Discovery of Novel Small-Molecule-Based Potential PD-L1/EGFR Dual Inhibitors with High Druggability for Glioblastoma Immunotherapy.

Based on the close relationship between programmed death protein ligand 1 (PD-L1) and epidermal growth factor receptor (EGFR) in glioblastoma (GBM), we designed and synthesized a series of small molecules as potential dual inhibitors of EGFR and PD-L1. Among them, compound EP26 exhibited the highest inhibitory activity against EGFR (IC50 = 37.5 nM) and PD-1/PD-L1 interaction (IC50 = 1.77 µM). In addition, EP26 displayed superior in vitro antiproliferative activities and in vitro immunomodulatory effects by promoting U87MG cell death in a U87MG/Jurkat cell coculture model. Furthermore, EP26 possessed favorable pharmacokinetic properties (F = 22%) and inhibited tumor growth (TGI = 92.0%) in a GBM mouse model more effectively than Gefitinib (77.2%) and NP19 (82.8%). Moreover, EP26 increased CD4+ cells and CD8+ cells in tumor microenvironment. Collectively, these results suggest that EP26 represents the first small-molecule-based PD-L1/EGFR dual inhibitor deserving further investigation as an immunomodulating agent for cancer treatment.

Discovery of novel resorcinol biphenyl ether-based macrocyclic small molecules as PD-1/PD-L1 inhibitors with favorable pharmacokinetics for cancer immunotherapy.

Programmed death protein 1 (PD-1)/programmed death protein ligand 1 (PD-L1) is one of the most promising immune checkpoints (ICs) in tumor immunology and has been actively pursued as a target for anticancer drug discovery. Based on our previous research in small molecule PD-1/PD-L1 modulators, we designed and synthesized a series of resorcinol biphenyl ether-bearing macrocyclic compounds and evaluated their anti-PD-1/PD-L1 activities. Among them, compound 8d exhibited the highest inhibitory activity against PD-1/PD-L1 interaction with IC50 of 259.7 nM in the homogenous time-resolved fluorescence (HTRF) assay. In addition, 8d displayed in vitro immunomodulatory effects by promoting HepG2 cell death in a HepG2/Jurkat cell co-culture model. Furthermore, 8d effectively inhibited tumor growth (TGI = 74.6% at 40 mg/kg) in a melanoma tumor model in mice without causing obvious toxicity. Moreover, 8d exhibited favorable pharmacokinetics [e.g. high stability, reasonable half-life, and good oral bioavailability (F = 21.5%)]. Finally, molecular modeling studies showed that 8d bound to PD-L1 with high affinity. These results suggest that 8d may serve as a starting point for further development of macrocyclic small molecule-based PD-1/PD-L1 inhibitors for cancer treatment.

M2 Polarization and Inhibition of Host Cell Glycolysis Contributes Intracellular Survival of Salmonella Strains in Chicken Macrophage HD-11 Cells.

Salmonella enterica is a group of facultative, gram-negative bacteria. Recently, new evidence indicated that Salmonella could reprogram the host metabolism to increase energy or metabolites available for intracellular replication. In this study, using a chicken-specific kinomic immunometabolism peptide array analysis, we found that infection by S. Enteritidis induced significant phosphorylation changes in many key proteins of the glycolytic pathway in chicken macrophage HD-11 cells, indicating a shift in glycolysis caused by Salmonella infection. Nitric oxide production and changes of glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) represented by extracellular acidification rate (ECAR) and oxygen consumption rate (OCR), respectively, were measured in chicken macrophages infected with three Salmonella strains (S. Enteritidis, S. Heidelberg, and S. Senftenberg). The infection reduced glycolysis and enhanced OXPHOS in chicken macrophages as indicated by changes of ECAR and OCR. Salmonella strains differentially affected macrophage polarization and glycolysis. Among three strains tested, S. Enteritidis was most effective in downregulating glycolysis and promoting M2 polarization as measured by ECAR, ORC, and NO production; while S. Senftenberg did not alter glycolysis and may promote M1 polarization. Our results suggested that downregulation of host cell glycolysis and increase of M2 polarization of macrophages may contribute to increased intracellular survival of S. Enteritidis.

Leukocyte Response to Campylobacter Intra-Abdominal Infection in One Day Old Leghorn Chickens.

