Sero-prevalence and risk factors for Severe Acute Respiratory Syndrome Coronavirus 2 infection in women and children in a rural district of Bangladesh: A cohort study.

Background: Bangladesh reported its first COVID-19 case on March 8, 2020. Despite lockdowns and promoting behavioural interventions, as of December 31, 2021, Bangladesh reported 1.5 million confirmed cases and 27 904 COVID-19-related deaths. To understand the course of the pandemic and identify risk factors for SARs-Cov-2 infection, we conducted a cohort study from November 2020 to December 2021 in rural Bangladesh. Methods: After obtaining informed consent and collecting baseline data on COVID-19 knowledge, comorbidities, socioeconomic status, and lifestyle, we collected data on COVID-like illness and care-seeking weekly for 54 weeks for women (n = 2683) and their children (n = 2433). Between March and July 2021, we tested all participants for SARS-CoV-2 antibodies using ROCHE's Elecsys® test kit. We calculated seropositivity rates and 95% confidence intervals (95% CI) separately for women and children. In addition, we calculated unadjusted and adjusted relative risk (RR) and 95% CI of seropositivity for different age and risk groups using log-binomial regression models. Results: Overall, about one-third of women (35.8%, 95% CI = 33.7-37.9) and one-fifth of children (21.3%, 95% CI = 19.2-23.6) were seropositive for SARS-CoV-2 antibodies. The seroprevalence rate doubled for women and tripled for children between March 2021 and July 2021. Compared to women and children with the highest household wealth (HHW) tertile, both women and children from poorer households had a lower risk of infection (RR, 95% CI for lowest HHW tertile women (0.83 (0.71-0.97)) and children (0.75 (0.57-0.98)). Most infections were asymptomatic or mild. In addition, the risk of infection among women was higher if she reported chewing tobacco (RR = 1.19,95% CI = 1.03-1.38) and if her husband had an occupation requiring him to work indoors (RR = 1.16, 95% CI = 1.02-1.32). The risk of infection was higher among children if paternal education was >5 years (RR = 1.37, 95% CI = 1.10-1.71) than in children with a paternal education of ≤5 years. Conclusions: We provided prospectively collected population-based data, which could contribute to designing feasible strategies against COVID-19 tailored to high-risk groups. The most feasible strategy may be promoting preventive care practices; however, collecting data on reported practices is inadequate. More in-depth understanding of the factors related to adoption and adherence to the practices is essential.

The use of viral vectors to promote repair after spinal cord injury.

Spinal cord injury (SCI) is a devastating event that can permanently disrupt multiple modalities. Unfortunately, the combination of the inhibitory environment at a central nervous system (CNS) injury site and the diminished intrinsic capacity of adult axons for growth results in the failure for robust axonal regeneration, limiting the ability for repair. Delivering genetic material that can either positively or negatively modulate gene expression has the potential to counter the obstacles that hinder axon growth within the spinal cord after injury. A popular gene therapy method is to deliver the genetic material using viral vectors. There are considerations when deciding on a viral vector approach for a particular application, including the type of vector, as well as serotypes, and promoters. In this review, we will discuss some of the aspects to consider when utilizing a viral vector approach to as a therapy for SCI. Additionally, we will discuss some recent applications of gene therapy to target extrinsic and/or intrinsic barriers to promote axon regeneration after SCI in preclinical models. While still in early stages, this approach has potential to treat those living with SCI.

Immunobiology of cancer-associated fibroblasts in the context of radiotherapy.

Radiotherapy (RT) still represents a mainstay of treatment in clinical oncology. Traditionally, the effectiveness of radiotherapy has been attributed to the killing potential of ionizing radiation (IR) over malignant cells, however, it has become clear that therapeutic efficacy of RT also involves activation of innate and adaptive anti-tumor immune responses. Therapeutic irradiation of the tumor microenvironment (TME) provokes profound cellular and biological reconfigurations which ultimately may influence immune recognition. As one of the major constituents of the TME, cancer-associated fibroblasts (CAFs) play central roles in cancer development at all stages and are recognized contributors of tumor immune evasion. While some studies argue that RT affects CAFs negatively through growth arrest and impaired motility, others claim that exposure of fibroblasts to RT promotes their conversion into a more activated phenotype. Nevertheless, despite the well-described immunoregulatory functions assigned to CAFs, little is known about the interplay between CAFs and immune cells in the context of RT. In this review, we go over current literature on the effects of radiation on CAFs and the influence that CAFs have on radiotherapy outcomes, and we summarize present knowledge on the transformed cellular crosstalk between CAFs and immune cells after radiation.

