Anti-inflammatory effect of Trichospira verticillata via suppression of the NLRP3 inflammasome in neutrophilic asthma.

Trichospira verticillata is an annual herb that belongs to the family Asteraceae. Trichospira verticillata extract (TVE) elicits anti-plasmodial activity; however, there has been no detailed report about its anti-inflammatory effects and molecular mechanisms. In addition, herbal plants exhibit anti-inflammatory effects by suppressing the NLRP3 inflammasome. Therefore, the primary goal of this study was to examine the effects of TVE on NLRP3 inflammasome activation by measuring interleukin-1ß (IL-1ß) secretion. We treated lipopolysaccharides (LPS)-primed J774A.1 and THP-1 cells with TVE, which attenuated NLRP3 inflammasome activation. Notably, TVE did not affect nuclear factor-kappa B (NF-κB) signalling or intracellular reactive oxygen species (ROS) production and potassium efflux, suggesting that it inactivates the NLRP3 inflammasome via other mechanisms. Moreover, TVE suppressed the formation of apoptosis-associated speck-like protein (ASC) speck and oligomerization. Immunoprecipitation data revealed that TVE reduced the binding of NLRP3 to NIMA-related kinase 7 (NEK7), resulting in reduced ASC oligomerization and speck formation. Moreover, TVE alleviated neutrophilic asthma (NA) symptoms in mice. This study demonstrates that TVE modulates the binding of NLPR3 to NEK7, thereby reporting novel insights into the mechanism by which TVE inhibits NLRP3 inflammasome. These findings suggest TVE as a potential therapeutic of NLRP3 inflammasome-mediated diseases, particularly NA.

Diet-Induced High Serum Levels of Trimethylamine-N-oxide Enhance the Cellular Inflammatory Response without Exacerbating Acute Intracerebral Hemorrhage Injury in Mice.

Trimethylamine-N-oxide (TMAO), an intestinal flora metabolite of choline, may aggravate atherosclerosis by inducing a chronic inflammatory response and thereby promoting the occurrence of cerebrovascular diseases. Knowledge about the influence of TMAO-related inflammatory response on the pathological process of acute stroke is limited. This study was designed to explore the effects of TMAO on neuroinflammation, brain injury severity, and long-term neurologic function in mice with acute intracerebral hemorrhage (ICH). We fed mice with either a regular chow diet or a chow diet supplemented with 1.2% choline pre- and post-ICH. In this study, we measured serum levels of TMAO with ultrahigh-performance liquid chromatography-tandem mass spectrometry at 24 h and 72 h post-ICH. The expression level of P38-mitogen-protein kinase (P38-MAPK), myeloid differentiation factor 88 (MyD88), high-mobility group box1 protein (HMGB1), and interleukin-1ß (IL-1ß) around hematoma was examined by western blotting at 24 h. Microglial and astrocyte activation and neutrophil infiltration were examined at 72 h. The lesion was examined on days 3 and 28. Neurologic deficits were examined for 28 days. A long-term choline diet significantly increased serum levels of TMAO compared with a regular diet at 24 h and 72 h after sham operation or ICH. Choline diet-induced high serum levels of TMAO did not enhance the expression of P38-MAPK, MyD88, HMGB1, or IL-1ß at 24 h. However, it did increase the number of activated microglia and astrocytes around the hematoma at 72 h. Contrary to our expectations, it did not aggravate acute or long-term histologic damage or neurologic deficits after ICH. In summary, choline diet-induced high serum levels of TMAO increased the cellular inflammatory response probably by activating microglia and astrocytes. However, it did not aggravate brain injury or worsen long-term neurologic deficits. Although TMAO might be a potential risk factor for cerebrovascular diseases, this exploratory study did not support that TMAO is a promising target for ICH therapy.

Development of prognostic predictive model with neutrophil-lymphocyte ratio (NLR) in patients with gastric signet ring carcinoma.

ABSTRACT: The risk factors have not been well-defined for prognosis in gastric signet ring cell carcinoma (GSRC) patients. This study is designed to prognosticate survival in GSRC patients by establishing and verifying a predictive model with neutrophil-lymphocyte ratio (NLR).A total of 147 GSRC patients from Department of Surgical Oncology, Neimenggu Baogang Hospital, Inner Mongolia Medical University were retrospectively reviewed. A predictive model was established using Cox proportional hazards. The performance of the model was evaluated by ROC curves.In present study, we found that overall survival (OS) (P < .001, Fig. 1A) and tumor recurrence rate (P = .036, Fig. 1B) in the NLR ≤ 2.8 group were significantly better than those in the NLR > 2.8 group. These results showed that NLR ≤ 2.8 was significant prognostic factor related with both OS and tumor recurrence in patients with GSRC. After adjusting for competing risk factors, NLR ≤ 2.8 (hazard ratio [HR]: 2.625, 95% confidence interval [CI]: 1.505-5.3166, P = .003), tumor size (HR: 3.024, 95% CI: 1.521-4.186, P = .005), and tumor metastasis (HR: 3.303, 95% CI: 1.25-4.525, P = .012) remained independent predictors of tumor recurrence rate and OS. Our results showed that comparing with the model without NLR (area under ROC curve: 0.798), the model with NLR (area under ROC curve: 0.826) had significant better predictive power than the model without NLR, which further confirmed the value of NLR in predicting prognosis of patients with GSRC.In conclusion, a high NLR value independently predicts poor survival in patients with GSRC after surgery. The NLR may help oncologists evaluate outcomes of patients received surgical resection and chemotherapy in order to choose alternative therapies for patients with high NLR value.