Using a previously characterized and described abdominal model to define the avian immune response to Salmonella intra-abdominal challenge in chickens, we have adapted this technique for the study of chickens' immune response to a Campylobacter intra-abdominal challenge. The intra-abdominal Campylobacter infection model facilitates the characterization of peripheral blood leukocyte dynamics and abdominal cell infiltrates. Day-of-hatch Leghorn chickens were injected intra-abdominally (IA) with Campylobacter jejuni [(CJ)1 × 108 colony-forming units (CFUs)]. Changes in peripheral blood leukocyte numbers and abdominal cell infiltrates were monitored at 0, 4, 8, and 24 h post-injection. Peripheral blood leukocyte numbers were also determined for 2 h post-injection. For mortality studies, birds were injected intra-abdominally with 1 × 108 CFUs CJ and mortalities were recorded for 72 h post-injection. In the peripheral blood of CJ-injected chicks, total white blood cell (WBC) numbers began increasing by 2 h post-injection, peaking at 4 h post-injection with the predominant cell type being polymorphonuclear leukocytes (heterophils). Total WBCs declined after 8 h and this decline continued at 24 h, with total WBC numbers approaching control values. The injection of CJ into the abdominal cavity caused a rapid rise in abdominal cell infiltrates with the predominant infiltrating leukocytes being heterophils. Peak abdominal heterophil infiltrates were observed at 8 h post-injection, declining only slightly by 24 h post-injection. Mortality in the CJ challenge groups reached 37%. Mortality in the Salmonella enteritidis positive control groups were greater than 50%. The data suggest that Campylobacter infection does stimulate the innate immune response in chickens when administered IA, however, the immune response and infection is not characterized with the high levels of mortality observed with a Salmonella infection. These data provide a basis for a more definitive characterization of chickens' immune response to Campylobacter and a model to evaluate intervention strategies to prevent the infection and colonization of poultry.

Addition of a protected complex of biofactors and antioxidants to breeder hen diets confers transgenerational protection against Salmonella enterica serovar Enteritidis in progeny chicks.

Addition of vitamins and antioxidants has been long associated with increased immunity and are commonly used in the poultry industry; however, less is known regarding their use in broiler breeder hens. The objective of this study was to determine if feeding a complex of protected biofactors and antioxidants composed of vitamins and fermentation extracts to broiler breeder hens conferred resistance against Salmonella enterica serovar Enteritidis (S. Enteritidis) in the progeny chicks. Three-day-old chicks from control- and supplement-fed hens were challenged with S. Enteritidis and necropsied 4- and 11-days postchallenge (dpc) to determine if there were differences in invasion and colonization. Serum and jejunum were evaluated for various cytokine and chemokine production. Fewer (P = 0.002) chicks from supplement-fed hens had detectable S. Enteritidis in the ceca (32.6%) compared to chicks from control-fed hens (64%). By 11 dpc, significantly (P < 0.001) fewer chicks from supplement-fed hens were positive for S. Enteritidis (liver [36%]; ceca [16%]) compared to chicks from the control hens (liver [76%]; ceca [76%]). The recoverable S. Enteritidis in the cecal content was also lower (P = 0.01) at 11 dpc. In additional to the differences in invasion and colonization, cytokine and chemokine production were distinct between the 2 groups of chicks. Chicks from supplement-fed hens had increased production of IL-16, IL-6, MIP-3α, and RANTES in the jejunum while IL-16 and MIP-1ß were higher in the serum of chicks from the control-fed hens. By 11 dpc, production of IFN-γ was decreased in the jejunum of chicks from supplement-fed hens. Collectively, these data demonstrate adding a protected complex of biofactors and antioxidants to the diet of broiler breeder hens offers a measure of transgenerational protection to the progeny against S. Enteritidis infection and reduces colonization that is mediated, in part, by a robust and distinct cytokine and chemokine response locally at the intestine and systemically in the blood.

Identification of the intersegmental plane by arterial ligation method during thoracoscopic segmentectomy.

