial for the initiation and growth of tumors in the body. Various cancer vaccines are created using stem cells (dendritic, whole cell, bacterial) and focus primarily on targeting tumor-related antigens. The ultimate objective of cancer vaccines is to immunize the patients by active artificial immunity against cancer, though. In this review, we primarily focused on existing immunotherapeutic options, immune checkpoint blockers, the latest progress in understanding the molecular mechanisms underlying resistance to immune checkpoint inhibitors (ICBs), advanced strategies to overcome resistance to ICBs, cancer stem cell antigens and molecular markers, ongoing clinical trials for BCs and cancer vaccines for breast cancer.

Phytomediated Silver Nanoparticles (AgNPs) Embellish Antioxidant Defense System, Ameliorating HLB-Diseased 'Kinnow' Mandarin Plants.

Citrus production is harmed worldwide by yellow dragon disease, also known as Huanglongbing (HLB), or citrus greening. As a result, it has negative effects and a significant impact on the agro-industrial sector. There is still no viable biocompatible treatment for Huanglongbing, despite enormous efforts to combat this disease and decrease its detrimental effects on citrus production. Nowadays, green-synthesized nanoparticles are gaining attention for their use in controlling various crop diseases. This research is the first scientific approach to examine the potential of phylogenic silver nanoparticles (AgNPs) to restore the health of Huanglongbing-diseased 'Kinnow' mandarin plants in a biocompatible manner. AgNPs were synthesized using Moringa oleifera as a reducing, capping, and stabilizing agent and characterized using different characterization techniques, i.e., UV-visible spectroscopy with a maximum average peak at 418 nm, scanning electron microscopy (SEM) with a size of 74 nm, and energy-dispersive spectroscopy (EDX), which confirmed the presence of silver ions along with different elements, and Fourier transform infrared spectroscopy served to confirm different functional groups of elements. Exogenously, AgNPs at various concentrations, i.e., 25, 50, 75, and 100 mgL-1, were applied against Huanglongbing-diseased plants to evaluate the physiological, biochemical, and fruit parameters. The findings of the current study revealed that 75 mgL-1 AgNPs were most effective in boosting the plants' physiological profiles, i.e., chl a, chl b, total chl, carotenoid content, MSI, and RWC up to 92.87%, 93.36%, 66.72%, 80.95%, 59.61%, and 79.55%, respectively; biochemical parameters, i.e., 75 mgL-1 concentration decreased the proline content by up to 40.98%, and increased the SSC, SOD, POD, CAT, TPC, and TFC content by 74.75%, 72.86%, 93.76%, 76.41%, 73.98%, and 92.85%, respectively; and fruit parameters, i.e., 75 mgL-1 concentration increased the average fruit weight, peel diameter, peel weight, juice weight, rag weight, juice pH, total soluble solids, and total sugarby up to 90.78%, 8.65%, 68.06%, 84.74%, 74.66%, 52.58%, 72.94%, and 69.69%, respectively. These findings enable us to develop the AgNP formulation as a potential citrus Huanglongbing disease management method.

Endodontic file separation and its management among dentists in Punjab, Pakistan: a cross-sectional study

The study aimed to find the incidence and awareness of endodontic instrument separation and its management among dental house officers, postgraduate trainees, demonstrators, consultants, and general dentists. Methods: This online questionnaire-based cross-sectional study was conducted with the approval of the IRB in private and public dental hospitals and dental clinics in Punjab. The authors developed the survey tool, which comprises 24 closed-ended items regarding demographics, the incidence of file separation, and awareness about its management. The data were analyzed using IBM SPSS version 24. The Chi-Square Test was used to compare percentages of categorical variables. Results: Postgraduate trainees experienced the most instrument separations (43.6%), made the most retrieval attempts (49.2%), and experienced the most secondary errors during retrieval (52.1%) (p<0.001). Around four out of ten respondents always informed the patients (39.6%) and department (41.6%) about errors. Manual files (69.8%), stainless steel files (75.8%), and short files (60.4%) were more frequently separated, and the most frequent cause was older fatigue files (57.7%). Manual files were more frequently broken in public dental institutes (p=0.003). Two-thirds of the file separations (72.5%) occurred during cleaning and shaping in the apical third of molars (65.1%), especially in mesiolingual canal (56.4%). Bypass attempt was the most common in symptomatic teeth (47.7%). Conclusions: Preventive approaches such as limiting file reuse and constructing a glide path can reduce the occurrence of file separation. Operators should be familiar with the number of uses of the instrument before fatigue and should be trained through workshops and refresher courses

