Burst Of Production Research Articles

Eosinophil expression of triggering receptor expressed on myeloid cells 1 (TREM-1) restricts type 2 lung inflammation.

Asthma affects 25 million Americans, and recent advances in treatment are effective for only a portion of severe asthma patients. TREM-1, an innate receptor that canonically amplifies inflammatory signaling in neutrophils and monocytes, plays a central role in regulating lung inflammation. It is unknown how TREM-1 contributes to allergic asthma pathology. Utilizing a murine model of asthma, flow cytometry revealed TREM-1+ eosinophils in the lung tissue and airway during allergic airway inflammation. TREM-1 expression was restricted to recruited, inflammatory eosinophils. Expression was induced on bone marrow-derived eosinophils by incubation with interleukin 33, lipopolysaccharide, or granulocyte-macrophage colony-stimulating factor. Compared to TREM-1- airway eosinophils, TREM-1+ eosinophils were enriched for proinflammatory gene sets, including migration, respiratory burst, and cytokine production. Unexpectedly, eosinophil-specific ablation of TREM-1 exacerbated airway interleukin (IL) 5 production, airway MUC5AC production, and lung tissue eosinophil accumulation. Further investigation of transcriptional data revealed apoptosis and superoxide generation-related gene sets were enriched in TREM-1+ eosinophils. Consistent with these findings, annexin V and caspase-3/7 staining demonstrated higher rates of apoptosis among TREM-1+ eosinophils compared to TREM-1- eosinophils in the inflammatory airway. In vitro, Trem1/3-/- bone marrow-derived eosinophils consumed less oxygen than wild-type in response to phorbol myristate acetate, suggesting that TREM-1 promotes superoxide generation in eosinophils. These data reveal protein-level expression of TREM-1 by eosinophils, define a population of TREM-1+ inflammatory eosinophils, and demonstrate that eosinophil TREM-1 restricts key features of type 2 lung inflammation.

Porcine Monocyte DNA Traps Formed during Infection with Pathogenic Clostridioides difficile Strains.

Clostridioides (Clostridium) difficile is an enteric pathogen of several mammalian species including man, frequently involving nosocomial resurgence, following oral administration of broad-spectrum antibiotics, but also with human-to-human infection occurring, and neonatal pigs with zoonotic transmission. To date, the immune response to C. difficile has mostly focused on neutrophils and cytokine/chemokines, particularly in human infection. The neonatal pig is now recognized as a valuable model for human infection. We show that porcine monocytes respond to C. difficile differently compared with many other bacterial infections. Infection of porcine monocytes with human C. difficile strains CD630 (Ribotype 078) or R20291 (Ribotype 027) for 3 or 24 h post-infection (pi) resulted in a lack of oxidative burst or nitrite ion production when compared to uninfected controls (p > 0.05). The survival dynamics of both CD630 and R20291 in monocytes were similar with intracellular bacterial numbers being similar at 3 h pi and 24 h pi (p > 0.05). However, we show that porcine monocytes entrap C. difficile via extracellular DNA traps. This process began as early as 3 h pi, and at 24 h pi the nuclei appeared to be depleted of DNA, although extracellular DNA was associated with the cell membrane. Our preliminary study also suggests that entrapment of C. difficile by extracellular DNA may occur via a process of monocyte etosis.

Cyrtocarpa edulis fruit and its immunostimulant effect on Almaco Jack Seriola rivoliana: in vitro, in vivo and ex vivo studies.

The present study investigates for the first time chemical, proximate analyses and immunostimulant effect of Cyrtocarpa edulis fruit (CeF). Three design experiments were carried out to evaluate immunostimulant effect of C. edulis fruit: in vitro, in vivo and ex vivo studies in juveniles Almaco jack Seriola rivoliana. In general, nutraceutical studies performed by gas chromatography/mass spectrometry (GC-MS) in CeF revealed a major quantity of the carbohydrate groups and phytosterols such as β-sitosterol. Their phytochemical and antioxidant values exposed a significant content of total phenols, flavonoids, and tannins, showing an antioxidant capacity against hydroxyl and superoxide radical. The in vitro results confirm that CeF is edible and enhanced the innate immune response in head-kidney leukocytes after 24h of immunostimulation. The in vivo results showed that myeloperoxidase, nitric oxide production, as well as antioxidant enzymes were enhanced in skin mucus of those fish fed with CeF. Interestingly in the intestine, IL-β, TNF-α, MARCO and Piscidin gene expression were up-regulated in fish fed with C. edulis after 4weeks. Finally, ex vivo experiments showed an important enhancement on cellular parameters (phagocytosis, respiratory burst, myeloperoxidase, and nitric oxide production) in head-kidney leukocytes of fish fed CeF and intraperitoneally infected with A. hydrophila. The results demonstrate that C. edulis fruit (0.5%) represents an available phytochemical and antioxidant rich alternative with great potential as fish immunostimulant additive.

