Optimal delivery of RNA interference by viral vectors for cancer therapy.

In recent years, there has been a surge in the innovative modification and application of the viral vector-based gene therapy field. Significant and consistent improvements in the engineering, delivery, and safety of viral vectors have set the stage for their application as RNA interference (RNAi) delivery tools. Viral vector-based delivery of RNAi has made remarkable breakthroughs in the treatment of several debilitating diseases and disorders (e.g., neurological diseases); however, their novelty has yet to be fully applied and utilized for the treatment of cancer. This review highlights the most promising and emerging viral vector delivery tools for RNAi therapeutics while discussing the variables limiting their success and suitability for cancer therapy. Specifically, we outline different integrating and non-integrating viral platforms used for gene delivery, currently employed RNAi targets for anti-cancer effect, and various strategies used to optimize the safety and efficacy of these RNAi therapeutics. Most importantly, we provide great insight into what challenges exist in their application as cancer therapeutics and how these challenges can be effectively navigated to advance the field.

Identification of FDA-approved bifonazole as a SARS-CoV-2 blocking agent following a bioreporter drug screen.

We established a split nanoluciferase complementation assay to rapidly screen for inhibitors that interfere with binding of the receptor binding domain (RBD) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein with its target receptor, angiotensin-converting enzyme 2 (ACE2). After a screen of 1,200 US Food and Drug Administration (FDA)-approved compounds, we identified bifonazole, an imidazole-based antifungal agent, as a competitive inhibitor of RBD-ACE2 binding. Mechanistically, bifonazole binds ACE2 around residue K353, which prevents association with the RBD, affecting entry and replication of spike-pseudotyped viruses as well as native SARS-CoV-2 and its variants of concern (VOCs). Intranasal administration of bifonazole reduces lethality in K18-hACE2 mice challenged with vesicular stomatitis virus (VSV)-spike by 40%, with a similar benefit after live SARS-CoV-2 challenge. Our screen identified an antiviral agent that is effective against SARS-CoV-2 and VOCs such as Omicron that employ the same receptor to infect cells and therefore has high potential to be repurposed to control, treat, or prevent coronavirus disease 2019 (COVID-19).

Single-dose replicating poxvirus vector-based RBD vaccine drives robust humoral and T cell immune response against SARS-CoV-2 infection.

The coronavirus disease 2019 (COVID-19) pandemic requires the continued development of safe, long-lasting, and efficacious vaccines for preventive responses to major outbreaks around the world, and especially in isolated and developing countries. To combat severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), we characterize a temperature-stable vaccine candidate (TOH-Vac1) that uses a replication-competent, attenuated vaccinia virus as a vector to express a membrane-tethered spike receptor binding domain (RBD) antigen. We evaluate the effects of dose escalation and administration routes on vaccine safety, efficacy, and immunogenicity in animal models. Our vaccine induces high levels of SARS-CoV-2 neutralizing antibodies and favorable T cell responses, while maintaining an optimal safety profile in mice and cynomolgus macaques. We demonstrate robust immune responses and protective immunity against SARS-CoV-2 variants after only a single dose. Together, these findings support further development of our novel and versatile vaccine platform as an alternative or complementary approach to current vaccines.

Branched-chain amino acids regulate intracellular protein turnover in porcine mammary epithelial cells.