BACKGROUND: Thoracoscopic segmentectomy is a common surgical procedure in thoracic surgery today. However, identifying the intersegmental plane is difficult in the surgical process. Therefore, we evaluated the feasibility of the arterial ligation method for determining the intersegmental plane and compared the demarcation status with the intravenous indocyanine green (ICG). METHODS: We retrospectively reviewed the records of 35 patients with peripheral small lung nodules who underwent thoracoscopic segmentectomy between May and December 2020. First, the preoperative three-dimensional reconstruction was performed to distinguish the location of lung nodules and the anatomical structures of targeted segmental arteries, veins, and bronchi. Second, the targeted segmental arteries were ligated, and the intersegmental plane was determined by the inflation-deflation technique. The waiting time for the appearance of the inflation-deflation line was recorded. Thirdly, the intersegmental plane was identified again using the ICG fluorescence method. Finally, the consistency of the two intersegmental planes was evaluated. RESULTS: The intersegmental planes were successfully observed in all patients using the arterial ligation method. Thirty-four patients underwent segmentectomy as planned, and one patient finally underwent lobectomy due to insufficient surgical margin. The waiting time for the appearance of the intersegmental plane by arterial ligation method was 13.7 ± 3.2 min (6-19 min). The intersegmental planes determined by the arterial ligation method and the ICG fluorescence method were comparable, with a maximum distance of no more than 5 mm between the two planes. The mean operative duration was 119.1 ± 34.9 min, and the mean blood loss was 76.9 ± 70.3 ml. No evident air leakage was found during the operation. Only one patient experienced a prolonged air leak (≥ 5 days) during the postoperative recovery. No atelectasis occurred in all cases. The chest tube duration was 3.1 ± 0.9 days. CONCLUSION: The arterial ligation method can efficiently and accurately identify the intersegmental plane, comparable to the ICG fluorescence method.

Chicken-Specific Kinome Analysis of Early Host Immune Signaling Pathways in the Cecum of Newly Hatched Chickens Infected With Salmonella enterica Serovar Enteritidis.

Poultry is a major source of human foodborne illness caused by broad host range Salmonella serovars (paratyphoid), and developing cost-effective, pre-harvest interventions to reduce these pathogens would be valuable to the industry and consumer. Host responses to infectious agents are often regulated through phosphorylation. However, proteomic mechanisms of Salmonella acute infection biology and host responses to the bacteria have been limited concentrating predominately on the genomic responses of the host to infection. Our recent development of chicken-specific peptide arrays for kinome analysis of host phosphorylation-based cellular signaling responses provided us with the opportunity to develop a more detailed understanding of the early (4-24 h post-infection) host-pathogen interactions during the initial colonization of the cecum by Salmonella. Using the chicken-specific kinomic immune peptide array, biological pathway analysis showed infection with S. Enteritidis increased signaling related to the innate immune response, relative to the non-infected control ceca. Notably, the acute innate immune signaling pathways were characterized by increased peptide phosphorylation (activation) of the Toll-like receptor and NOD-like receptor signaling pathways, the activation of the chemokine signaling pathway, and the activation of the apoptosis signaling pathways. In addition, Salmonella infection induced a dramatic alteration in the phosphorylation events of the JAK-STAT signaling pathway. Lastly, there is also significant activation of the T cell receptor signaling pathway demonstrating the initiation of the acquired immune response to Salmonella infection. Based on the individual phosphorylation events altered by the early Salmonella infection of the cecum, certain conclusions can be drawn: (1) Salmonella was recognized by both TLR and NOD receptors that initiated the innate immune response; (2) activation of the PPRs induced the production of chemokines CXCLi2 (IL-8) and cytokines IL-2, IL-6, IFN-α, and IFN-γ; (3) Salmonella infection targeted the JAK-STAT pathway as a means of evading the host response by targeting the dephosphorylation of JAK1 and TYK2 and STAT1,2,3,4, and 6; (4) apoptosis appears to be a host defense mechanism where the infection with Salmonella induced both the intrinsic and extrinsic apoptotic pathways; and (5) the T cell receptor signaling pathway activates the AP-1 and NF-κB transcription factor cascades, but not NFAT.

Targeting Pulmonary Artery Infusion of Nuclear-Targeted Plasmid-Based Short Hairpin RNA (ShRNA) to Hypoxia Inducible Factor-1α3 (pshHIF-1α3) Nano-Microspheres for Treatment of Implanted Lung Cancer in Rats.