Development, characterization, and immunomodulation performance of spray-dried Moringa oleifera seed extract in Longfin yellowtail Seriola rivoliana.

Moringa oleifera is one of the most promising plants in aquaculture because it improves the health status, zootechnical parameters and resistance against diseases. This research evaluates the physicochemical, antioxidant values of spray-dried Moringa oleifera seed extract microencapsulates obtained at 140 and 180°C with whey protein concentrate (WPC) and maltodextrin (MD) as wall materials in two different proportions: WPC 100% and WPC-MD (3:1). Also, immune response of peripheral blood leukocytes (PBL) of Longfin yellowtail Seriola rivoliana stimulated with spray-dried Moringa oleifera seed for 24h was assessed. The physicochemical parameters show that the recovery yield for all the treatments was of 65% and microencapsulates demonstrated to be stable in the physicochemical tests with low solubilization times and protection against humidity. For WPC-MD (3:1)/140°C, bioactive compound retention and antioxidant potential were higher than in other combinations. The immunological test show that any treatments was non-cytotoxic against peripheral blood leukocytes. WPC-MD (3:1)/140°C treatment enhanced immune parameters as phagocytosis, respiratory burst, myeloperoxidase activities and nitric oxide production. Immune related genes as IL-1β and TNF-α were up-regulated in those stimulated leukocytes with WPC-MD (3:1)/140°C. The results suggest that this combination may be a good alternative for animal health as a medicinal and immunostimulant additive.

Isolation of Mouse Neutrophils.

Neutrophils represent the first line of defense against bacterial and fungal pathogens. Indeed, patients with inherited or acquired qualitative and quantitative neutrophil defects are at high risk for developing bacterial and fungal infections and suffering adverse outcomes from these infections. Therefore, research aiming at defining the molecular factors that modulate neutrophil effector function under homeostatic conditions and during infection is essential for devising strategies to augment neutrophil function and improve the outcomes of infected individuals. This article describes reproducible density-gradient-centrifugation-based as well as positive and negative immunomagnetic selection protocols that can be applied in any laboratory to harvest large numbers of highly enriched and highly viable neutrophils from the bone marrow of mice. In another protocol, we also present a method that combines gentle enzymatic tissue digestion with a positive immunomagnetic selection technique or fluorescence-activated cell sorting (FACS) to harvest highly pure and highly viable preparations of neutrophils directly from mouse tissues such as the kidney, the liver, or the spleen. Mouse neutrophils isolated by these protocols can be used to examine several aspects of cellular function ex vivo, including pathogen binding, phagocytosis, and killing, neutrophil chemotaxis, oxidative burst, degranulation, and cytokine production, and for performing neutrophil adoptive transfer experiments. © 2023 Wiley Periodicals LLC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA. Basic Protocol 1: Isolation of Neutrophils from Mouse Bone Marrow Using Positive Immunomagnetic Separation Alternate Protocol 1: Purification of Neutrophils from Bone Marrow Using Negative Immunomagnetic Separation Alternate Protocol 2: Purification of Neutrophils from Bone Marrow Using Histopaque-Based Density Gradient Centrifugation Basic Protocol 2: Isolation of Neutrophils from Mouse Tissues Using Positive Immunomagnetic Separation Alternate Protocol 3: Isolation of Neutrophils from Mouse Tissues Using FACS.