Essential metals, vitamins and antioxidant enzyme activities in COVID-19 patients and their potential associations with the disease severity.

The role of micronutrient deficiency in the pathogenesis of COVID-19 has been reviewed in the literature; however, the data are limited and conflicting. This study investigated the association between the status of essential metals, vitamins, and antioxidant enzyme activities in COVID-19 patients and disease severity. We recruited 155 patients, who were grouped into four classes based on the Adults guideline for the Management of Coronavirus Disease 2019 at King Faisal Specialist & Research Centre (KFSH&RC): asymptomatic (N = 16), mild (N = 49), moderate (N = 68), and severe (N = 22). We measured serum levels of copper (Cu), zinc (Zn), selenium (Se), vitamin D3, vitamin A, vitamin E, total antioxidant capacity, and superoxide dismutase (SOD). Among the patients, 30%, 25%, 37%, and 68% were deficient in Se (< 70.08 µg/L), Zn (< 0.693 µg/mL), vitamin A (< 0.343 µg/mL), and vitamin D3 (< 20.05 µg/L), respectively, and SOD activity was low. Among the patients, 28% had elevated Cu levels (> 1.401 µg/mL, KFSH&RC upper reference limit). Multiple regression analysis revealed an 18% decrease in Se levels in patients with severe symptoms, which increased to 30% after adjusting the model for inflammatory markers. Regardless of inflammation, Se was independently associated with COVID-19 severity. In contrast, a 50% increase in Cu levels was associated with disease severity only after adjusting for C-reactive protein, reflecting its possible inflammatory and pro-oxidant role in COVID-19 pathogenesis. We noted an imbalance in the ratio between Cu and Zn, with ~ 83% of patients having a Cu/Zn ratio > 1, which is an indicator of inflammation. Cu-to-Zn ratio increased to 45% in patients with mild symptoms and 34%-36% in patients with moderate symptoms compared to asymptomatic patients. These relationships were only obtained when one of the laboratory parameters (lymphocyte or monocyte) or inflammatory markers (neutrophil-to-lymphocyte ratio) was included in the regression model. These findings suggest that Cu/Zn might further exacerbate inflammation in COVID-19 patients and might be synergistically associated with disease severity. A 23% decrease in vitamin A was seen in patients with severe symptoms, which disappeared after adjusting for inflammatory markers. This finding may highlight the potential role of inflammation in mediating the relationship between COVID-19 severity and vitamin A levels. Despite our patients' low status of Zn, vitamin D3, and antioxidant enzyme (SOD), there is no evidence of their role in COVID-19 progression. Our findings reinforce that deficiency or excess of certain micronutrients plays a role in the pathogenesis of COVID-19. More studies are required to support our results.

Results of Complex Immunopharmacological Analysis of the Mechanisms of Action of Bioregulation Agents.

Elucidation of the pharmacodynamic mechanisms of drugs capable of potentiating the effects of non-steroidal anti-inflammatory drugs is an important task. In this in vitro study, the ability of Traumeel S to influence the innate and acquired immunity was evaluated. Traumeel S was found to reduce activities of NADPH oxidase and neutrophil extracellular traps, as well as to evoke anti-inflammatory activity of lymphocyte subpopulations.

Evaluation of Berberine as an Adjunct to TB Treatment.

Tuberculosis (TB) is the global health problem with the second highest number of deaths from a communicable disease after COVID-19. Although TB is curable, poor health infrastructure, long and grueling TB treatments have led to the spread of TB pandemic with alarmingly increasing multidrug-resistant (MDR)-TB prevalence. Alternative host modulating therapies can be employed to improve TB drug efficacies or dampen the exaggerated inflammatory responses to improve lung function. Here, we investigated the adjunct therapy of natural immune-modulatory compound berberine in C57BL/6 mouse model of pulmonary TB. Berberine treatment did not affect Mtb growth in axenic cultures; however, it showed increased bacterial killing in primary murine bone marrow-derived macrophages and human monocyte-derived macrophages. Ad libitum berberine administration was beneficial to the host in combination with rifampicin and isoniazid. Berberine adjunctive treatment resulted in decreased lung pathology with no additive or synergistic effects on bacterial burdens in mice. Lung immune cell flow cytometry analysis showed that adjunctive berberine treatment decreased neutrophil, CD11b+ dendritic cell and recruited interstitial macrophage numbers. Late onset of adjunctive berberine treatment resulted in a similar phenotype with consistently reduced numbers of neutrophils both in lungs and the spleen. Together, our results suggest that berberine can be supplemented as an immunomodulatory agent depending on the disease stage and inflammatory status of the host.