Dietary supplementation with branched-chain amino acids (BCAAs) to lactating sows has been reported to enhance their milk production, but the underlying mechanisms remain largely unknown. This study was conducted with porcine mammary epithelial cells (PMECs) to test the hypothesis that individual BCAAs or their mixture stimulates protein synthesis and inhibit proteolysis in PMECs. Cells were cultured at 37 °C in customized Dulbecco's modified Eagle medium containing 5 mmol/L D-glucose, 1 mmol/L L-phenylalanine, L-[ring-2,4-3H]phenylalanine, 0.1 (control), 0.25, 0.5, 1, or 2 mmol/L L-leucine, L-isoleucine or L-valine or an equimolar mixture of the three BCAAs. The culture medium also contained physiological concentrations of other amino acids found in the plasma of lactating sows. Proliferation, protein synthesis, proteolysis, ß-casein production, the mechanistic target of rapamycin (mTOR) signaling, and the ubiquitin-proteasome pathway were determined for PMECs. Cell proliferation and abundances of phosphorylated mTOR, eukaryotic translation initiation factor 4E-binding protein 1, and ribosomal protein S6 kinase ß-1 proteins increased (P < 0.05), but abundances of ubiquitinated protein and 20S proteasome decreased (P < 0.05) when extracellular concentrations of L-leucine, L-isoleucine, L-valine, or an equimolar mixture of BCAAs were increased from 0.1 to 2 mmol/L. Compared with the control, 0.25, 0.5, 1 or 2 mmol/L BCAAs enhanced (P < 0.01) protein (including ß-casein) synthesis, while decreasing (P < 0.05) proteolysis in PMECs in a dose-dependent manner. Collectively, our results indicate that physiological concentrations of BCAAs regulate protein turnover in mammary epithelial cells to favor net protein synthesis through stimulating the mTOR signaling pathway and inhibiting the ubiquitin-proteasome pathway.

Dietary supplementation with monosodium glutamate enhances milk production by lactating sows and the growth of suckling piglets.

This study was conducted to test the hypothesis that increasing dietary content of glutamate through addition of monosodium glutamate (MSG) enhances milk production by lactating sows and the growth of their offspring. Thirty multiparous sows (Landrace × Large White) were assigned randomly into one of three dietary groups: control (a corn- and soybean meal-based diet), the basal diet + 1% MSG, and the basal diet + 2% MSG. Diets were made isonitrogenous by the addition of appropriate amounts of L-alanine. Lactating sows had free access to drinking water and were fed twice daily their respective diets. The number of live-born piglets was standardized to 9 per sow at day 0 of lactation (the day of farrowing). On days 3, 15, and 29 of lactation, body weight and milk consumption of piglets were measured, and blood samples obtained from sows and piglets at 2 h and 1 h after feeding, respectively. Feed intake of sows did not differ (P > 0.05) among the three groups of sows. Concentrations of aspartate, glutamine, citrulline, arginine, tryptophan, proline, branched-chain amino acids, and glutamate were greater (P < 0.05) in the plasma of MSG-supplemented sows and their piglets than for controls. When compared with the control, dietary supplementation with 1-2% MSG increased (P < 0.05): concentrations of many free amino acids (including glutamate plus glutamine) and all protein-bound amino acids in milk; the milk intake of piglets by 14-25%; and daily weight gains of piglets by 23-44%. These results indicate that dietary supplementation with 1-2% MSG to lactating sows enhances milk production to support the growth of sow-reared piglets.

Nanoluciferase complementation-based bioreporter reveals the importance of N-linked glycosylation of SARS-CoV-2 S for viral entry.

The ongoing COVID-19 pandemic has highlighted the immediate need for the development of antiviral therapeutics targeting different stages of the SARS-CoV-2 life cycle. We developed a bioluminescence-based bioreporter to interrogate the interaction between the SARS-CoV-2 viral spike (S) protein and its host entry receptor, angiotensin-converting enzyme 2 (ACE2). The bioreporter assay is based on a nanoluciferase complementation reporter, composed of two subunits, large BiT and small BiT, fused to the S receptor-binding domain (RBD) of the SARS-CoV-2 S protein and ACE2 ectodomain, respectively. Using this bioreporter, we uncovered critical host and viral determinants of the interaction, including a role for glycosylation of asparagine residues within the RBD in mediating successful viral entry. We also demonstrate the importance of N-linked glycosylation to the RBD's antigenicity and immunogenicity. Our study demonstrates the versatility of our bioreporter in mapping key residues mediating viral entry as well as screening inhibitors of the ACE2-RBD interaction. Our findings point toward targeting RBD glycosylation for therapeutic and vaccine strategies against SARS-CoV-2.