Glycation derived AuNPs bioconjugated novel herbal drug isoferulic acid: As a potential anti-glycation, anti-diabetic and antineoplastic agent

Based on the significance and impact of serum protein glycation and the possibility that AGEs play a role in diabetes-related complications and cancer, this study examined the anti-glycation, anti-cancer, and α-amylase inhibition activities of isoferulic acid (IFA) and its gold nanoparticle (IFA-AuNPs) bioconjugated formulations. In comparison to the control (glycated) samples, IFA and IFA-AuNPs were found to show a significant reduction in the KM moieties, carbonyl content, and HMF content that were produced during the glycation, demonstrating that the glycation reaction generated less superoxide radicals, had fewer intermediates, and did not cause as many structural perturbations to BSA. The IFA and IFA-AuNPs treated samples had a higher proportion of unreacted lysine and arginine deposits than the control (glycated) samples, suggesting that the inhibitors prevented the association of negatively charged arginine deposits with 2′-Deoxyribose. Purified IFA (IC50 = 4.3 mM) decreased α-amylase activity insignificantly compared to acarbose (IC50 = 0.118 mM). Therefore, IFA may not be suited for targeting α-amylase. However, IFA and its analogs have been reported to inhibit several malignancies, including leukemia, breast, colon, and lung cancer. Therefore, the cytotoxicity study was conducted using human keratinocytes (HaCat cells), and the IC50 values for IFA and IFA-AuNPs were determined to be 6062.4 µM and 151.6 µM, respectively. Their cytotoxic effects on human lung cancer epithelial cells (A549), however, were estimated to be 5593 µM and 117.5 µM. Our study confirmed that IFA-treated A549 cells at a greater concentration (5593 µM) caused less disruption of mitochondrial membrane potential (ΔΨm) and production of ROS burst than IFA-AuNPs at a significantly lower concentration (117,5 µM). When ΔΨm was disrupted, Cyt-c-mediated apoptosis was elicited, which then allowed IFA and AuNPs to work synergistically to degrade DNA. It was discovered that IFA-AuNPs target cancer cells through different pathways, with AuNPs potentiating the effect of IFA.

Polymorphism in neutrophil cytosolic factor 4 (NCF4) of dairy cows had mastitis in previous lactations, and the relationship with the respiratory burst

Nicotinamide adenine dinucleotide phosphate oxidase (NADPH oxidase), as a key factor in innate immunity, consists of several components, one of them is p40phox which is encoded by neutrophil cytosolic factor 4 (NCF4). Respiratory burst and reactive oxygen species (ROS) production are antimicrobial mechanisms associated with NADPH oxidase. This study evaluated the effects of g.18174 A > G and g.18270C > T single-nucleotide polymorphisms (SNP) in NCF4 on bovine mastitis and the respiratory burst capacity of neutrophils. SNPs of 160 dairy cattle were determined using a novel PCR-RFLP protocol by employing restriction enzymes, MboI and FokI. Also, the flow cytometry measured respiratory burst in 82 blood samples. Our results indicated that only g.18174 A > G SNP reduced the respiratory burst capacity. However, both SNPs were not significantly correlated with clinical mastitis. We concluded that g.18174 A > G decreases the function of NADPH oxidase. However, both SNPs were not significantly correlated with clinical mastitis.

Slow negative feedback enhances robustness of square-wave bursting

Square-wave bursting is an activity pattern common to a variety of neuronal and endocrine cell models that has been linked to central pattern generation for respiration and other physiological functions. Many of the reduced mathematical models that exhibit square-wave bursting yield transitions to an alternative pseudo-plateau bursting pattern with small parameter changes. This susceptibility to activity change could represent a problematic feature in settings where the release events triggered by spike production are necessary for function. In this work, we analyze how model bursting and other activity patterns vary with changes in a timescale associated with the conductance of a fast inward current. Specifically, using numerical simulations and dynamical systems methods, such as fast-slow decomposition and bifurcation and phase-plane analysis, we demonstrate and explain how the presence of a slow negative feedback associated with a gradual reduction of a fast inward current in these models helps to maintain the presence of spikes within the active phases of bursts. Therefore, although such a negative feedback is not necessary for burst production, we find that its presence generates a robustness that may be important for function.