Comparative Study of Antimicrobial and Antioxidant Potential of Olea ferruginea Fruit Extract and Its Mediated Selenium Nanoparticles.

Nanotechnology, the science of the recent era, has diverse applications in agriculture. Selenium (Se) is a non-metal and an essential micronutrient for animals and humans. In this study, selenium nanoparticles (SeNPs) were biosynthesized by using Olea ferruginea fruit extracts. The size, shape, chemical nature, and identification of functional groups involved in the synthesis of SeNPs were studied by UV-visible spectroscopy, Scanning Electron Microscope (SEM), and Fourier Transform Infra-Red (FTIR) spectrometry. SeNP synthesis was confirmed by an absorption peak at 258 nm by UV-visible spectroscopy. SEM showed that SeNPs were spherical, smooth, and between 60 and 80 nm in size. FTIR spectrometry confirmed the presence of terpenes, alcohols, ketones, aldehydes, and esters as well as phyto-constituents, such as alkaloids and flavonoids, that possibly act as reducing or capping agents of SeNPs in an aqueous solution of Olea ferruginea. Antimicrobial activity was examined against bacterial pathogens, such as Klebsiella pneumonia, Escherichia coli, Staphylococcus aureus, and Staphylococcus epidermitis, as well as fungal pathogens, such as Aspergillus niger and Fusarium oxysporum, by using the well-diffusion method. Antioxidant activity was observed using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, ABTs assay, and reducing power assay. At a higher concentration of 400 ppm, biosynthesized SeNPs showed an inhibition zone of 20.5 mm, 20 mm, 21 mm, and 18.5 mm against Klebsiella pneumonia, Escherichia coli, Staphylococcus aureus, and Staphylococcus epidermitis, respectively. Similarly, SeNPs also demonstrated a zone of inhibition against Aspergillus niger and Fusarium oxysporum of 17.5 and 21 mm, respectively. In contrast to Olea ferruginea fruit extracts, Olea ferruginea-mediated SeNPs demonstrated strong antimicrobial activity. By performing the DPPH, ABTs, and reducing power assay, SeNPs showed 85.2 ± 0.009, 81.12 ± 0.007, and 80.37 ± 0.0035% radical scavenging potential, respectively. The present study could contribute to the drug development and nutraceutical industries.

A Machine Learning Approach to Achieving Energy Efficiency in Relay-Assisted LTE-A Downlink System.

In recent years, Energy Efficiency (EE) has become a critical design metric for cellular systems. In order to achieve EE, a fine balance between throughput and fairness must also be ensured. To this end, in this paper we have presented various resource block (RB) allocation schemes in relay-assisted Long Term Evolution-Advanced (LTE-A) networks. Driven by equal power and Bisection-based Power Allocation (BOPA) algorithm, the Maximum Throughput (MT) and an alternating MT and proportional fairness (PF)-based SAMM (abbreviated with Authors' names) RB allocation scheme is presented for a single relay. In the case of multiple relays, the dependency of RB and power allocation on relay deployment and users' association is first addressed through a k-mean clustering approach. Secondly, to reduce the computational cost of RB and power allocation, a two-step neural network (NN) process (SAMM NN) is presented that uses SAMM-based unsupervised learning for RB allocation and BOPA-based supervised learning for power allocation. The results for all the schemes are compared in terms of EE and user throughput. For a single relay, SAMM BOPA offers the best EE, whereas SAMM equal power provides the best fairness. In the case of multiple relays, the results indicate SAMM NN achieves better EE compared to SAMM equal power and BOPA, and it also achieves better throughput fairness compared to MT equal power and MT BOPA.

