Application Of Lipopolysaccharide Research Articles

Histopathological and biochemical Effects of 18β-glycyrrhetinic acid application on lipopolysaccharide-induced kidney toxicity in rats

Lipopolysaccharide (LPS) is an endotoxin found in the wall of gram-negative bacteria and causes acute inflammation when it enters the tissues. 18β-glycyrrhetinic acid (18β-GA) is a substance found in licorice root and is responsible for this plant's antiallergic, antioxidant, and anti-inflammatory activity. This study aimed to examine the possible effects of 18β-glycyrrhetinic acid on the damage caused by LPS in kidney tissue. The study divided 40 Sprague Dawley adult male rats into 5 equal groups (n = 8). The groups were created as follows; the control group; the group that received 1cc physiological saline throughout the experiment was the DMSO group; DMSO, an intraperitoneal carrier substance, was given. LPS group; A single dose of 7.5 mg/kg intraperitoneal (i.p) LPS was administered. 18β-GA50+LPS group; 18β-glycyrrhetinic acid was given by gavage at 50 mg/kg daily for 10 days, followed by a single dose of 7.5 mg/kg i.p. LPS was administered. 18β-GA100+LPS group; 18β-glycyrrhetinic acid was administered by gavage at 100 mg/kg daily for 10 days, followed by a single dose of 7.5 mg/kg i.p. LPS was administered. 18β-GA100 group; 18β-glycyrrhetinic was given by gavage at 100 mg/kg daily for 10 days. 24 hours after LPS application to all groups, the kidney tissues of the rats were removed under anesthesia and placed in 10% formaldehyde. Histopathological and oxidative stress parameters analyses were performed in kidney tissue.

Octreotide attenuates intestinal barrier damage by maintaining basal autophagy in Caco2 cells.

The intestinal mucosal barrier is of great importance for maintaining the stability of the internal environment, which is closely related to the occurrence and development of intestinal inflammation. Octreotide (OCT) has potential applicable clinical value for treating intestinal injury according to previous studies, but the underlying molecular mechanisms have remained elusive. This article is based on a cell model of inflammation induced by lipopolysaccharide (LPS), aiming to explore the effects of OCT in protecting intestinal mucosal barrier function. A Cell Counting Kit‑8 assay was used to determine cell viability and evaluate the effectiveness of OCT. Gene silencing technology was used to reveal the mediated effect of somatostatin receptor 2 (SSTR2). The changes in intestinal permeability were detected through trans‑epithelial electrical resistance and fluorescein isothiocyanate‑dextran 4 experiments, and the alterations in tight junction proteins were detected using immunoblotting and reverse transcription fluorescence‑quantitative PCR technology. Autophagosomes were observed by electron microscopy and the dynamic changes of the autophagy process were characterized by light chain (LC)3‑II/LC3‑I conversion and autophagic flow. The results indicated that SSTR2‑dependent OCT can prevent the decrease in cell activity. After LPS treatment, the permeability of monolayer cells decreased and intercellular tight junctions were disrupted, resulting in a decrease in tight junction protein zona occludens 1 in cells. The level of autophagy‑related protein LC3 was altered to varying degrees at different times. These abnormal changes gradually returned to normal levels after the combined application of LPS and SSTR2‑dependent OCT, confirming the role of OCT in protecting intestinal barrier function. These experimental results suggest that OCT maintains basal autophagy and cell activity mediated by SSTR2 in intestinal epithelial cells, thereby preventing the intestinal barrier dysfunction in inflammation injury.

Murine sterile fecal filtrate is a potent pharmacological agent that exerts age-independent immunomodulatory effects in RAW264.7 macrophages