Investigating the biological degradation of the drug ß-blocker atenolol from wastewater using the SBR.

Drug compounds are one of the main contributors to the entry of micro-pollutants into the environment, known as a constant threat to environmental stability. Atenolol is a type of beta-blocker extensively used to cure cardiovascular disorders. The residues of this compound have been continuously detected in aquatic environments because it is a polar and poorly biodegradable compound. Thus, removing atenolol from wastewater is essential before discharging into the environment. Biological processes are considered the most important removal process for polar drugs in wastewater treatment plants. Accordingly, for the first time in this study, the SBR performance was investigated in the biodegradation and mineralization of atenolol under different concentrations (50-600 mg/L) and hydraulic retention times (48-32 h). Based on the results, the time required for the acclimation of biomass to atenolol (C: 50 mg/L and the HRT: 48 h) was 80 days. The SBR efficiencies under optimum conditions (C: 400 mg/L and HRT: 40 h) in removing the atenolol and COD were 91% and 87%, respectively. For the first time in this study, one of the main pathways of the atenolol biodegradation was identified. Based on the review and comparison of the results of this study with existing literature showing that the SBR used in this study was able to remove higher concentrations with better efficiencies than other processes. Therefore, it can be concluded that the SBR used in this study could be considered an efficient and promising technique for treating wastewaters containing atenolol and other beta-blocker group drugs.

Characterization of Critical Determinants of ACE2-SARS CoV-2 RBD Interaction.

Despite sequence similarity to SARS-CoV-1, SARS-CoV-2 has demonstrated greater widespread virulence and unique challenges to researchers aiming to study its pathogenicity in humans. The interaction of the viral receptor binding domain (RBD) with its main host cell receptor, angiotensin-converting enzyme 2 (ACE2), has emerged as a critical focal point for the development of anti-viral therapeutics and vaccines. In this study, we selectively identify and characterize the impact of mutating certain amino acid residues in the RBD of SARS-CoV-2 and in ACE2, by utilizing our recently developed NanoBiT technology-based biosensor as well as pseudotyped-virus infectivity assays. Specifically, we examine the mutational effects on RBD-ACE2 binding ability, efficacy of competitive inhibitors, as well as neutralizing antibody activity. We also look at the implications the mutations may have on virus transmissibility, host susceptibility, and the virus transmission path to humans. These critical determinants of virus-host interactions may provide more effective targets for ongoing vaccines, drug development, and potentially pave the way for determining the genetic variation underlying disease severity.

miR-455-5p downregulation promotes inflammation pathways in the relapse phase of relapsing-remitting multiple sclerosis disease.

MicroRNA-455-5p (miR-455-5p) seems to have an anti-inflammatory role in the immune system since its expression is induced by IL-10 cytokine. Multiple sclerosis (MS) is a chronic demyelinating neurodegenerative disease of the central nervous system that is caused by an autoimmune inflammatory attack against the myelin insulation of neurons. The expression level of miR-455-5p and its role in MS pathogenesis has yet to be elucidated. We found that miR-455-5p expression was highly correlated with disease severity in MS patients. miR-455-5p expression inversely correlates with its inflammatory-predicted targets (MyD88 and REL) in relapse- and remitting-phase patients. Luciferase assays confirm that MyD88 and REL are direct targets of miR-455-5p. This study represents the first report of the miR-455-5p acts as an anti-inflammatory role in MS, at least partially through targeting MyD88 and REL. This study may provide important information for the use of miR-455-5p as a novel strategy to improve the severity of disease and control inflammation and attack in MS patients.

Effect of algae acclimation to the wastewater medium on the growth kinetics and nutrient removal capacity.