The corepressor NCOR1 regulates the survival of single-positive thymocytes.

Nuclear receptor corepressor 1 (NCOR1) is a transcriptional regulator bridging repressive chromatin modifying enzymes with transcription factors. NCOR1 regulates many biological processes, however its role in T cells is not known. Here we show that Cd4-Cre-mediated deletion of NCOR1 (NCOR1 cKOCd4) resulted in a reduction of peripheral T cell numbers due to a decrease in single-positive (SP) thymocytes. In contrast, double-positive (DP) thymocyte numbers were not affected in the absence of NCOR1. The reduction in SP cells was due to diminished survival of NCOR1-null postselection TCRßhiCD69+ and mature TCRßhiCD69- thymocytes. NCOR1-null thymocytes expressed elevated levels of the pro-apoptotic factor BIM and showed a higher fraction of cleaved caspase 3-positive cells upon TCR stimulation ex vivo. However, staphylococcal enterotoxin B (SEB)-mediated deletion of Vß8+ CD4SP thymocytes was normal, suggesting that negative selection is not altered in the absence of NCOR1. Finally, transgenic expression of the pro-survival protein BCL2 restored the population of CD69+ thymocytes in NCOR1 cKOCd4 mice to a similar percentage as observed in WT mice. Together, these data identify NCOR1 as a crucial regulator of the survival of SP thymocytes and revealed that NCOR1 is essential for the proper generation of the peripheral T cell pool.

Effect of protonation on the mechanism of phosphate monoester hydrolysis and comparison with the hydrolysis of nucleoside triphosphate in biomolecular motors.

Hydrolysis of phosphate groups is a crucial reaction in living cells. It involves the breaking of two strong bonds, i.e. the OaH bond of the attacking water molecule, and the POl bond of the substrate (Oa and Ol stand for attacking and leaving oxygen atoms). Mechanism of the hydrolysis reaction can proceed either by a concurrent or a sequential mechanism. In the concurrent mechanism, the breaking of OaH and POl bonds occurs simultaneously, whereas in the sequential mechanism, the OaH and POl bonds break at different stages of the reaction. To understand how protonation affects the mechanism of hydrolysis of phosphate monoester, we have studied the mechanism of hydrolysis of protonated and deprotonated phosphate monoester at M06-2X/6-311+G**//M06-2X/6-31+G*+ZPE level of theory (where ZPE stands for zero point energy). Our calculations show that in both protonated and deprotonated cases, the breaking of the water OaH bond occurs before the breaking of the POl bond. Because the two events are not separated by a stable intermediate, the mechanism can be categorized as semi-concurrent. The overall energy barrier is 41kcalmol-1 in the unprotonated case. Most (5/6th) of this is due to the initial breaking of the water OaH bond. This component is lowered from 34 to 25kcalmol-1 by adding one proton to the phosphate. The rest of the overall energy barrier comes from the subsequent breaking of the POl bond and is not sensitive to protonation. This is consistent with previous findings about the effect of triphosphate protonation on the hydrolysis, where the equivalent protonation (on the γ-phosphate) was seen to lower the barrier of breaking the water OaH bond and to have little effect on the POl bond breaking. Hydrolysis pathways of phosphate monoester with initial breaking of the POl bond could not be found here. This is because the leaving group in phosphate monoester cannot be protonated, unlike in triphosphate hydrolysis, where protonation of the ß- and γ-phosphates had been shown to promote a mechanism where the POl bond breaks before the OaH bond does. We also point out that the charge shift due to POl bond breaking during sequential ATP hydrolysis in bio-molecular motors onsets the week unbinding of hydrolysis product that finally leads to the product release during power stroke.

A novel Cd8-cis-regulatory element preferentially directs expression in CD44hiCD62L+ CD8+ T cells and in CD8αα+ dendritic cells.