BackgroundSterile fecal filtrate (SFF) is being considered a safer alternative to fecal microbiota transplantation (FMT) therapy; however, its bioactive potency is very little understood. The present study thus assessed the age-dependent immunostimulatory and immunomodulatory attributes of murine SFF in vitro.MethodsSFF from young (Y-SFF) and old (O-SFF) Swiss albino mice were prepared. Immunostimulatory and immunomodulatory effects of SFF were evaluated in resting and lipopolysaccharide (LPS) stimulated macrophage cells by measuring intracellular reactive oxygen species (ROS), nitric oxide (NO) production, inflammatory cytokines profile, as well as gene expression of oxidative and inflammatory transcription factors. SFF were also evaluated for native antioxidant capacity by measuring DPPH and ABTS free radical scavenging activity. Bioactive components present in SFF were also determined by GC/MS analysis.ResultsBoth Y-SFF and O-SFF induced potent immunostimulatory effects characterized by changes in cell morphology, a significant increase in NO production, ROS levels, and an increased ratio of pro-inflammatory (IL-6, TNF-α, IL-1β) to anti-inflammatory (IL-10) secretory proteins although no significant aggravation in the transcription of NF-κB and Nrf-2 could be observed. Application of LPS to cells significantly augmented a pro-oxidative and pro-inflammatory response which was much higher in comparison to Y-SFF or O-SFF application alone and mediated by strong suppression of Nrf-2 gene expression. Pre-treatment of macrophages with both Y-SFF and O-SFF robustly attenuated cellular hyperresponsiveness to LPS characterized by significantly decreased levels of NO, ROS, and inflammatory cytokines while a concomitant increase in anti-inflammatory protein (IL-10) was observed. Further, both Y-SFF and O-SFF strongly resisted LPS-induced downregulation of Nrf-2 expression although O-SFF appeared to protect cells slightly better from the overall LPS threat. Neat SFF samples exhibited moderate antioxidant capacity and GC/MS analysis of SFF revealed diverse volatile organic compounds characterized by alkanes, organosulphur compounds, furans, amides, amino acids, and antimicrobial elements.ConclusionOur results indicate that SFF is a potent stimulant of macrophages and confers strong anti-inflammatory effects regardless of donor age thereby suggesting its therapeutic efficacy in lieu of FMT therapy.

Antioxidant Polyphenols from Lespedeza bicolor Turcz. Honey: Anti-Inflammatory Effects on Lipopolysaccharide-Treated RAW 264.7 Macrophages.

Although the honey produced by Lespedeza bicolor Turcz. is precious because of its medicinal value, its pharmacological mechanism is still unclear. Here, its anti-inflammatory and antioxidant functions on lipopolysaccharide (LPS)-treated murine RAW 264.7 macrophages were analyzed using targeted and non-targeted metabolomics. Results showed that twelve polyphenols were identified in L. bicolor honey using UHPLC-QQQ-MS/MS. L. bicolor honey extract could scavenge the free radicals DPPH• and ABTS+ and reduce Fe3+. Furthermore, pretreatment with L. bicolor honey extract significantly decreased NO production; suppressed the expression of COX-2, IL-10, TNF-α, and iNOS; and upregulated HO-1's expression in the cells with LPS application. UHPLC-Q-TOF-MS/MS-based metabolomics results revealed that L. bicolor honey extract could protect against inflammatory damage caused by LPS through the reduced activation of sphingolipid metabolism and necroptosis pathways. These findings demonstrate that L. bicolor honey possesses excellent antioxidant and anti-inflammatory activities.

Acute phase reaction to lipopolysaccharide-induced mastitis in early lactation dairy cows fed nitrogenic, glucogenic, or lipogenic diets