The pshHIF-1α3 stealth nanospheres have been studied if they have the function of arterial targeted drug delivery to provide a new arterial targeted drug delivery method for interventional therapy of lung cancer. The study is also aimed at exploring therapeutic effect of the checked drug delivery on lung cancer. The tested groups were designed as follows: Group I: blank control group (pulmonary artery perfusion of 0.5 mL 0.9% saline); group II: tail vein injection of pshHIF-1α3 nano-microsphere; group III: pshHIF-1α3 nano-microsphere pulmonary artery perfusion group. In vitro experiment assessed the effects of pulmonary artery perfusion of pshHIF-1α3 nanospheres on proliferation, apoptosis and colony forming ability of lung cancer A549 cells, which were all evaluated by using MTT method, flow cytometry and colony formation experiments, respectively. In vivo experiment tumor xenotransplantation was used to observe the effect of pulmonary artery perfusion of pshHIF-1α3 nanospheres on treatment of lung cancer. Both the In vivo pulmonary artery perfusion experiment and In vitro experiments in A549 cells confirmed that the pulmonary artery perfusion of pshHIF-1α3 nano-microspheres can inhibit the proliferation of lung cancer tissues and cells, promoting apoptosis and inhibiting migration, leading to enhanced therapeutic effect of lung cancer. One of characteristics of nanomaterials is their large surface area, high dispersion, specific adhesion, tumor-specific affinity and adhesion, thereby prolonging their circulation time in the body. Through aggregation of nanodrug delivery system in tumor cells, the local concentration of the drug is increased, thereby improving selectivity of chemotherapeutic drugs. The results from this study therefore suggest that pulmonary artery perfusion of pshHIF-1α3 may be used in arterial targeted drug delivery for treatment of lung cancer, providing a new and efficient targeted drug delivery arterial route for interventional therapy of lung cancer.

A blend of microencapsulated organic acids and botanicals reduces necrotic enteritis via specific signaling pathways in broilers.

Necrotic enteritis (NE) is a devastating disease that has seen a resurgence of cases following the removal of antibiotics from feed resulting in financial loss and significant animal health concerns across the poultry industry. The objective was to evaluate the efficacy of a microencapsulated blend of organic (25% citric and 16.7% sorbic) acids and botanicals (1.7% thymol and 1% vanillin [AviPlusP]) to reduce clinical NE and determine the signaling pathways associated with any changes. Day-of-hatch by-product broiler breeder chicks were randomly assigned to a control (0) or supplemented (500 g/MT) diet (n = 23-26) and evaluated in a NE challenge model (n = 3). Birds were administered 2X cocci vaccine on d 14 and challenged with a cocktail of Clostridium perfringens strains (107) on d 17 to 19. On d 20 to 21 birds were weighed, euthanized, and scored for NE lesions. Jejunal tissue was collected for kinome analysis using an immuno-metabolism peptide array (n = 5; 15/treatment) to compare tissue from supplement-fed birds to controls. Mortality and weight were analyzed using Student's t test and lesion scores analyzed using F-test two-sample for variances (P < 0.05). The kinome data was analyzed using PIIKA2 peptide array analysis software and fold-change between control and treated groups determined. Mortality in the supplemented group was 47.4% and 70.7% in controls (P = 0.004). Lesions scores were lower (P = 0.006) in supplemented birds (2.47) compared to controls (3.3). Supplement-fed birds tended (P = 0.19) to be heavier (848.6 g) than controls (796.2 g). Kinome analysis showed T cell receptor, TNF and NF-kB signaling pathways contributed to the improvements seen in the supplement-fed birds. The following peptides were significant (P < 0.05) in all 3 pathways: CHUK, MAP3K14, MAP3K7, and NFKB1 indicating their importance. Additionally, there were changes to IL6, IL10, and IFN- γ mRNA expression in tissue between control- and supplement-fed chickens. In conclusion, the addition of a microencapsulated blend of organic acids and botanicals to a broiler diet reduced the clinical signs of NE that was mediated by specific immune-related pathways.

Antagonistic Effects of Lipids Against the Anti-Escherichia coli and Anti-Salmonella Activity of Thymol and Thymol-ß-d-Glucopyranoside in Porcine Gut and Fecal Cultures In Vitro.