CD8 coreceptor expression is dynamically regulated during thymocyte development and is tightly controlled by the activity of at least 5 different cis-regulatory elements. Despite the detailed characterization of the Cd8 loci, the regulation of the complex expression pattern of CD8 cannot be fully explained by the activity of the known Cd8 enhancers. In this study, we revisited the Cd8ab gene complex with bioinformatics and transgenic reporter gene expression approaches to search for additional Cd8 cis-regulatory elements. This led to the identification of an ECR (ECR-4), which in transgenic reporter gene expression assays, directed expression preferentially in CD44(hi)CD62L(+) CD8(+) T cells, including innate-like CD8(+) T cells. ECR-4, designated as Cd8 enhancer E8VI, was bound by Runx/CBFß complexes and Bcl11b, indicating that E8VI is part of the cis-regulatory network that recruits transcription factors to the Cd8ab gene complex in CD8(+) T cells. Transgenic reporter expression was maintained in LCMV-specific CD8(+) T cells upon infection, although short-term, in vitro activation led to a down-regulation of E8VI activity. Finally, E8VI directed transgene expression also in CD8αα(+) DCs but not in CD8αα-expressing IELs. Taken together, we have identified a novel Cd8 enhancer that directs expression in CD44(hi)CD62L(+) CD8(+) T cells, including innate-like and antigen-specific effector/memory CD8(+) T cells and in CD8αα(+) DCs, and thus, our data provide further insight into the cis-regulatory networks that control CD8 expression.

CD4(+) T cell lineage integrity is controlled by the histone deacetylases HDAC1 and HDAC2.

Molecular mechanisms that maintain lineage integrity of helper T cells are largely unknown. Here we show histone deacetylases 1 and 2 (HDAC1 and HDAC2) as crucial regulators of this process. Loss of HDAC1 and HDAC2 during late T cell development led to the appearance of major histocompatibility complex (MHC) class II-selected CD4(+) helper T cells that expressed CD8-lineage genes such as Cd8a and Cd8b1. HDAC1 and HDAC2-deficient T helper type 0 (TH0) and TH1 cells further upregulated CD8-lineage genes and acquired a CD8(+) effector T cell program in a manner dependent on Runx-CBFß complexes, whereas TH2 cells repressed features of the CD8(+) lineage independently of HDAC1 and HDAC2. These results demonstrate that HDAC1 and HDAC2 maintain integrity of the CD4 lineage by repressing Runx-CBFß complexes that otherwise induce a CD8(+) effector T cell-like program in CD4(+) T cells.

The transcription factor MAZR preferentially acts as a transcriptional repressor in mast cells and plays a minor role in the regulation of effector functions in response to FcεRI stimulation.

Mast cells are key players in type I hypersensitivity reactions in humans and mice and their activity has to be tightly controlled. Previous studies implicated the transcription factor MAZR in the regulation of mast cell function. To study the role of MAZR in mast cells, we generated a conditional Mazr allele and crossed Mazr (F/F) mice with the Vav-iCre deleter strain, which is active in all hematopoietic cells. MAZR-null BM-derived mast cells (BMMC) were phenotypically indistinguishable from wild-type BMMCs, although the numbers of IL-3 generated Mazr (F/F) Vav-iCre BMMCs were reduced in comparison to Mazr (F/F) BMMCs, showing that MAZR is required for the efficient generation of BMMC in vitro. A gene expression analysis revealed that MAZR-deficiency resulted in the dysregulation of 128 genes, with more genes up- than down-regulated in the absence of MAZR, indicating that MAZR acts as a transcriptional repressor in mast cells. Among the up-regulated genes were the chemokines Ccl5, Cxcl10, Cxcl12, the chemokine receptor Ccr5 and the cytokine IL18, suggesting an immunoregulatory role for MAZR in mast cells. Enforced expression of MAZR in mature Mazr-deficient BMMCs rescued the altered expression pattern of some genes tested, suggesting direct regulation of these genes by MAZR. Upon FcεRI stimulation, Mazr expression was transiently down-regulated in BMMCs. However, early and late effector functions in response to FcεRI-mediated stimulation were not impaired in the absence of MAZR, with the exception of IL-6, which was slightly decreased. Taken together, out data indicate that MAZR preferentially acts as a transcriptional repressor in mast cells, however MAZR plays only a minor role in the transcriptional networks that regulate early and late effector functions in mast cells in response to FcεRI stimulation.