The availability of certain macronutrients is likely to influence the capacity of the immune system. Therefore, we investigated the acute phase response to intramammary (i.mam.) lipopolysaccharide (LPS) in dairy cows fed a nitrogenic diet (n = 10) high in crude protein, a glucogenic diet (n = 11) high in carbohydrates and glucogenic precursors, or a lipogenic diet (n = 11) high in lipids. Thirty-two dairy cows were fed one of the dietary concentrates directly after calving until the end of trial at 27 ± 3 days in milk (mean ± standard deviation). In wk 3 of lactation, 20 µg of LPS was i.mam. injected in one quarter, and sterile NaCl (0.9%) in the contralateral quarter. Milk samples of the LPS-challenged and control quarter were taken hourly from before (0 h) until 9 h after LPS challenge and analyzed for milk amyloid A (MAA), haptoglobin (HP), and IL-8. In addition, blood samples were taken in the morning, and composite milk samples at morning and evening milkings, from 1 d before until 3 d after LPS challenge, and again on d 9, to determine serum amyloid A (SAA) and HP in blood, and MAA and HP in milk. The mRNA abundance of various immunological and metabolic factors in blood leukocytes was quantified by quantitative reverse-transcription PCR from samples taken at -18, -1, 6, 9, and 23 h relative to LPS application. The dietary concentrates did not affect any of the parameters in blood, milk, and leukocytes. The IL-8 was increased from 2 h, HP from 2 to 3 h, and MAA from 6 h relative to the LPS administration in the milk of the challenged quarter and remained elevated until 9 h. The MAA and HP were also increased at 9 h after LPS challenge in whole-udder composite milk, whereas HP and SAA in blood were increased only after 23 h. All 4 parameters were decreased again on d 9. Similar for all groups, the mRNA abundance of HP and the heat shock protein family A increased after the LPS challenge, whereas the mRNA expression of the tumor necrosis factor α and the leukocyte integrin β 2 subunit (CD18) were decreased at 6 h after LPS challenge. The glucose transporter (GLUT)1 mRNA abundance decreased after LPS, whereas that of the GLUT3 increased, and that of the GLUT4 was not detectable. The mRNA abundance of GAPDH was increased at 9 h after LPS and remained elevated. The acute phase protein response was detected earlier in milk compared with blood indicating mammary production. However, immunological responses to LPS were not affected by the availability of specific macronutrients provided by the different diets.

THE NEURONAL ANTIINFLAMMATORY EFFECTS OF APELIN-13

Objective: Apelin, a peptide with growth factor characteristic, is expressed in many cells and acts by binding to the APJ receptor. Apelin mRNA was detected in the central nervous system (CNS), macrophages and many peripheral tissues. According to recent studies, apelin protects neurons from hypoxia and glutamate-mediated excitotoxicity and can protect cells from oxidative stres (stress) (OS) damage by stimulating catalase activity. Pituitary and neurodegenerative diseases are associated with progressive neuronal loss in the CNS. Neuroinflammation and OS-induced cell damage, caused by microglia and astrocytes activation, have an important role in the pathogenesis and prognosis of these diseases. The aim of this study is to investigate the effects of apelin-13 molecule on the development of inflammation and OS caused by bacterial endotoxin in an invitro astrocyte cell culture medium. Methods: The effect of apelin -13 on cell viability changes in lipopolysaccharide (LPS) induced astrocyte inflammation was analyzed by enzymatic MTT test. The anti-inflammatory effects were determined by qRT-PCR and ELISA analysis at the level of TNF-α, IL-1β, IL-6 and mRNA transcription and protein. The effects of apelin on endotoxin mediated OS and possible DNA damage, lipid peroxidation, and nitric oxide (NO) production were evaluated by ELISA method. Results: In our study, LPS application to astrocytes caused inflammation and a significant decrease in inflammation was found with apelin treatment.
 Conclusion: The results obtained from this study reveal the antiinflammatory, antiapoptotic and antioxidant effects of apelin, that have widely expressed receptors and role in the development of immune response on the development of neuroinflammation and oxidative cell damage.

A key role for platelet GPVI in neutrophil recruitment, migration, and NETosis in the early stages of acute lung injury

AbstractAcute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are associated with high morbidity and mortality. Excessive neutrophil infiltration into the pulmonary airspace is the main cause for the acute inflammation and lung injury. Platelets have been implicated in the pathogenesis of ALI/ARDS, but the underlying mechanisms are not fully understood. Here, we show that the immunoreceptor tyrosine-based activation motif–coupled immunoglobulin-like platelet receptor, glycoprotein VI (GPVI), plays a key role in the early phase of pulmonary thrombo-inflammation in a model of lipopolysaccharide (LPS)-induced ALI in mice. In wild-type (WT) control mice, intranasal LPS application triggered severe pulmonary and blood neutrophilia, hypothermia, and increased blood lactate levels. In contrast, GPVI-deficient mice as well as anti-GPVI–treated WT mice were markedly protected from pulmonary and systemic compromises and showed no increased pulmonary bleeding. High-resolution multicolor microscopy of lung sections and intravital confocal microcopy of the ventilated lung revealed that anti-GPVI treatment resulted in less stable platelet interactions with neutrophils and overall reduced platelet–neutrophil complex (PNC) formation. Anti-GPVI treatment also reduced neutrophil crawling and adhesion on endothelial cells, resulting in reduced neutrophil transmigration and alveolar infiltrates. Remarkably, neutrophil activation was also diminished in anti-GPVI–treated animals, associated with strongly reduced formation of PNC clusters and neutrophil extracellular traps (NETs) compared with that in control mice. These results establish GPVI as a key mediator of neutrophil recruitment, PNC formation, and NET formation (ie, NETosis) in experimental ALI. Thus, GPVI inhibition might be a promising strategy to reduce the acute pulmonary inflammation that causes ALI/ARDS.