In vitro study of deltamethrin-induced extracellular traps in hemocytes of Ruditapes philippinarum

Deltamethrin (DLM), a broad-spectrum pesticide, has been proven to have toxic effects on aquatic organisms. Here, we detected the formation of extracellular traps (ETosis) formation in Manila clam (Ruditapes philippinarum) hemocytes stimulated by three concentrations of DLM (0.01, 0.1 and 1 μg/mL) in vitro, and explored the underlying mechanisms induced by this pesticide. Extracellular DNA structure observation and quantitative results indicated that DLM exposure could obviously induce hemocytes ETosis, especially under high concentration of DLM induction. Moreover, DLM increased the levels of myeloperoxidase (MPO) and reactive oxygen species (ROS) in a dose-dependent manner, and enhanced the mRNA expression of several ROS-related genes. DPI (NADPH oxidase inhibitor) and ABAH (MPO inhibitor) could substantially inhibit DLM-induced extracellular traps (ETs), suggesting that the induced ETs release was caused by the induction of the ROS burst and MPO production. In addition, three concentrations of DLM-induced ETs were also accompanied by mitochondrial dysfunction, such as increasing the production of mitochondrial ROS, leading to a decrease in mitochondrial membrane potential (MMP) and activation of mitochondrial permeability transition pore (MPTP). Taken together, these results will shed new light on the immunotoxicity of DLM in clams and perhaps lays the foundation for health assessment in bivalves.

Hydroxy-Selenomethionine, an Organic Selenium Source, Increases Selenoprotein Expression and Positively Modulates the Inflammatory Response of LPS-Stimulated Macrophages.

The role of 2-hydroxy-(4-methylseleno)butanoic acid (OH-SeMet), a form of organic selenium (Se), in selenoprotein synthesis and inflammatory response of THP1-derived macrophages stimulated with lipopolysaccharide (LPS) has been investigated. Glutathione peroxidase (GPX) activity, GPX1 gene expression, selenoprotein P (SELENOP) protein and gene expression, and reactive oxygen species (ROS) production were studied in Se-deprived conditions (6 and 24 h). Then, macrophages were supplemented with OH-SeMet for 72 h and GPX1 and SELENOP gene expression were determined. The protective effect of OH-SeMet against oxidative stress was studied in H2O2-stimulated macrophages, as well as the effect on GPX1 gene expression, oxidative stress, cytokine production (TNFα, IL-1β and IL-10), and phagocytic and killing capacities after LPS stimulation. Se deprivation induced a reduction in GPX activity, GPX1 gene expression, and SELENOP protein and gene expression at 24 h. OH-SeMet upregulated GPX1 and SELENOP gene expression and decreased ROS production after H2O2 treatment. In LPS-stimulated macrophages, OH-SeMet upregulated GPX1 gene expression, enhanced phagocytic and killing capacities, and reduced ROS and cytokine production. Therefore, OH-SeMet supplementation supports selenoprotein expression and controls oxidative burst and cytokine production while enhancing phagocytic and killing capacities, modulating the inflammatory response, and avoiding the potentially toxic insult produced by highly activated macrophages.

Noise reduction by upstream open reading frames

Gene expression is prone to burst production, making it a highly noisy process that requires additional controls. Upstream open reading frames (uORFs) are widely present in the 5′ leader sequences of 30–50% of eukaryotic messenger RNAs1–3. The translation of uORFs can repress the translation efficiency of the downstream main coding sequences. Whether the low translation efficiency leads to a different variation, or noise, in gene expression has not been investigated, nor has the direct biological impact of uORF-repressed translation. Here we show that uORFs achieve low but precise protein production in plant cells, possibly by reducing the protein production rate. We also demonstrate that, by buffering a stable TIMING OF CAB EXPRESSION 1 (TOC1) protein production level, uORFs contribute to the robust operation of the plant circadian clock. Our results provide both an action model and the biological impact of uORFs in translational control to mitigate transcriptional noise for precise protein production.

Protection and Pathology in Leishmania braziliensis Infection.

Leishmania killing is mediated by IFN-γ-activated macrophages, but IFN-γ production and macrophage activation are insufficient to control L. braziliensis infection. In American tegumentary leishmaniasis (ATL), pathology results from an exaggerated inflammatory response. This report presents an overview of our contributions regarding ATL pathogenesis, highlighting future directions to improve the management of L. braziliensis infection. Monocytes and lymphocytes from individuals exposed to L. braziliensis but who do not develop CL, i.e., subclinical infection (SC), exhibit lower respiratory burst and IFN-γ production, yet more efficiently kill L. braziliensis. As vaccines aimed at inducing IL-12 and IFN-γ do not sufficiently prevent CL, the elucidation of how subjects with SC infection kill Leishmania may lead to new approaches to controlling ATL. While inflammation arising from the recruitment of inflammatory cells via chemokines induced by IFN-γ and TNF or IL-17 is observed and contributes to pathology, cytotoxic CD8+ T cells and NK cells play a key role in the pathogenesis of L. braziliensis infection. The increased transcription of genes related to inflammation and cytotoxicity, e.g., granzyme A, granzyme B, NLRP3 and IL-1β, has been documented in CL tissue samples. The release of products by killed cells leads to NLRP3 inflammasome activation, IL-1β production and additional damage to skin and mucosal tissues. The use of drugs that downmodulate the inflammatory response in combination with chemotherapy improves the ATL cure rate and decreases healing time.