Converting an Anti-Mouse CD4 Monoclonal Antibody into an scFv Positron Emission Tomography Imaging Agent for Longitudinal Monitoring of CD4+ T Cells.

Immuno-positron emission tomography (PET), a noninvasive imaging modality, can provide a dynamic approach for longitudinal assessment of cell populations of interest. Transformation of mAbs into single-chain variable fragment (scFv)-based PET imaging agents would allow noninvasive tracking in vivo of a wide range of possible targets. We used sortase-mediated enzymatic labeling in combination with PEGylation to develop an anti-mouse CD4 scFv-based PET imaging agent constructed from an anti-mouse CD4 mAb. This anti-CD4 scFv can monitor the in vivo distribution of CD4+ T cells by immuno-PET. We tracked CD4+ and CD8+ T cells in wild-type mice, in immunodeficient recipients reconstituted with monoclonal populations of OT-II and OT-I T cells, and in a B16 melanoma model. Anti-CD4 and -CD8 immuno-PET showed that the persistence of both CD4+ and CD8+ T cells transferred into immunodeficient mice improved when recipients were immunized with OVA in CFA. In tumor-bearing animals, infiltration of both CD4+ and CD8+ T cells increased as the tumor grew. The approach described in this study should be readily applicable to convert clinically useful Abs into the corresponding scFv PET imaging agents.

Occurrence, fates, and carcinogenic risks of substituted polycyclic aromatic hydrocarbons in two coking wastewater treatment systems.

This paper reports for the first time the occurrence, fates, and carcinogenic risks of 20 substituted polycyclic aromatic hydrocarbons (SPAHs) and 16 priority PAH species in two coking wastewater treatment plants (WWTPs) (plant E and central WWTP). The measured total concentrations of PAHs and SPAHs in raw wastewater of coking plant E were 3700 and 1200 µg·L-1, respectively, with naphthalene (1400 µg·L-1), and fluoranthene (353 µg·L-1) as dominant PAH species and 2-methylnaphthalene (167 µg·L-1), anthraquinone (133 µg·L-1), and 1-methylnaphthalene (132 µg·L-1) as dominant SPAHs. For the 11 methyl-PAHs (MPAHs), 4 oxygenated-PAHs (OPAHs), and 5 nitrated-PAHs (NPAHs) investigated, the biological wastewater treatment process removed 98.6% MPAHs, 83.9% OPAHs, and 89.1% NPAHs. Mass balance analysis result revealed that transformation was the major mechanism to remove low-molecular-weight (LMW) MPAHs (59.9-77.3%), a large part of OPAHs, including anthraquinone, methylanthraquinone, and 9-fluorenone (46.7-49.6%), and some NPAHs, including 2-nitrofluorene and 9-nitroanthrancene (52.9-59.1%). Adsorption by activated sludge mainly accounted for removing high-molecular-weight (HMW) SPAHs (59.6-71.01%). The relatively high concentrations of SPAHs in excess sludge (15,000 µg·g-1) and treated effluent (104 µg·L-1) are of great concern for their potential adverse ecological impacts. SPAHS exhibited similar behaviors in central WWTP, though the influent concentrations were much lower. The concentration levels of SPAHs in the ambient air of coking plant E and central WWTP may also pose potential lung cancer risks (LCR) to the workers through inhalation, where all studied SPAHs except 3-nitrofluoranthene and 7-nitrobenz[a]anthracene exceeded the acceptable cancer risk standards (>10-6) recommended by U.S EPA. This study could help identify the ecological and healthy risks during coking wastewater treatment and provide useful information for policy-making.