Evaluation of the protective role of resveratrol against sepsis caused by LPS via TLR4/NF-κB/TNF-α signaling pathways: Experimental study.

The development and progression of sepsis are multifactorial and influencethe immunological, endocrine, and cardiovascular systems of the body. Our knowledge of the key mechanisms involved in the pathogenesis of sepsis has expanded exponentially, yet this still needs to be translated into effective targeted therapeutic regimes. In the present study, we aimed to determine whether resveratrol has positive effects in the experimental sepsis rat model. Twenty-eightmale Spraque-Dawley rats were randomly divided into four groups (n = 7) as follows:control, lipopolysaccharide (LPS) (30 mg/kg dose), resveratrol, and LPS and resveratrol. After the experiment, liver and kidney tissues were collected for histopathological evaluation, blood serums were collected to measure malondialdehyde levels with enyzme-linked immunosorbent assay, and Toll-like receptor-4 (TLR4), tumor necrosis factor-α (TNF-α), nuclear factor-κB (NF-κB) immunoreactivity density was evaluated immunohistochemically. In addition, messenger RNA expression levels for TLR4, TNF-α, NF-κB, interleukin-1β, and interleukin 6 were measured. In addition, the damage observed in liver and kidney tissue was determined by AgNOR (argyrophilic nucleolar organizer regions) staining. LPS application caused severe tissue damage, oxidative stress, and increased the expressions of proinflammatory proteins and genes we evaluated, while resveratrol application eliminated these negativities. Resveratrol has been proven to suppress the TLR4/NF-κB/TNF-α pathway, a possible therapeutic signaling pathway that is important in initiating the inflammatory response in an animal model of sepsis.

The effect of mitochondria-targeted slow hydrogen sulfide releasing donor AP39-treatment on airway inflammation

Mitochondrial dysfunction has been shown to contribute to the pathophysiology of airway diseases. Therefore, mitochondria are targeted in the development of new therapeutic approaches. Hydrogen sulfide (H2S) has been shown to be involved in the pathophysiological processes of airway inflammation. We aimed to evaluate the effect of mitochondria-targeted slow H2S releasing donor AP39 [(10-oxo-10-(4-(3-thioxo-3H-1,2-dithiol5yl)phenoxy)decyl)triphenylphosphoniumbromide)] on lipopolysaccharide (LPS)-induced airway inflammation in mice. LPS was applied to female Balb/c mice by intranasal (i.n.) route to induce airway inflammation and the subgroups of mice were treated with i.n. AP39 (250–1000 nmol/kg). 48 h after LPS administration airway reactivity was evaluated in vivo, then bronchoalveolar lavage (BAL) fluid and lungs were collected. LPS application led to bronchial hyperreactivity and neutrophil infiltration into the lung tissues along with increased TNF-α, IL-1β and IL-6 levels in BAL fluid. LPS also induced an increase in the rate of glycolysis, glycogenolysis and Krebs-cycle. AP39 treatment prevented the LPS-induced bronchial hyperreactivity and reversed the increase in TNF-α and IL-6 levels in BAL fluid. The increase in neutrophil numbers in BAL fluid was also prevented by AP39 treatment at the highest dose. Our results indicate that AP39 can prevent bronchial hyperreactivity and decrease airway inflammation. Targeting H2S to the mitochondria may be a new therapeutic approach in airway inflammation.