Strategies are sought to reduce the carriage and dissemination of zoonotic pathogens and antimicrobial-resistant microbes within food-producing animals and their production environment. Thymol (an essential oil) is a potent bactericide in vitro but in vivo efficacy has been inconsistent, largely due to its lipophilicity and absorption, which limits its passage and subsequent availability in the distal gastrointestinal tract. Conjugation of thymol to glucose to form thymol-ß-d-glucopyranoside can decrease its absorption, but in vivo passage of effective concentrations to the lower gut remains suboptimal. Considering that contemporary swine diets often contain 5% or more added fat (to increase caloric density and reduce dustiness), we hypothesized that there may be sufficient residual fat in the distal intestinal tract to sequester free or conjugated thymol, thereby limiting the availability and subsequent effectiveness of this biocide. In support of this hypothesis, the anti-Salmonella Typhimurium effects of 6 mM free or conjugated thymol, expressed as log10-fold reductions of colony-forming units (CFU) ml-1, were diminished 90 and 58%, respectively, following 24-h in vitro anaerobic fecal incubation (at 39°C) with 3% added vegetable oil compared to reductions achieved during culture without added oil (6.1 log10 CFU ml-1). The antagonistic effect of vegetable oil and the bactericidal effect of free and conjugated thymol against Escherichia coli K88 tested similarly were diminished 86 and 84%, respectively, compared to reductions achieved in cultures incubated without added vegetable oil (5.7 log10 CFU ml-1). Inclusion of taurine (8 mg/ml), bile acids (0.6 mg/ml), or emulsifiers such as polyoxyethylene-40 stearate (0.2%), Tween 20, or Tween 80 (each at 1%) in the in vitro incubations had little effect on vegetable oil-caused inhibition of free or conjugated thymol. Based on these results, it seems reasonable to suspect that undigested lipid in the distal gut may limit the effectiveness of free or conjugated thymol. Accordingly, additional research is warranted to learn how to overcome obstacles diminishing bactericidal activity of free and conjugated thymol in the lower gastrointestinal tract of food-producing animals.

Evaluation on the Distribution of EGFR, KRAS and BRAF Genes and the Expression of PD-L1 in Different Types of Lung Cancer.

OBJECTIVE: To evaluate the distribution of high frequency mutant genes and the expression of PDL1 in different types of lung cancer. METHODS: This retrospective analysis was conducted on 330 patients who were diagnosed with primary lung cancer and treated in our hospital from October 2018 to October 2020. The patients were listed into non-small cell carcinoma group (101 cases), squamous carcinoma group (28 cases) and adenocarcinoma group (201 cases) according to their pathological results. The gene mutations were detected using EGFR, KRAS, and BRAF gene mutation detection kits, and the expression of PDL1 was detected by immunostaining. The mutation of EGFR, KRAS and BRAF genes and PDL1 expression in patients with different types of lung cancer were compared. RESULTS: The patients in the adenocarcinoma group had the highest incidence of EGFR gene mutation, the mutation rate of the gene whose mutation location was exon 18 was significantly higher, and the difference between each group was statistically significant (P < 0.05). The patients in the adenocarcinoma group had the highest incidence of KRAS gene mutation, the mutation rate of the gene whose mutation location was exon 2 was obviously the highest, exon 15 was the lowest, and the difference between each group was statistically significant (P < 0.05). There was no significant difference in the distribution of BRAF gene mutations among groups, and all mutations occurred on exon 15, with no statistically significant difference between each group (P > 0.05). PD-L1 expression in NSCLC patients was significantly higher than that in other lung cancer patients (P < 0.05). CONCLUSION: EGFR and KRAS genes showed obvious specific expressions in patients with different types of lung cancer and they were more common in patients with lung adenocarcinoma. Gene mutation and PDL1 expression are high in patients with lung adenocarcinoma.

Controlling the Colonization of Clostridium perfringens in Broiler Chickens by an Electron-Beam-Killed Vaccine.

Clostridium perfringens (Cp) is a Gram-positive anaerobe that is one of the causative agents of necrotic enteritis (NE) in chickens, which leads to high mortality. Owing to the ban of administering antibiotics in feed to chickens, there has been an increase in the number of NE outbreaks all over the world, and the estimated loss is approximately 6 billion U.S. dollars. The best alternative method to control NE without antibiotics could be vaccination. In this study, we exposed three different strains of Cp to electron beam (eBeam) irradiation to inactivate them and then used them as a killed vaccine to control the colonization of Cp in broiler chickens. The vaccine was delivered to 18-day old embryos in ovo and the chickens were challenged with the respective vaccine strain at two different time points (early and late) to test the protective efficacy of the vaccine. The results indicate that an effective eBeam dose of 10 kGy inactivated all three strains of Cp, did not affect the cell membrane or epitopes, induced significant levels of IgY in the vaccinated birds, and further reduced the colonization of Cp strains significantly (p < 0.0001) in late challenge (JGS4064: 4 out of 10; JGS1473: 0 out of 10; JGS4104: 3 out of 10). Further studies are necessary to enhance the efficacy of the vaccine and to understand the mechanism of vaccine protection.