Secretion rates and protein composition of extracellular vesicles released by cancer-associated fibroblasts after radiation.

Reciprocal communication between the malignant and non-malignant cellular elements in tumors is essential for cancer sustainability and plays an important role in the response of cancers to treatments. Some of this cellular crosstalk takes place via secretion of vesicles that are actively released into the extracellular space by most cell types in tumors. Recent studies have demonstrated radiation-induced changes in the secretion rate and composition of extracellular vesicles (EVs), with impact on radiation-related cellular communication. However, little is known about the effects of different radiation regimens on the release of EVs by cells of the tumor microenvironment. In this study, we provide a comprehensive molecular characterization of EVs released by cultured primary lung tumor fibroblasts. We explore the quantitative and morphological changes triggered by ionizing radiation (IR), delivered as a single dose of 18 Gy or three consecutive daily medium-doses of 6 Gy. Cancer-associated fibroblasts (CAFs) secrete EVs with sizes ranging from 80 to 200 nm, expressing some of the classical exosome markers. Exposing CAFs to a single-high radiation dose (1 × 18 Gy) or fractionated medium-dose did not alter the release of CAF-EVs. The protein composition of CAF-EVs was analyzed by LC-MS/MS proteomics and revealed that CAF-EVs are enriched with heat shock proteins, integrins, tetraspanins, proteinases, collagens, growth factors and an array of molecules involved in the regulation of cell migration and the immune system. Quantitative proteomic analyses revealed minor changes in the protein composition of CAF-EVs after radiation exposure. Taken together, this study presents original data on lung tumor CAF-EV composition and reveals that release and protein cargo of CAF-EVs are largely unaltered after exposing CAFs to IR.

Emerging concerns of VOCs and SVOCs in coking wastewater treatment processes: Distribution profile, emission characteristics, and health risk assessment.

In this study, the distribution profiles, emission characteristics, and health risks associated with 43 volatile and semi-volatile organic compounds, including 15 phenols, 18 polycyclic aromatic hydrocarbons (PAHs), 6 BTEX, and 4 other compounds, were determined in the wastewater treatment plant (WWTP) of a coking factory (plant C) and the succeeding final WWTP (central WWTP). Total phenols with a concentration of 361,000 µg L-1 were the predominant compounds in the influent wastewater of plant C, whereas PAHs were the major compounds in the final effluents of both coking WWTPs (84.4 µg L-1 and 30.7 µg L-1, respectively). The biological treatment process in plant C removed the majority of volatile organic pollutants (94.1%-99.9%). A mass balance analysis for plant C showed that biodegradation was the main removal pathway for all the target compounds (56.6%-99.9%) except BTEX, chlorinated phenols, and high molecular weight (MW) PAHs. Chlorinated phenols and high MW PAHs were mainly removed via sorption to activated sludge (51.8%-73.2% and 60.2%-75.9%, respectively). Air stripping and volatilization were the dominant mechanisms for removing the BTEX compounds (59.8%-73.8%). The total emission rates of the detected volatile pollutants from plant C and the central WWTP were 1,640 g d-1 and 784 g d-1, respectively. Benzene from the equalization basins of plant C and the central WWTP corresponded to the highest inhalation carcinogenic risks (1.4 × 10-3 and 3.2 × 10-4, respectively), which exceeded the acceptable level for human health (1 × 10-6) recommended by the United States Environmental Protection Agency. The results showed that BaP exhibited the highest inhalation non-cancer risk, with a hazard index ratio of 70 and 30 for plant C and the central WWTP, respectively. Moreover, the excess sludge generated during wastewater treatment should also be carefully handled because it adsorbed abundant PAHs and chlorinated phenols at coking plant C (58,000 µg g-1 and 3,500 µg g-1) and the central WWTP (622 µg g-1 and 54 µg g-1).

Hippocampal knockdown of α2 nicotinic or M1 muscarinic acetylcholine receptors in C57BL/6J male mice impairs cued fear conditioning.