Effects of Flavobacterium psychrophilum extracellular products and lipopolysaccharide antigens on the antioxidant enzyme system activity of rainbow trout (Oncorhynchus mykiss) fry

The aim of this study was to investigate the response of different doses of lipopolysaccharide (LPS) and extracellular product (ECP) antigens (25, 50, 100, 250, and 500 µg) obtained from Flavobacterium psychrophilum to changes in oxidative stress parameters (catalase [CAT], superoxide dismutase [SOD], glutathione peroxidase [GPx], glucose 6-phosphate dehydrogenase [G6PD], glutathione reductase [GR], and malondialdehyde [MDA]) in juvenile rainbow trout. For this purpose, 6 different doses of F. psychrophilum (102–106) were applied to the fish, and the lethal dose 50 % (LD50) value was determined as 1.33·104. Changes in MDA levels and antioxidant enzymes were determined by applying 25 % of the LD50 value to fish for 14 days. Accordingly, while F. psychrophilum increased GR, G6PD, GPx, and SOD activity, it decreased MDA levels and CAT activity. Initially, 25, 50, 100, 250, and 500 µg doses of ECP and LPS were applied to the fish; after 21 days, a 25 % dose of LD50 was applied to these fish. Fourteen days later, the fish were taken, and the changes in MDA levels and antioxidant enzymes were examined. As a result, it was determined that while ECP and LPS application decreased GR, G6PD, GPx, and SOD activity, there was an increase in MDA levels and CAT activity.

Dexpanthenol Inhibits Inflammation and Apoptosis in LPS-Induced Acute Lung Injury by Reducing Increased VCAM-1 and Caspase-3 Expressions in Rats

This study aims to investigate the effects of Dexpanthenol (Dex), a stable alcoholic analogue of D-pantothenic acid which has anti-oxidant, antiapoptotic, and antiinflammatory properties, on lipopolysaccharide (LPS)-induced lung damage via caspase-3 (cas-3) and vascular cell adhesion molecule-1 (VCAM-1) levels. According to the experimental plan of study, thirty-two Wistar Albino species rats were distributed randomly into four groups as control, LPS (5 mg/kg, intraperitoneally (i.p), single dose), LPS (30 minutes before last Dex treatment) + Dex (500 mg/kg, i.p, for 3 days) and Dex. After six hours of LPS application, lung tissues of the rats were taken for histopathological, immunohistochemical and biochemical examinations. According to results of the study, LPS causes hyperemia, neutrophil leukocyte chemotaxis and thickened septal tissue on lung. Inducing inflammation by increasing VCAM-1 levels and triggers apoptosis by increasing cas-3 levels in lung tissue. In addition, LPS decreased total antioxidant status levels, which is a marker of anti-oxidant capacity, and increased oxidative stress index and total oxidant status values, which are indicators of oxidative stress. Dex has shown its effect by reversing all these alterations and normalizing the values. These results suggest that Dex can be used as a preservative to reduce LPS-induced acute toxicity in the lung.

In vivo evaluation of a lipopolysaccharide-induced ear vascular leakage model in mice using photoacoustic microscopy.

Sepsis is caused by dysregulated host inflammatory response to infection. During sepsis, early identification and monitoring of vascular leakage are pivotal for improved diagnosis, treatment, and prognosis. However, there is a lack of research on noninvasive observation of inflammation-related vascular leakage. Here, we investigate the use of photoacoustic microscopy (PAM) for in vivo visualization of lipopolysaccharide (LPS)-induced ear vascular leakage in mice using Evans blue (EB) as an indicator. A model combining needle pricking on the mouse ear, topical smearing of LPS on the mouse ear, and intravenous tail injection of EB is developed. Topical application of LPS is expected to induce local vascular leakage in skin. Inflammatory response is first validated by ex vivo histology and enzyme-linked immunosorbent assay. Then, local ear vascular leakage is confirmed by ex vivo measurement of swelling, thickening, and EB leakage. Finally, PAM for in vivo identification and evaluation of early vascular leakage using the model is demonstrated. For PAM, common excitation wavelength of 532 nm is used, and an algorithm is developed to extract quantitative metrics for EB leakage. The results show potential of PAM for noninvasive longitudinal monitoring of peripheral skin vascular leakage, which holds promise for clinical sepsis diagnosis and management.

Activation of Humoral Immunity during the Pathogenesis of Experimental Chronic Lung Allograft Dysfunction.