Evaluation of Thymol-ß-d-Glucopyranoside as a Potential Prebiotic Intervention to Reduce Carriage of Zoonotic Pathogens in Weaned and Feeder Pigs.

The gut of food-producing animals is a reservoir for foodborne pathogens. Thymol is bactericidal against foodborne pathogens but rapid absorption of thymol from the proximal gut precludes the delivery of effective concentrations to the lower gut where pathogens mainly colonize. Thymol-ß-d-glucopyranoside is reported to be more resistant to absorption than thymol in everted jejunal segments and could potentially function as a prebiotic by resisting degradation and absorption in the proximal gut but being hydrolysable by microbial ß-glycosidase in the distal gut. Previous in vitro studies showed bactericidal effects of thymol-ß-d-glucopyranoside against Campylobacter, Escherichia coli, and Salmonella enterica serovar Typhimurium in the presence but not absence of intestinal microbes expressing ß-glycosidase activity, indicating that hydrolysis was required to obtain antimicrobial activity. Presently, the oral administration of thymol-ß-d-glucopyranoside was studied to examine the effects on intestinal carriage of Campylobacter, E. coli, and S. Typhimurium in swine. The effects of thymol-ß-d-glucopyranoside or thymol on antimicrobial sensitivity of representative E. coli isolates and characterized Salmonella strains were also explored. Results from two in vivo studies revealed little antimicrobial effects of thymol-ß-d-glucopyranoside on Campylobacter, E. coli, or S. Typhimurium in swine gut. These findings add credence to current thinking that hydrolysis and absorption of thymol-ß-d-glucopyranoside and thymol may be sufficiently rapid within the proximal gut to preclude delivery to the distal gut. Antibiotic susceptibilities of selected bacterial isolates and strains were mainly unaffected by thymol. Further research is warranted to overcome obstacles, preventing the delivery of efficacious amounts of thymol-ß-d-glucopyranoside to the lower gut.

Inhibition of Salmonella Binding to Porcine Intestinal Cells by a Wood-Derived Prebiotic.

Numerous Salmonella enterica serovars can cause disease and contamination of animal-produced foods. Oligosaccharide-rich products capable of blocking pathogen adherence to intestinal mucosa are attractive alternatives to antibiotics as these have potential to prevent enteric infections. Presently, a wood-derived prebiotic composed mainly of glucose-galactose-mannose-xylose oligomers was found to inhibit mannose-sensitive binding of select Salmonella Typhimurium and Escherichia coli strains when reacted with Saccharomyces boulardii. Tests for the ability of the prebiotic to prevent binding of a green fluorescent protein (GFP)-labeled S. Typhimurium to intestinal porcine epithelial cells (IPEC-J2) cultured in vitro revealed that prebiotic-exposed GFP-labeled S. Typhimurium bound > 30% fewer individual IPEC-J2 cells than did GFP-labeled S. Typhimurium having no prebiotic exposure. Quantitatively, 90% fewer prebiotic-exposed GFP-labeled S. Typhimurium cells were bound per individual IPEC-J2 cell compared to non-prebiotic exposed GFP-labeled S. Typhimurium. Comparison of invasiveness of S. Typhimurium DT104 against IPEC-J2 cells revealed greater than a 90% decrease in intracellular recovery of prebiotic-exposed S. Typhimurium DT104 compared to non-exposed controls (averaging 4.4 ± 0.2 log10 CFU/well). These results suggest compounds within the wood-derived prebiotic bound to E. coli and S. Typhimurium-produced adhesions and in the case of S. Typhimurium, this adhesion-binding activity inhibited the binding and invasion of IPEC-J2 cells.

A microencapsulated feed additive containing organic acids, thymol, and vanillin increases in vitro functional activity of peripheral blood leukocytes from broiler chicks.