Acetylcholine (ACh) signaling in the hippocampus is important for behaviors related to learning, memory and stress. In this study, we investigated the role of two ACh receptor subtypes previously shown to be involved in fear and anxiety, the M1 mAChR and the α2 nAChR, in mediating the effects of hippocampal ACh on stress-related behaviors. Adeno-associated viral vectors containing short-hairpin RNAs targeting M1 or α2 were infused into the hippocampus of male C57BL/6J mice, and behavior in a number of paradigms related to stress responses and fear learning was evaluated. There were no robust effects of hippocampal M1 mAChR or α2 nAChR knockdown (KD) in the light/dark box, tail suspension, forced swim or novelty-suppressed feeding tests. However, effects on fear learning were observed in both KD groups. Short term learning was intact immediately after training in all groups of mice, but both the M1 and α2 hippocampal knock down resulted in impaired cued fear conditioning 24 h after training. In addition, there was a trend for a deficit in contextual memory the M1 mAChR KD group 24 h after training. These results suggest that α2 nicotinic and M1 muscarinic ACh receptors in the hippocampus contribute to fear learning and could be relevant targets to modify brain circuits involved in stress-induced reactivity to associated cues.

Irradiated Tumor Fibroblasts Avoid Immune Recognition and Retain Immunosuppressive Functions Over Natural Killer Cells.

Recent studies have demonstrated that radiotherapy is able to induce anti-tumor immune responses in addition to mediating direct cytotoxic effects. Cancer-associated fibroblasts (CAFs) are central constituents of the tumor stroma and participate actively in tumor immunoregulation. However, the capacity of CAFs to influence immune responses in the context of radiotherapy is still poorly understood. This study was undertaken to determine whether ionizing radiation alters the CAF-mediated immunoregulatory effects on natural killer (NK) cells. CAFs were isolated from freshly resected non-small cell lung cancer tissues, while NK cells were prepared from peripheral blood of healthy donors. Functional assays to study NK cell immune activation included proliferation rates, expression of cell surface markers, secretion of immunomodulators, cytotoxic assays, as well as production of intracellular activation markers such as perforin and granzyme B. Our data show that CAFs inhibit NK cell activation by reducing their proliferation rates, the cytotoxic capacity, the extent of degranulation, and the surface expression of stimulatory receptors, while concomitantly enhancing surface expression of inhibitory receptors. Radiation delivered as single high-dose or in fractioned regimens did not reverse the immunosuppressive features exerted by CAFs over NK cells in vitro, despite triggering enhanced surface expression of several checkpoint ligands on irradiated CAFs. In summary, CAFs mediate noticeable immune inhibitory effects on cytokine-activated NK cells during co-culture in a donor-independent manner. However, ionizing radiation does not interfere with the CAF-mediated immunosuppressive effects.

Fibroblast-Mediated Immunoregulation of Macrophage Function Is Maintained after Irradiation.

The abilities of cancer-associated fibroblasts (CAFs) to regulate immune responses in the context of radiotherapy remain largely unknown. This study was undertaken to determine whether ionizing radiation alters the CAF-mediated immunoregulatory effects on macrophages. CAFs were isolated from freshly-resected non-small cell lung cancer tumors, while monocyte-derived macrophages were prepared from peripheral blood of healthy donors. Experimental settings included both (CAF-macrophage) co-cultures and incubations of M0 and M1-macrophages in the presence of CAF-conditioned medium (CAF-CM). Functional assays to study macrophage polarization/activation included the expression of cell surface markers, production of nitric oxide, secretion of inflammatory cytokines and migratory capacity. We show that CAFs promote changes in M0-macrophages that harmonize with both M1-and M2-phenotypes. Additionally, CAFs inhibit pro-inflammatory features of M1-macrophages by reducing nitric oxide production, pro-inflammatory cytokines, migration, and M1-surface markers expression. Radiation delivered as single-high dose or in fractioned regimens did not modify the immunoregulatory features exerted by CAFs over macrophages in vitro. Protein expression analyses of CAF supernatants showed that irradiated and non-irradiated CAFs produce approximately the same protein levels of immunoregulators. Thus, CAF-derived soluble factors mediate measurable changes on uncommitted macrophages and down-regulate pro-inflammatory features of M1-polarized macrophages. Notably, ionizing radiation does not curtail the CAF-mediated immunosuppressive effects.