Algal treatment methods have been widely used in nutrient removal studies. However, in most cases, the experimental conditions have not been fully complied with actual conditions. For instance, the effect of algae acclimation to wastewater medium on cell growth and removal efficiency has generally been ignored in laboratory scale experiments. This paper investigates the effect of acclimation on cell growth and nutrient uptake rates of Arthrospira platensis and Chlorella vulgaris. For this purpose, batch reactors, which contained the synthetic secondary effluent, had been inoculated by acclimated algae cells and the growth parameters were measured daily, as well as nutrient concentration. A significant decrease (P < 0.05) in chlorophyll-a content of acclimated A. platensis was observed, although there was no significant change in specific growth rate (µ) and doubling time (dt), in comparison with the non-acclimated ones. Moreover, the acclimation process changed the chlorophyll-a content and kinetic parameters of Chlorella vulgaris. Furthermore, t test results showed a significant increase in removal rate of nitrogen compounds through the acclimation. Residence time of A. platensis and C. vulgaris was also reduced through the acclimation by approximately 50% and 25%, respectively.

Posterior-only surgery with preoperative skeletal traction for management of severe scoliosis.

PURPOSE: The surgical treatment of severe adolescent spinal deformities is challenging and carries substantial risks of mortality and morbidity. To mitigate this risk, surgeons have employed various methods as this study designed to evaluate the safety and effectiveness of preoperative halo-femoral or halo gravity traction (HGT) followed by posterior-only surgery in the management of severe scoliosis. METHOD: A total number of 23 patients with severe scoliosis treated by preoperative skeletal traction (halo gravity or halo femoral) followed by posterior fusion and instrumentation in one stage. All patients were followed for a minimum of 2 years after surgery. RESULTS: The average age of the patients was 12.7 years at the time of surgery. Mean of the Cobb angle improved from 99.9° ± 8.2° preoperatively to 75.3° ± 8° post-traction and 49.5° ± 7.7° postoperatively. Kyphosis angle corrected from 56.4° ± 9.5° to 38.6° ± 5.8°. The preop-FVC% was 41 ± 6.1% and after 1 year follow-up FVC% was 45.7 ± 7.7%. No patients required an anterior release due to amount of their deformity. DISCUSSION: Despite the benefits of modern instrumentation procedures, the treatment of severe scoliosis can be very competing. We think that by applying preoperative halo femoral traction and halo-gravity traction, managing severe scoliosis will be in safe and easy manner and can lead to better deformity correction and less neurological complications and facilitate to avoid anterior operation for severe scoliosis and its related complications.

Safety of long-term dietary supplementation with L-arginine in pigs.

This study was conducted with a swine model to determine the safety of long-term dietary supplementation with L-arginine-HCl or L-arginine free base. Beginning at 30 days of age, pigs were fed a corn- and soybean meal-based diet (31.5 g/kg body weight/day) supplemented with 0, 1.21, 1.81 or 2.42 % L-arginine-HCl (Experiment 1) or with 0, 1, 1.5 or 2 % L-arginine (Experiment 2). The supplemental doses of 0, 1, 1.5, and 2 % L-arginine provided pigs with 0, 315, 473, and 630 mg L-arginine/kg body weight/day, respectively, which were equivalent to 0, 286, 430, and 573 mg L-arginine/kg body weight/day, respectively, in humans. At 121 days of age (91 days after initiation of supplementation), blood samples were obtained from the jugular vein of pigs at 1 and 4 h after feeding for hematological and clinical chemistry tests. Dietary supplementation with L-arginine increased plasma concentrations of arginine, ornithine, proline, albumin and reticulocytes, while reducing plasma concentrations of ammonia, free fatty acids, triglyceride, cholesterol, and neutrophils. L-Arginine supplementation enhanced protein gain and reduced white-fat deposition in the body. Other variables in standard hematology and clinical chemistry tests, serum concentrations of insulin, growth hormone and insulin-like growth factor-I did not differ among all the groups of pigs. These results indicate that dietary supplementation with L-arginine (up to 630 mg/kg body weight/day) is safe in pigs for at least 91 days. Our findings help guide clinical studies to determine the safety of long-term oral administration of L-arginine to humans.

Primary extraperitoneal hydatid cyst, a rare differential diagnosis of subdiaphragmatic mass: A case report.