The Antibacterial Activity of Erythrocytes From Goose (Anser domesticus) Can Be Associated With Phagocytosis and Respiratory Burst Generation.

In the lumen of blood vessels, there are large numbers of erythrocytes, which are approximately 95% of the total blood cells. Although the function of erythrocytes is to transport oxygen in the organism, recent studies have shown that mammalian and teleost erythrocytes are involved in the immune response against bacterial infections. However, the immune mechanisms used by avian erythrocytes are not yet clear. Here, we demonstrated that erythrocytes from goose have the ability to phagocytose as well as conduct antimicrobial activity. Firstly, we revealed the phagocytosis or adhesion activity of goose erythrocytes for latex beads 0.1-1.0 μm in diameter by fluorescence microscopy, and scanning and transmission electron microscopy. The low cytometry results also proved that goose erythrocytes had a wide range of phagocytic or adhesion activity for different bacteria. Followed, the low cytometry analysis data further explored that the goose erythrocytes contain the ability to produce reactive oxygen species (ROS) and inducible nitric oxide synthase (iNOS) in response to bacterial stimulation, and also up-regulated the expression of NOX family includes NOX1 and NOX5. Finally, we also found that goose erythrocytes showed a powerful antibacterial activity against all the three bacteria, meanwhile the stimulation of three kinds of bacteria up-regulated the expression of inflammatory factors, and increased the production of antioxidant enzymes to protect the cells from oxidative damage. Herein, our results demonstrate that goose Erythrocytes possess a certain phagocytic capacity and antioxidant system, and that the antimicrobial activity of erythrocytes can occurred through the production of unique respiratory burst against foreign pathogenic bacteria, which provides new clues to the interaction between bacteria and avian erythrocytes.

Blockade of TLR2 and TLR4 Attenuates Inflammatory Response and Parasite Load in Cutaneous Leishmaniasis.

Human cutaneous leishmaniasis (CL) caused by Leishmania braziliensis is characterized by a pronounced inflammatory response associated with ulcer development. Monocytes/macrophages, the main cells harboring parasites, are largely responsible for parasite control. Toll-like receptor (TLR) signaling leads to the transcription of inflammatory mediators, such as IL-1β and TNF during innate immune response. TLR antagonists have been used in the treatment of inflammatory disease. The neutralization of these receptors may attenuate an exacerbated inflammatory response. We evaluated the ability of TLR2 and TLR4 antagonists to modulate host immune response in L. braziliensis-infected monocytes and cells from CL patient skin lesions. Following TLR2 and TLR4 neutralization, decreased numbers of infected cells and internalized parasites were detected in CL patient monocytes. In addition, reductions in oxidative burst, IL-1β, TNF and CXCL9 production were observed. TNF production by cells from CL lesions also decreased after TLR2 and TLR4 neutralization. The attenuation of host inflammatory response after neutralizing these receptors suggests the potential of TLR antagonists as immunomodulators in association with antimonial therapy in human cutaneous leishmaniasis.

The Pleiades are a cluster of fungal effectors that inhibit host defenses.

Biotrophic plant pathogens secrete effector proteins to manipulate the host physiology. Effectors suppress defenses and induce an environment favorable to disease development. Sequence-based prediction of effector function is impeded by their rapid evolution rate. In the maize pathogen Ustilago maydis, effector-coding genes frequently organize in clusters. Here we describe the functional characterization of the pleiades, a cluster of ten effector genes, by analyzing the micro- and macroscopic phenotype of the cluster deletion and expressing these proteins in planta. Deletion of the pleiades leads to strongly impaired virulence and accumulation of reactive oxygen species (ROS) in infected tissue. Eight of the Pleiades suppress the production of ROS upon perception of pathogen associated molecular patterns (PAMPs). Although functionally redundant, the Pleiades target different host components. The paralogs Taygeta1 and Merope1 suppress ROS production in either the cytoplasm or nucleus, respectively. Merope1 targets and promotes the auto-ubiquitination activity of RFI2, a conserved family of E3 ligases that regulates the production of PAMP-triggered ROS burst in plants.