Quadruplex-forming oligonucleotide targeted to the VEGF promoter inhibits growth of non-small cell lung cancer cells.

BACKGROUND: Vascular endothelial growth factor (VEGF) is commonly overexpressed in a variety of tumor types including lung cancer. As a key regulator of angiogenesis, it promotes tumor survival, growth, and metastasis through the activation of the downstream protein kinase B (AKT) and extracellular signal-regulated kinase (ERK 1/2) activation. The VEGF promoter contains a 36 bp guanine-rich sequence (VEGFq) which is capable of forming quadruplex (four-stranded) DNA. This sequence has been implicated in the down-regulation of both basal and inducible VEGF expression and represents an ideal target for inhibition of VEGF expression. RESULTS: Our experiments demonstrate sequence-specific interaction between a G-rich quadruplex-forming oligonucleotide encoding a portion of the VEGFq sequence and its double stranded target sequence, suggesting that this G-rich oligonucleotide binds specifically to its complementary C-rich sequence in the genomic VEGF promoter by strand invasion. We show that treatment of A549 non-small lung cancer cells (NSCLC) with this oligonucleotide results in decreased VEGF expression and growth inhibition. The VEGFq oligonucleotide inhibits proliferation and invasion by decreasing VEGF mRNA/protein expression and subsequent ERK 1/2 and AKT activation. Furthermore, the VEGFq oligonucleotide is abundantly taken into cells, localized in the cytoplasm/nucleus, inherently stable in serum and intracellularly, and has no effect on non-transformed cells. Suppression of VEGF expression induces cytoplasmic accumulation of autophagic vacuoles and increased expression of LC3B, suggesting that VEGFq may induce autophagic cell death. CONCLUSION: Our data strongly suggest that the G-rich VEGFq oligonucleotide binds specifically to the C-rich strand of the genomic VEGF promoter, via strand invasion, stabilizing the quadruplex structure formed by the genomic G-rich sequence, resulting in transcriptional inhibition. Strand invading oligonucleotides represent a new approach to specifically inhibit VEGF expression that avoids many of the problems which have plagued the therapeutic use of oligonucleotides. This is a novel approach to specific inhibition of gene expression.

Plagiochiline A Inhibits Cytokinetic Abscission and Induces Cell Death.

We previously reported on the isolation and biological activities of plagiochiline A (1), a 2,3-secoaromadendrane-type sesquiterpenoid from the Peruvian medicinal plant, Plagiochila disticha. This compound was found to have antiproliferative effects on a variety of solid tumor cell lines, as well as several leukemia cell lines. Other researchers have also noted the cytotoxicity of plagiochiline A (isolated from different plant species), but there are no prior reports regarding the mechanism for this bioactivity. Here, we have evaluated the effects of plagiochiline A on cell cycle progression in DU145 prostate cancer cells. A cell cycle analysis indicated that plagiochiline A caused a significant increase in the percentage of cells in the G2/M phase when compared with control cells. When cells were stained and observed by fluorescence microscopy to examine progress through the mitotic phase, we found a significant increase in the proportion of cells with features of late cytokinesis (cells connected by intercellular bridges) in the plagiochiline A-treated samples. These results suggest that plagiochiline A inhibits cell division by preventing completion of cytokinesis, particularly at the final abscission stage. We also determined that plagiochiline A reduces DU145 cell survival in clonogenic assays and that it induces substantial cell death in these cells.

Microencapsulation of fish oil using supercritical antisolvent process.