Key Clinical Message: Hydatid cyst is a sly disease that can manifest with a spectrum of symptoms in almost every part of the human body, so it is crucial to be familiar with different scenarios that a patient may present. Abstract: The echinococcus granulosus parasite causes hydatid disease and is common in areas with animal husbandry and agriculture. Here, we report a middle age woman who presented with abdominal pain that further investigation revealed a cyst in subdiaphragmatic area.

Effectiveness of autologous fibrin glue in preventing post-thoracotomy air leaks: a randomized controlled trial.

Introduction: Post-thoracotomy air leaks remain a significant challenge in thoracic surgery. Aim: This randomized controlled trial assessed the efficacy of autologous fibrin glue in reducing air leaks following thoracotomy procedures. Material and methods: Conducted as a single-center, single-blind, randomized clinical trial, the study enrolled adult patients undergoing lung resection or decortication at a thoracic surgery clinic. Participants were randomly assigned to either the intervention group, receiving autologous fibrin glue application during surgery, or the control group, undergoing standard surgical procedures without glue application. Key inclusion criteria were adult patients undergoing elective thoracotomy for lung resection or decortication, while exclusion criteria included patients with severe comorbidities or contraindications to fibrin glue. Results: A total of 40 patients were enrolled and randomized equally to the two groups. The group treated with autologous fibrin glue demonstrated a significant reduction in the duration of air leakage and chest tube drainage, along with a shorter hospital stay, compared to the control group. There were no statistically significant differences in postoperative complications between the groups. Conclusions: The application of autologous fibrin glue during thoracotomy procedures significantly reduces postoperative air leaks and hospitalization duration without increasing complication rates. This finding suggests a beneficial role of fibrin glue in thoracic procedures requiring lung resection or decortication.

A systematic review of novel cannabinoids and their targets: Insights into the significance of structure in activity.

To provide a comprehensive framework of the current information on the potency and efficacy of interaction between phyto- and synthetic cannabinoids and their respective receptors, an electronic search of the PubMed, Scopus, and EMBASE literature was performed. Experimental studies included reports of mechanistic data providing affinity, efficacy, and half-maximal effective concentration (EC50). Among the 108 included studies, 174 structures, and 16 targets were extracted. The most frequent ligands belonged to the miscellaneous category with 40.2% followed by phytocannabinoid-similar, indole-similar, and pyrrole-similar structures with an abundance of 17.8%, 16.6%, and 12% respectively. 64.8% of structures acted as agonists, 17.1 % appeared as inverse agonists, 10.8% as antagonists, and 7.2% of structures were reported with antagonist/inverse agonist properties. Our outcomes identify the affinity, EC50, and efficacy of the interactions between cannabinoids and their corresponding receptors and the subsequent response, evaluated in the available evidence. Considering structures' significance and very important effects of on the activities, the obtained results also provide clues to drug repurposing.

PD-L1 promotes oncolytic virus infection via a metabolic shift that inhibits the type I IFN pathway.

While conventional wisdom initially postulated that PD-L1 serves as the inert ligand for PD-1, an emerging body of literature suggests that PD-L1 has cell-intrinsic functions in immune and cancer cells. In line with these studies, here we show that engagement of PD-L1 via cellular ligands or agonistic antibodies, including those used in the clinic, potently inhibits the type I interferon pathway in cancer cells. Hampered type I interferon responses in PD-L1-expressing cancer cells resulted in enhanced efficacy of oncolytic viruses in vitro and in vivo. Consistently, PD-L1 expression marked tumor explants from cancer patients that were best infected by oncolytic viruses. Mechanistically, PD-L1 promoted a metabolic shift characterized by enhanced glycolysis rate that resulted in increased lactate production. In turn, lactate inhibited type I IFN responses. In addition to adding mechanistic insight into PD-L1 intrinsic function, our results will also help guide the numerous ongoing efforts to combine PD-L1 antibodies with oncolytic virotherapy in clinical trials.

Dietary supplementation with monosodium glutamate is safe and improves growth performance in postweaning pigs.