Alloreactive and autoreactive antibodies have been associated with the development of chronic lung allograft dysfunction (CLAD), but their pathogenic role is disputed. Orthotopic left lung transplantation was performed in the Fischer-344 to Lewis rat strain combination followed by the application of ciclosporine for 10 days. Four weeks after transplantation, lipopolysaccharide (LPS) was instilled into the trachea. Lungs were harvested before (postoperative day 28) and after LPS application (postoperative days 29, 33, 40, and 90) for histopathological, immunohistochemical, and Western blot analyses. Recipient serum was collected to investigate circulating antibodies. Lung allografts were more strongly infiltrated by B cells and deposits of immunoglobulin G and M were more prominent in allografts compared to right native lungs or isografts and increased in response to LPS instillation. LPS induced the secretion of autoreactive antibodies into the circulation of allograft and isograft recipients, while alloreactive antibodies were only rarely detected. Infiltration of B cells and accumulation of immunoglobulin, which is observed in allografts treated with LPS but not isografts or native lungs, might contribute to the pathogenesis of experimental CLAD. However, the LPS-induced appearance of circulating autoreactive antibodies does not seem to be related to CLAD, because it is observed in both, isograft and allograft recipients.

Effect of topical application of lipopolysaccharide on contact hypersensitivity

Lipopolysaccharide (LPS) is the principal component of the outer membrane of gram-negative bacteria. The prior oral administration of LPS attenuates inflammatory responses, such as intestinal injury and atopic dermatitis, in mouse models; however, the underlying mechanism remains unclear. Here, we examined the effect of topical LPS application on allergic contact dermatitis and its mechanism of action using a murine contact hypersensitivity (CHS) model. Prolonged LPS application to the skin significantly suppressed 2,4-dinitrofluorobenzene (DNFB)-induced CHS. LPS application to the skin also reduced the phagocytosis of fluorescein isothiocyanate (FITC)-dextran by Langerhans and dendritic cells. Cutaneous cell migration into the skin-draining lymph nodes (LNs) induced by FITC painting was reduced by LPS application. During the CHS response, DNFB application induced T-cell proliferation and inflammatory cytokine production in skin-draining LNs, whereas prolonged LPS application inhibited DNFB-induced T-cell growth and interferon gamma production, indicating suppression of DNFB-induced sensitization. These results suggest that prolonged LPS application suppressed DNFB-induced sensitization and subsequently CHS response. Our findings imply that topical application of LPS may prevent allergic dermatitis such as CHS.

Antiinflammatory effects of adalimumab, tocilizumab, and steroid on lipopolysaccharide-induced lung injury.

Background/aimAcute lung injury (ALI) is a major cause of death in the intensive care unit. Lipopolysaccharide (LPS) induced lung injury is the most widely used experimental ALI model and provides opportunities for new targeting therapy. In this study, we investigated the effects of tocilizumab, adalimumab, and methylprednisolone in LPS-induced acute lung injury.Materials and methodsLung injury was established by intratracheal instillation of LPS. The rats were randomly divided into six groups: LPS, control, and treatment groups (adalimumab, tocilizumab, methylprednisolone, adalimumab + tocilizumab). Bronchoalveolar lavage (BAL) and lung tissues were collected at 48 h and 96 h following LPS administration from each group. For histological analysis, hematoxylin–eosin (H&E) staining was performed. The sections were obtained for immunohistochemical analysis. IL-6 and TNF-alpha immunoreactivity were measured. ResultsIntratracheal LPS application resulted in inflammatory cell infiltration of interstitial and alveolar spaces and thickening of the alveolar wall. All treatment groups showed significantly amelioration compared to LPS at 48 h. Interestingly, adalimumab and adalimumab + tocilizumab groups showed a significant amelioration of the lung histoarchitecture, compared to the prednisolone group at 96 h (p = 0.028, p = 0.025, respectively). Compared to the control group, LPS stimulation resulted in a significant increase in IL-6 and TNF-alpha immunoreactivity (p < 0.001). IL-6 and TNF-alpha expression were markedly reduced in all treatment groups at 48 h but the reduction was greater in the adalimumab and tocilizumab group than in the steroid. Administration with adalimumab and/or tocilizumab effectively decreased expression of TNF-alpha (p = 0.001) and IL-6 (p < 0.001) at 96 h, but prednisolone did not exert an effective decrease (p > 0.05). ConclusionAdalimumab and/or tocilizumab significantly reduce the release of proinflammatory cytokines and improve the tissue inflammation in the experimental model of ALI. Our results suggest that adalimumab and/or tocilizumab have a more potent antiinflammatory effect on lung injury than the steroid.