Leishmania donovani Targets Host Transcription Factor NRF2 To Activate Antioxidant Enzyme HO-1 and Transcriptional Repressor ATF3 for Establishing Infection

We showed previously that antioxidant enzyme heme oxygenase 1 (HO-1) is critical for Leishmania survival in visceral leishmaniasis. HO-1 inhibits host oxidative burst and inflammatory cytokine production, leading to parasite persistence. In the present study, screening of reported HO-1 transcription factors revealed that infection upregulated (4.1-fold compared to control [P < 0.001]) nuclear factor erythroid 2 (NFE2)-related factor 2 (NRF2). Silencing of NRF2 reduced both HO-1 expression and parasite survival. Investigation revealed that infection-induced transient reactive oxygen species (ROS) production dissociated NRF2 from its inhibitor KEAP1 and enabled phosphorylation-dependent nuclear translocation. Both NRF2 and HO-1 silencing in infection increased production of proinflammatory cytokines. But the level was greater in NRF2-silenced cells than in HO-1-silenced ones, suggesting the presence of other targets of NRF2. Another stress responsive transcription factor ATF3 is also induced (4.6-fold compared to control [P < 0.001]) by NRF2 during infection. Silencing of ATF3 reduced parasite survival (59.3% decrease compared to control [P < 0.001]) and increased proinflammatory cytokines. Infection-induced ATF3 recruited HDAC1 into the promoter sites of tumor necrosis factor alpha (TNF-α) and interleukin 12b (IL-12b) genes. Resulting deacetylated histones prevented NF-κB promoter binding, thereby reducing transcription of inflammatory cytokines. Administering the NRF2 inhibitor trigonelline hydrochloride to infected BALB/c mice resulted in reduced HO-1 and ATF3 expression, decreased spleen and liver parasite burdens, and increased proinflammatory cytokine levels. These results suggest that Leishmania upregulates NRF2 to activate both HO-1 and ATF3 for disease progression.

A sensitive chemiluminescence method for quantification of the oxidative burst in grapevine cells and rice roots

Roots are prominent plant-microbe interaction sites and of great biological relevance for many studies. The root response is of interest when searching for potential systemic resistance inducers. Screening of elicitors often focuses on the oxidative burst, the rapid and transient production of Reactive Oxygen Species (ROS). However, to our knowledge, no high-throughput, sensitive methods have been developed for the quantification of ROS released by roots. Here, we report on the development of an L-012-based chemiluminescence bioassay to quantitatively determine the oxidative burst following elicitation events in roots. Rice and grapevine were used as monocot and dicot models. We demonstrate that chitosan, a recognized elicitor in rice cells, was able to elicit ROS production in rice roots. Chitosan also triggered a strong oxidative burst in grapevine cell suspension cultures, while grapevine roots were not responsive. Although this method is broadly applicable, the L-012 probe requires careful consideration of solvents and plant species. Insufficient extracellular ROS, quenching, and the interference of solvents with the probe can undermine the assay sensitivity.

Candida Albicans Activated Splenocytes Promote Strong Immune Responses in a Murine Model of Breast Cancer

Background: The potential of Candida albicans to modulate antigen-presenting cells maturation has been documented in past studies. Dendritic cells are critical modulators in the orchestration of adaptive immune responses alongside myeloid subtypes, which play an important role in the presentation of antigens to T cells. The aim of this study was to evaluate the efficacy of splenocytes activated with the extract of heated 4T1 cells and the yeast form of C. albicans against breast cancer growth in vivo. Methods: 4T1 cells were subcutaneously injected into the left flanks of female BALB/c mice (n=40). At a time when palpable tumors had developed, experimental groups were immunized twice at one-week interim with either activated splenocytes with the extract of heated 4T1 or the killed preparation of yeast form of C. albicans or a combination of the two-One week after the second injection, one-half of animals (n=20) were euthanized to investigate the immune response profile. Results: Administration of activated splenocytes with the combination protocol caused a favorable survival curve and slower rates of tumor development compared to other tumor-bearing mice. Moreover, combination therapy significantly increased the secretion of IFN-γ, respiratory burst and nitric oxide production and conversely diminished the secretion of IL-4, IL-10 and TGF-β in the splenocyte population. Conclusion: Since the murine 4T1 cell line is similar to the final stage of human breast carcinoma, we postulate that activated splenocytes with the extract of heated 4T1 cells and yeast form of C. albicans can reduce tumor development in tumor-bearing mice.