Cd8 enhancer E8I and Runx factors regulate CD8α expression in activated CD8+ T cells.

Cd8a and Cd8b1 coreceptor gene (Cd8) expression is tightly controlled during T-cell development by the activity of five Cd8 enhancers (E8(I)-E8(V)). Here we demonstrate a unique transcriptional program regulating CD8 expression during CD8(+) effector T-cell differentiation. The Cd8 enhancer E8(I) and Runx/core-binding factor-ß (CBFß) complexes were required for the establishment of this regulatory circuit, because E8(I)-, Runx3-, or CBFß-deficient CD8(+) T cells down-regulated CD8α expression during activation. This finding correlated with enhanced repressive histone marks at the Cd8a promoter in the absence of E8(I), and the down-regulation of CD8α expression could be blocked by treating E8(I)-, Runx3-, or CBFß-deficient CD8(+) T cells with the histone deacetylase inhibitor trichostatin A. Moreover, Runx/CBFß complexes bound the Cd8ab gene cluster in activated CD8(+) T cells, suggesting direct control of the Cd8a locus. However, CD8(+) effector T cells maintained high levels of CD8α when CBFß was conditionally deleted after activation. Thus, our data suggest an E8(I)- and Runx3/CBFß-dependent epigenetic programming of the Cd8a locus during T-cell activation, leading to Runx/CBFß complex-independent maintenance of CD8α expression in effector T cells.

Prenatal diagnosis of beta-thalassemia in Southern Punjab, Pakistan.

Pakistan has a large population of more than 150 million people with an overall carrier frequency of approximately 5.6% for beta-thalassemia. Punjab is the largest province of the country having more than 50% of the population. The state of beta-thalassemia is alarming as consanguinity is very high (>81%) and the literacy rate is low in South Punjab. A thalassemia prevention program is the need of the hour in this part of Pakistan. In this study, we initiated awareness, screening, and characterization of the mutations causing beta-thalassemia as well as a genetic counseling program mainly in the districts of Faisalabad and D.G. Khan to establish prenatal diagnosis, a facility previously unavailable in this region for disease prevention. A total of 248 unrelated transfusion-dependent children and the available members of their families were screened to characterize the mutations and identify the carriers. Genetic counseling was provided to these families and prenatal diagnosis offered. In the samples analyzed, 11 beta-thalassemia mutations and three hemoglobin variants were detected mainly by using the Monoplex and Multiplex ARMS-PCR. First-trimester prenatal diagnosis was carried out through chorionic villus sampling (CVS) in seven pregnancies at risk. As a result of our campaign, 145 carrier couples planning to have more children gave their consent to have retrospective prenatal diagnosis in every pregnancy in future. A cooperative trend and a positive attitude toward the prevention of beta-thalassemia were noticed in the families with affected children and in the general population.

Spectrum of beta-thalassemia mutations in various regions of Punjab and Islamabad, Pakistan: establishment of prenatal diagnosis.

We present here an analysis of 888 unrelated beta-thal chromosomes consisting of 444 transfusion dependent children from various regions of Punjab and Islamabad Pakistan. By using Multiplex ARMS- PCR, restriction endonuclease analysis, allele specific oligonucleotide (ASO) hybridization and sequencing, 17 beta-thal mutations and 3 Hb variants were detected in 99.5 % (884/888) of the chromosomes analyzed. First trimester prenatal diagnosis by chorionic villus sampling (CVS) was also carried out in seven pregnancies at risk of beta-thalassemia. Our results indicate that three most common mutations accounted for 86.8% of the beta-thal alleles in this region. These findings have important implications for prevention of beta-thalassemia through genetic counseling and prenatal diagnosis in this part of Pakistan.