During the first week after hatch, young chicks are vulnerable to pathogens as the immune system is not fully developed. The objectives of this study were to determine if supplementing the starter diet with a microencapsulated feed additive containing citric and sorbic acids, thymol, and vanillin affects in vitro functional activity of peripheral blood leukocytes (PBLs). Day-old chicks (n = 800) were assigned to either a control diet (0 g/metric ton [MT]) or a diet supplemented with 500 g/MT of the microencapsulated additive. At 4 D of age, peripheral blood was collected (100 birds per treatment), and heterophils and monocytes isolated (n = 4). Heterophils were assayed for the ability to undergo degranulation and production of an oxidative burst response while nitric oxide production was measured in monocytes. Select cytokine and chemokine mRNA expression levels were also determined. Statistical analysis was performed using Student t test comparing the supplemented diet to the control (P ≤ 0.05). Heterophils isolated from chicks fed the microencapsulated citric and sorbic acids, thymol, and vanillin had higher (P ≤ 0.05) levels of degranulation and oxidative burst responses than those isolated from chicks on the control diet. Heterophils from the supplemented chicks also had greater (P ≤ 0.05) expression of IL10, IL1ß, and CXCL8 mRNA than those from control-fed chicks. Similarly, nitric oxide production was significantly (P ≤ 0.05) higher in monocytes isolated from birds fed the supplement. The cytokine and chemokine profile in monocytes from the supplement-fed chicks showed a significant (P ≤ 0.05) drop in IL10 mRNA expression while IL1ß, IL4, and CXCL8 were unchanged. In conclusion, 4 D of supplementation with a microencapsulated blend made up of citric and sorbic acids, thymol, and vanillin enhanced the in vitro PBL functions of degranulation, oxidative burst, and nitric oxide production compared with the control diet. Collectively, the data suggest feeding broiler chicks a diet supplemented with a microencapsulated blend of citric and sorbic acids, thymol, and vanillin may prime key immune cells making them more functionally efficient and acts as an immune-modulator to boost the inefficient and undeveloped immune system of young chicks.

Evaluation of the Effect of Fiber Type, Length, and Content on Asphalt Properties and Asphalt Mixture Performance.

Fiber-reinforced asphalt mixture has been widely used in pavement engineering to not only prevent asphalt binder leakage but also improve engineering properties of asphalt mixture. However, the research on three key parameters, namely fiber type, fiber length, and fiber content, which significantly affect the performance of fiber-reinforced asphalt mixture, have seldom been conducted systematically. To determine these three key parameters in the support of the application of fibers in mixture scientifically, three commonly used fibers were selected, basalt fiber, polyester fiber, and lignin fiber, and the testing on fibers, fiber-reinforced asphalt binders, and fiber-reinforced asphalt mixtures was conducted afterwards. The results showed: the favorable fiber type was basalt fiber; the favorable basalt fiber length was 6mm; the engineering properties including high temperature stability, low temperature crack resistance, and water susceptibility were clearly improved by the added basalt fiber, and the optimum basalt fiber content was 0.4 wt.%. The obtained results may be valuable from a practical point of view to engineers and practitioners.

Trends in the incidence and survival of patients with esophageal cancer: A SEER database analysis.

BACKGROUND: Recent studies have indicated that the incidence of esophageal cancer has declined in the past decade in the U.S. However, trends in the incidence and survival have not been thoroughly examined. METHODS: Data from 46 063 patients with esophageal cancer between 1973 and 2015 were collected from the Surveillance, Epidemiology, and End Results database. The trends in the age-adjusted incidence and survival were analyzed using joinpoint regression models. RESULTS: The age-adjusted incidence of esophageal cancer increased from 5.55 to 7.44 per 100 000 person-years between 1973 and 2004. Later, it decreased at an annual percentage change of 1.23%. In the last 40 years, the strong male predominance increased slightly. Importantly, the percentage of patients with localized stage of squamous cell cancer decreased. It was observed that the incidence of esophageal squamous cell carcinoma declined since 1986, while the incidence of esophageal adenocarcinoma sharply increased since 1973 and surpassed the rate of squamous cell cancer, mainly due to the increase in the incidence among men. Consistently, the estimated 40-year limited-duration prevalence of esophageal adenocarcinoma was higher than that of esophageal squamous cell carcinoma. Additionally, we observed a modest but significant improvement in survival during the study period. CONCLUSION: The incidence of esophageal squamous cell carcinoma has decreased significantly over the past four decades in the U.S., while the incidence of adenocarcinoma has increased, particularly among men. Overall, the long-term survival of patients with esophageal cancer is poor but it has improved over the past decades, especially for the localized disease. KEY POINTS: Significant findings of the study The incidence of esophageal cancer has decreased at an annual percentage change of 1.23% since 2004. The incidence of esophageal adenocarcinoma has sharply increased since 1973 and surpassed the rate of squamous cell cancer, mainly due to the increase in the incidence among men. What this study adds There has been a shift in the prevalence of esophageal cancer histological subtypes over the past decades in the U.S. We found that the incidence of esophageal squamous cell carcinoma has continued to decrease, while the esophageal adenocarcinoma rate has continued to increase.