In order to improve the encapsulation process, a newly supercritical antisolvent process was developed to encapsulate fish oil using hydroxypropyl methyl cellulose as a polymer. Three factors, namely, temperature, pressure, and feed emulsion rate were optimized using response surface methodology. The suitability of the model for predicting the optimum response value was evaluated at the conditions of temperature at 60°C, pressure at 150 bar, and feed rate at 1.36 mL/min. At the optimum conditions, particle size of 58.35 µm was obtained. The surface morphology of the micronized fish oil was also evaluated using field emission scanning electron microscopy where it showed that particles formed spherical structures with no internal voids. Moreover, in vitro release of oil showed that there are significant differences of release percentage of oil between the formulations and the results proved that there was a significant decrease in the in vitro release of oil from the powder when the polymer concentration was high.

c-Myc and cancer metabolism.

The processes of cellular growth regulation and cellular metabolism are closely interrelated. The c-Myc oncogene is a "master regulator" which controls many aspects of both of these processes. The metabolic changes which occur in transformed cells, many of which are driven by c-Myc overexpression, are necessary to support the increased need for nucleic acids, proteins, and lipids necessary for rapid cellular proliferation. At the same time, c-Myc overexpression results in coordinated changes in level of expression of gene families which result in increased cellular proliferation. This interesting duality of c-Myc effects places it in the mainstream of transformational changes and gives it a very important role in regulating the "transformed phenotype." The effects induced by c-Myc can occur either as a "primary oncogene" which is activated by amplification or translocation or as a downstream effect of other activated oncogenes. In either case, it appears that c-Myc plays a central role in sustaining the changes which occur with transformation. Although efforts to use c-Myc as a therapeutic target have been quite frustrating, it appears that this may change in the next few years.

Genomic c-Myc quadruplex DNA selectively kills leukemia.

c-Myc, a key regulator of cell cycle and proliferation, is commonly overexpressed in leukemia and associated with poor prognosis. Conventional antisense oligonucleotides targeting c-myc may attenuate leukemic cell growth, however, are poorly taken into cells, rapidly degraded, and have unwanted effects on normal cells. The c-myc promoter contains a guanine-rich sequence (PU27) capable of forming quadruplex (four-stranded) DNA, which may negatively regulate c-myc transcription. However, its biological significance is unknown. We show that treatment of leukemia with an oligonucleotide encoding the genomic PU27 sequence induces cell-cycle arrest and death by oncotic necrosis due to PU27-mediated suppression of c-myc mRNA/protein expression. Furthermore, PU27 is abundantly taken into cells, localized in the cytoplasm/nucleus, inherently stable in serum and intracellularly, and has no effect on normal cells. Suppression of c-myc expression by PU27 caused significant DNA damage, cell and mitochondrial swelling, and membrane permeability characteristic of oncotic necrosis. Induction of oncosis caused mitochondrial dysfunction, depletion of cellular ATP levels, and enhanced oxidative stress. This novel antileukemic strategy addresses current concerns of oligonucleotide therapeutics including problems with uptake, stability, and unintentional effects on normal cells and is the first report of selective cancer cell killing by a genomic DNA sequence.

Survivin modulates microtubule dynamics and nucleation throughout the cell cycle.

Survivin is a member of the chromosomal passenger complex implicated in kinetochore attachment, bipolar spindle formation, and cytokinesis. However, the mechanism by which survivin modulates these processes is unknown. Here, we show by time-lapse imaging of cells expressing either green fluorescent protein (GFP)-alpha-tubulin or the microtubule plus-end binding protein GFP-EB1 that depletion of survivin by small interfering RNAs (siRNAs) increased both the number of microtubules nucleated by centrosomes and the incidence of microtubule catastrophe, the transition from microtubule growth to shrinking. In contrast, survivin overexpression reduced centrosomal microtubule nucleation and suppressed both microtubule dynamics in mitotic spindles and bidirectional growth of microtubules in midbodies during cytokinesis. siRNA depletion or pharmacologic inhibition of another chromosomal passenger protein Aurora B, had no effect on microtubule dynamics or nucleation in interphase or mitotic cells even though mitosis was impaired. We propose a model in which survivin modulates several mitotic events, including spindle and interphase microtubule organization, the spindle assembly checkpoint and cytokinesis through its ability to modulate microtubule nucleation and dynamics. This pathway may affect the microtubule-dependent generation of aneuploidy and defects in cell polarity in cancer cells, where survivin is commonly up-regulated.