Specific parameters of infrared LED irradiation promote the inhibition of oxidative stress in dental pulp cells

ObjectivesThe present study aimed to assess the oxidative stress and the viability of dental pulp cells stimulated by lipopolysaccharide (LPS) and submitted to photobiomodulation (PBM) with infrared light-emitting diode (LED, 850 nm). DesignThree healthy primary teeth (n = 3) were collected and seeded in 24-well plates with 10 µg/mL of LPS to induce inflammatory mediator formation. The cells were irradiated (850 nm, 40 mW/cm2 and 80 mW/cm2) at the proposed radiant exposures of 0 (control), 4, 15, and 30 J/cm2 shortly after LPS supplementation. The tests were performed 24 h after irradiation to assess mitochondrial activity (MTT assay), the number of viable cells (Trypan Blue), cell morphology (Scanning Electron Microscopy - SEM), and the quantification of Nitric Oxide (NO) and Reactive Oxygen Species (ROS). The data were analyzed using Kruskal–Wallis and Dunn’s tests (p < 0.05). ResultsThe irradiated groups showed larger viable cells number than the non-irradiated group with LPS (p < 0.0001). All irradiation parameters decreased ROS concentrations after LPS application compared to the non-irradiated group (p < 0.05). All irradiation parameters enhanced the NO values compared to those of the control group (p < 0.05). The SEM images showed cells with regular morphology that adhered to the substrate. ConclusionsAccording to the parameters used in this study, the radiant exposure of 15 J/cm2 and irradiance of 40 mW/cm2 were the most effective irradiation parameters to stimulate and modulate oxidative stress in the primary teeth-derived dental pulp cells.

Ferulago pauciradiata Boiss. &amp; Heldr. Effects of Prantschimgin Compound Isolated from the Root of the Plant on LPS-Induced Inflammation Model in the L929 Cell Line

Investigation of the effects of prantchimgin (Prn) compound isolated from the root part of the Ferulago pauciradiata (Apiaceae) plant in the lipopolysaccharide (LPS) cytotoxicity model created in the L929 cell line. L929 cell lines were applied in a 5% CO2 incubator at 37 °C, using standard cell culture procedures, LPS was applied with Prn pure substance with a final concentration of 2 µL and 1 µL after 1 hour. Following the LPS application, the MTT protocol, a colorimetric method, was applied to determine cell viability at the end of the required 24th, 48th, and 72nd hours incubation times. IC50 values were calculated as 0.28 µg/mL for Pnr and 1 µg/mL for LPS. Application of LPS to L929 cell lines caused a significant decrease in cellular index depending on time. Also, in the Prn + LPS groups, it was found that the decreased cell index significantly increased even closer to the control compared to the LPS applied group. It was found that cyctoxicity and cell damage caused by LPS applied to the L929 cell line improved after Pnr application.

Prophylactic effect of myricetin and apigenin against lipopolysaccharide-induced acute liver injury.

Liver has an important role in the initiation and progression of multiple organ failure that occurs in sepsis. Many natural active substances can be used to reduce the liver injury caused by sepsis. For this aim, the effects of myricetin and apigenin on mice model of acute liver injury was evaluated in this study. Thirty-six mice were randomly divided into six groups as; control, lipopolysaccharide (LPS) (5mg/kg), LPS + myricetin (100mg/kg), LPS + myricetin (200mg/kg), LPS + apigenin (100mg/kg), and LPS + apigenin (200mg/kg) groups. Myricetin and apigenin were administered orally for 7days, and LPS was administered intraperitoneally only on the 7th day of the study. 24h after LPS application, all animals were sacrificed and serum biochemical parameters, histopathology and oxidative stress and inflammation markers of liver tissue were examined. Myricetin and apigenin pre-treatments increased serum albumin and total protein levels, liver GSH level and catalase and SOD activities and decreased serum ALT, AST, ALP, γ-GT, CRP, total and direct bilirubin levels, liver MPO activity, MDA, NOx, PGE2, TNF-α, IL-1β, and IL-6 levels, iNOS and COX-2 mRNA levels, phosphorylation of NF-κB p65, IκB, and IKK proteins but not p38, ERK, and JNK proteins in LPS-treated mice. Myricetin and apigenin administration also regained the hepatic architecture disrupted during LPS application. Myricetin and apigenin pre-treatments led to reduction of liver injury indices and oxidative stress and inflammatory events and these flavonoids has probably hepatoprotective effects in acute liver injury.