Genetic polymorphisms of IL-10, IL-18 and IL12B are associated with risk of non-small cell lung cancer in a Chinese Han population.

PURPOSE: IL-10, IL-18 and IL-12 are reported to participate in the inflammation process. The potential influences of IL-10, IL-18 and IL-12 polymorphisms on non-small cell lung cancer (NSCLC) risk were explored in this study. METHODS: Six candidate SNPs from 500 NSCLC patients and 500 controls were genotyped. The correlation between the SNPs and NSCLC risk was evaluated by logistic regression analysis. RESULTS: Comparisons of the allele and genotype frequencies showed that five SNPs were correlated with NSCLC risk. The minor allele 'G' of IL-18 rs5744256 and rs1834481 and IL-10 rs3021094 was correlated with a decreased risk of NSCLC (p < 0.05). In contrast, the minor allele 'T' of IL-18 rs5744224 and the minor allele 'G' of IL-12B rs3212227 were correlated with an increased risk of NSCLC (p < 0.05). By genetic model analysis, we found that rs5744256 and rs1834481 were associated with a decreased risk of NSCLC under dominant and log-additive models (p < 0.05). Rs3021094 was correlated with a decreased risk of NSCLC under all three models (p < 0.05). In contrast, rs5744224 was associated with an increased risk of NSCLC under the recessive model (p = 0.005), and rs3212227 was associated with an increased risk of NSCLC under all three models (p < 0.05). Finally, the GGA haplotype of rs5744256, rs1834481 and rs5744224 and the GT haplotype of rs3021094 and rs3790622 were associated with a decreased risk of NSCLC (p < 0.05). CONCLUSION: Our results provided new candidate SNPs for the prediction and prevention of NSCLC.

Administration of a Postbiotic Causes Immunomodulatory Responses in Broiler Gut and Reduces Disease Pathogenesis Following Challenge.

With the reemergence of poultry diseases such as necrotic enteritis following the restriction of in-feed antibiotics, the search for antibiotic alternatives has become critically important. Postbiotics are non-viable bacterial products or metabolic byproducts from probiotic microorganisms that have positive effects on the host or microbiota. These are a promising alternative to antibiotics. Here, we describe the mechanism of action of a postbiotic in the context of a Clostridium perfringens (C. perfringens) challenge model. By using performance measurements and a peptide array kinome analysis, we describe the kinotypes and signal transduction changes elicited by the postbiotic with and without C. perfringens challenge. The postbiotic improves lesion scores, C. perfringens counts and mortality compared to challenge groups without the postbiotic, and it improves weight gain in the most severely challenged birds. The postbiotic predominantly affects the innate immune response and appears immunomodulatory. In the context of infection, it reduces the proinflammatory responses and generates a homeostatic-like response. This postbiotic is a viable alternative to antibiotics to improve poultry health in the context of C. perfringens pathogen challenge.

Inhibition of calmodulin increases intracellular survival of Salmonella in chicken macrophage cells.

Calcium (Ca2+) is a pivotal intracellular second messenger and calmodulin (CaM) acts as a multifunctional Ca2+-binding protein that regulates downstream Ca2+ dependent signaling. Together they play an important role in regulating various cellular functions, including gene expression, maturation of phagolysosome, apoptosis, and immune response. Intracellular Ca2+ has been shown to play a critical role in Toll-like receptor-mediated immune response to microbial agonists in the HD11 chicken macrophage cell line. The role of that the Ca2+/CaM pathway plays in the intracellular survival of Salmonella in chicken macrophages has not been reported. In this study, kinome peptide array analysis indicated that the Ca2+/CaM pathway was significantly activated when chicken macrophage HD11 cells were infected with S. Enteritidis or S. Heidelberg. Further study demonstrated that treating cells with a pharmaceutical CaM inhibitor W-7, which disrupts the formation of Ca2+/CaM, significantly inhibited macrophages to produce nitric oxide and weaken the control of intracellular Salmonella replication. These results strongly indicate that CaM plays an important role in the innate immune response of chicken macrophages and that the Ca2+/CaM mediated signaling pathway is critically involved in the host cell response to Salmonella infection.