Dietary intake of glutamate by postweaning pigs is markedly reduced due to low feed consumption. This study was conducted to determine the safety and efficacy of dietary supplementation with monosodium glutamate (MSG) in postweaning pigs. Piglets were weaned at 21 days of age to a corn and soybean meal-based diet supplemented with 0, 0.5, 1, 2, and 4 % MSG (n = 25/group). MSG was added to the basal diet at the expense of cornstarch. At 42 days of age (21 days after weaning), blood samples (10 mL) were obtained from the jugular vein of 25 pigs/group at 1 and 4 h after feeding for hematological and clinical chemistry tests; thereafter, pigs (n = 6/group) were euthanized to obtain tissues for histopathological examinations. Feed intake was not affected by dietary supplementation with 0-2 % MSG and was 15 % lower in pigs supplemented with 4 % MSG compared with the 0 % MSG group. Compared with the control, dietary supplementation with 1, 2 and 4 % MSG dose-dependently increased plasma concentrations of glutamate, glutamine, and other amino acids (including lysine, methionine, phenylalanine and leucine), daily weight gain, and feed efficiency in postweaning pigs. At day 7 postweaning, dietary supplementation with 1-4 % MSG also increased jejunal villus height, DNA content, and antioxidative capacity. The MSG supplementation dose-dependently reduced the incidence of diarrhea during the first week after weaning. All variables in standard hematology and clinical chemistry tests, as well as gross and microscopic structures, did not differ among the five groups of pigs. These results indicate that dietary supplementation with up to 4 % MSG is safe and improves growth performance in postweaning pigs.

Reactive Blue 19 dye removal by UV-LED/chlorine advanced oxidation process.

In recent years, advanced oxidation processes (AOPs) have indicated the greatest potential in the removal of stable organic compounds, including dyes. In this study, the ultraviolet light-emitting diodes (UV-LEDs) combined with chlorine was evaluated to remove Reactive Blue 19 (RB19) dye from aqueous solution. The effect of key experimental parameters including pH, initial chlorine concentration, initial dye concentration, and reaction time on the performance of UV-LED irradiation, UV-LED/chlorine, and the chlorination method for the removal of RB19 was studied in this research. Results showed that, more than 99% of RB19 was removed after 30 min of reaction time under optimized conditions (pH = 5, [chlorine] = 300 µM, and [RB19] = 20 mg L-1) with apparent kinetic rate constant (kapp) of 17.1 × 10-2 min-1 in UV-LED/chlorine process. However, for the chlorination method, removal efficiency was 64.7% (kapp = 3.41 × 10-2 min-1) with an apparent kinetic rate constant of 0.0341 min-1. Results also showed that UV-LED irradiation is not effective at all in removing RB19. The scavenging assay showed that OH• radicals (67.23%) had the highest contribution in RB19 removal in UV-LED/chlorine process while Cl• (17.82%) and [Formula: see text] (8.56%) had a minor role in the degradation of the dye. The RB19 degradation kinetics analysis revealed that the processes of UV-LED/chlorine and chlorination degradation followed the pseudo-first-order kinetic model. In this study, the impact of chloride, nitrate, bicarbonate, carbonate, sulfate, and sulfite anions on the performance of the process was investigated. It indicated that sulfite anion has the most negative impact on the RB19 removal process. By evaluating the synergistic effect between UV-LED lamp and chlorine, a synergy index of 5.0 was obtained for the UV-LED/chlorine process. The results presented that the UV-LED/chlorine process has a better performance than each of them alone and has the necessary efficiency for RB19 removal. Measuring COD reported its removal efficiency of 98% during the UV-LED/chlorine process under optimized conditions. Experiments continued with textile factory wastewater and indicated 30.9% of its COD removed after treatment when 1.0 µM chlorine was used.

An accessory mediastinal thyroid: A case report.

Various differential diagnoses and etiologies exist for mediastinal masses. One of the most uncommon is the ectopic mediastinal goiter. It is crucial for a clinician, especially cardiothoracic surgeons, to think about it when encountering such masses.