LPS decreases CFTR open probability and mucociliary transport through generation of reactive oxygen species

Lipopolysaccharide (LPS) serves as the interface between gram-negative bacteria (GNB) and the innate immune response in respiratory epithelial cells (REC). Herein, we describe a novel biological role of LPS that permits GNB to persist in the respiratory tract through inducing CFTR and mucociliary dysfunction. LPS reduced cystic fibrosis transmembrane conductance regulater (CFTR)-mediated short-circuit current in mammalian REC in Ussing chambers and nearly abrogated CFTR single channel activity (defined as forskolin-activated Cl- currents) in patch clamp studies, effects of which were blocked with toll-like receptor (TLR)-4 inhibitor. Unitary conductance and single-channel amplitude of CFTR were unaffected, but open probability and number of active channels were markedly decreased. LPS increased cytoplasmic and mitochondrial reactive oxygen species resulting in CFTR carbonylation. All effects of exposure were eliminated when reduced glutathione was added in the medium along with LPS. Functional microanatomy parameters, including mucociliary transport, in human sinonasal epithelial cells in vitro were also decreased, but restored with co-incubation with glutathione or TLR-4 inhibitor. In vivo measurements, following application of LPS in the nasal cavities showed significant decreases in transepithelial Cl- secretion as measured by nasal potential difference (NPD) – an effect that was nullified with glutathione and TLR-4 inhibitor. These data provide definitive evidence that LPS-generated reactive intermediates downregulate CFTR function in vitro and in vivo which results in cystic fibrosis-type disease. Findings have implications for therapeutic approaches intent on stimulating Cl- secretion and/or reducing oxidative stress to decrease the sequelae of GNB airway colonization and infection.

Bacterial Lipase Neutralized Toxicity of Lipopolysaccharide on Chicken Embryo Cardiac Tissue

It has been shown that near all organs, especially the cardiovascular system, are affected by bacterial lipopolysaccharide via the activation of Toll-like receptor signaling pathways. Here, we tried to find the blunting effect of bacterial lipase on lipopolysaccharide (LPS)-induced cardiac tissue toxicity in chicken embryos. 7-day fertilized chicken eggs were divided randomly into different groups as follows; Control, Normal Saline, LPS (0.1, 0.5 and 1mg/kbw), and LPS (0.1, 0.5 and 1mg/kbw) plus 5mg/ml Lipase. On day 17, the hearts were sampled. The expression of genes such as GATA4, NKX2.5, EGFR, TRIF, and NF-ƙB was monitored using real-time PCR analysis. Using western blotting, we measured NF-ƙB protein level. Total antioxidant capacity, glutathione peroxidase, and Catalase activity were also studied. Microvascular density and anterior wall thickness were monitored in histological samples using H&E staining. High dose of LPS (1mg/kbw) increased the expression of TRIF but not NF-ƙB compared to the control group (p < 0.05). We found a statistically significant reduction in groups that received LPS + Lipase compared to the control and LPS groups (p < 0.05). Western blotting revealed that the injection of Lipase could reduce LPS-induced NF-ƙB compared to the control group (p < 0.05). The expression of GATA4, NKx2.5, and EGFR was not altered in the LPS group, while the simultaneous application of LPS and Lipase significantly reduced GATA4, NKx2.5, and EGFR levels below the control (p < 0.05). We found non-significant differences in glutathione peroxidase, and Catalase activity in all groups (p > 0.05), while total antioxidant capacity was increased in groups that received LPS + Lipase. Anterior wall thickness was diminished in LPS groups and the use of both lipase and LPS returned near-to-control values (p < 0.05). Despite a slight increase in microvascular density, we found statistically non-significant differences in all groups (p > 0.05). Bacterial lipase reduces detrimental effects of LPS on chicken embryo heart induced via Toll-like receptor signaling pathway.