Circulating Neutrophil Dysfunction in HBV-Related Acute-on-Chronic Liver Failure.

Background and AimsAcute-on-chronic liver failure (ACLF) is characterized by systemic inflammation accompanied by defective anti-bacterial immunity. The role of neutrophils in immune derangement of ACLF has not been fully elucidated. This study is aimed to characterize the role of circulating neutrophils in HBV-related ACLF patients.MethodsQuantitative, phenotypic, transcriptomic, and functional alterations of circulating neutrophils were compared in ACLF and non-ACLF subjects and analyzed for associations with short-term outcomes. Interventional experiments were performed to test the impact on ACLF-patient neutrophil function in vitro.ResultsCirculating absolute neutrophil count was significantly increased in patients with ACLF and was an independent risk factor for 28-day mortality. ACLF-patient neutrophils differentially expressed a panel of surface markers (include TLR-1, TLR-2, TLR-4, CEACAM-1 and FPR1), as well as a distinct transcriptomic signature. ACLF-neutrophils displayed significantly impaired phagocytosis but an increased capacity to form neutrophil extracellular traps (NETs), which was more pronounced in patients with poor outcome. Healthy neutrophils mimicked functional characteristics of ACLF counterpart after co-cultured with plasma from ACLF patients. The oxidative burst and cytokine production capacities remained unchanged. Plasma GM-CSF, IL-6, IL-8, IL-10, and IP-10 levels, as well as lipopolysaccharide (LPS) concentration, were markedly elevated in ACLF patients but not DAMP molecules HMGB-1 and HSP70. Finally, a glycolysis inhibitor, 2-deoxy-glucose, reduced NET formation of ACLF patients’ neutrophils.ConclusionsCirculating ACLF-patient neutrophils exhibit alterations in number, phenotype, gene expression and function, which was associated with poor outcome and shaped by the ACLF circulatory environment. Inhibiting glycolysis can reverse neutrophil dysfunction in ACLF patients.

Combination of Propranolol and Heated 4T1 Cells Elicits Beneficial Response Against Mouse Model of Breast Cancer

Background: Induction of Th1 responses against tumor antigens may be a useful strategy to control malignancy. In this respect, previous studies have shown that beta-adrenoreceptor antagonists can promote cellular immune responses. Objectives: This survey was done to evaluate the beneficiary of a new immunotherapy method against breast cancer made by mixing heated 4T1 cells and propranolol, as an adjuvant. Methods: Subcutaneously injected live 4T1 cells (1 × 104) were used to induce breast cancer in six to eight-week-old female Balb/c mice. when all mice had a palpable tumor, immunotherapy was dawned. Mice in the treatment groups were vaccinated, twice at a one-week interval, with the extract of heated 4T1(1 × 105) either alone or in combination with propranolol (6 mg/kg). Negative control mice received phosphate-buffered saline (PBS) as the same schedule. One-half of the mice were euthanized one week after the last vaccination to investigate the immune response profile. Other animals were kept until death occurred spontaneously. Results: Combined immunotherapy with propranolol and extract of heated 4T1 had synergistic effects, causing a more desirable survival curve and slower tumor growth when compared to other tumor-bearing mice receiving only heated 4T1 or PBS. Furthermore, combined immunotherapy significantly augmented the production of IFN-γ nitric oxide production, respiratory burst, and cytotoxicity of natural killer cells in the splenocyte culture of tumor-bearing mice. Conversely, combined immunotherapy significantly regressed the production of TGF-β and IL-10 in the splenocyte population compared to cytokine production by splenocytes from other groups. Conclusions: Combined heated 4T1 cells with propranolol promote beneficial outcomes in the animal model of breast cancer.