Targeting DCs for Tolerance Induction: Don't Lose Sight of the Neutrophils.

Chronic inflammatory disorders (CID), such as autoimmune diseases, are characterized by overactivation of the immune system and loss of immune tolerance. T helper 17 (Th17) cells are strongly associated with the pathogenesis of multiple CID, including psoriasis, rheumatoid arthritis, and inflammatory bowel disease. In line with the increasingly recognized contribution of innate immune cells to the modulation of dendritic cell (DC) function and DC-driven adaptive immune responses, we recently showed that neutrophils are required for DC-driven Th17 cell differentiation from human naive T cells. Consequently, recruitment of neutrophils to inflamed tissues and lymph nodes likely creates a highly inflammatory loop through the induction of Th17 cells that should be intercepted to attenuate disease progression. Tolerogenic therapy via DCs, the central orchestrators of the adaptive immune response, is a promising strategy for the treatment of CID. Tolerogenic DCs could restore immune tolerance by driving the development of regulatory T cells (Tregs) in the periphery. In this review, we discuss the effects of the tolerogenic adjuvants vitamin D3 (VD3), corticosteroids (CS), and retinoic acid (RA) on both DCs and neutrophils and their potential interplay. We briefly summarize how neutrophils shape DC-driven T-cell development in general. We propose that, for optimization of tolerogenic DC therapy for the treatment of CID, both DCs for tolerance induction and the neutrophil inflammatory loop should be targeted while preserving the potential Treg-enhancing effects of neutrophils.

Effect of the Fungal Immunomodulatory Protein FIP-fve in the Neutrophilic Asthma Animal Model.

BACKGROUND: Asthma animal models provide valuable information about the pathogenesis and the treatment of asthma. An ovalbumin (OVA)/complete Freund's adjuvant (CFA)-sensitized model was developed to induce neutrophil-dominant asthma and to investigate whether fungal immunomodulatory peptide-fve (FIP-fve) could improve asthma features in the OVA/CFA-sensitized model. METHODS: We used female BALB/c mice and sensitized them intraperitoneally with OVA/CFA on days 1, 2, and 3. On days 14, 17, 21, 24, and 27, they were challenged with intranasal OVA. The airway hyper-responsiveness (AHR) was detected by BUXCO, inflammatory cells were stained with Liu's stain, the cytokines were detected using ELISA, and the airway inflammation was analyzed with hematoxylin and eosin stain. RESULTS: According to the results, OVA/CFA sensitization could induce AHR, high levels of IgE, and inflammatory cells especially neutrophils infiltration in the lung and airway inflammation. IL-4, IL-5, IL-6, IL-8, IL-10, IL-13, IL-17, IL-25, IL-33, and transforming growth factor-ß (TGF-ß) increased in the OVA/CFA-sensitized mice. OVA/CFA-sensitized mice treated with FIP-fve not only increased IL-12 and IFN-γ but also decreased IL-4, IL-5, IL-6, IL-8, IL-13, IL-17, IL-25, IL-33, and TGF-ß in the bronchoalveolar lavage fluid. Moreover, FIP-fve significantly decreased neutrophil infiltration in the lung. CONCLUSION: The OVA/CFA model induced neutrophilic asthma successfully, and FIP-fve improved neutrophil-dominant asthma.

Anti-Inflammatory and Healing Activity of the Hydroalcoholic Fruit Extract of Solanum diploconos (Mart.) Bohs.

BACKGROUND: Solanum diploconos (Mart.) Bohs is a native Brazilian plant belonging to the Solanaceae family, popularly known as "tomatinho do mato" and poorly investigated. Herein, we presented for the first time evidence for the anti-inflammatory and wound healing activities of S. diploconos fruit hydroalcoholic extract. Material and Methods. In vitro fMLP-induced chemotaxis, LPS-induced inflammatory mediator levels (cytokines by ELISA and NO release by Griess reaction), and adhesion molecule expression (CD62L, CD49d, and CD18, by flow-cytometry) were assessed in neutrophils treated with different concentrations of the extract. Inflammation resolution was measured by the efferocytosis assay and the healing activity by in vivo and in vitro assays. The air pouch model of carrageenan-induced inflammation in Swiss mice was used to investigate the in vivo anti-inflammatory effects of the extract. Leukocyte influx (by optical microscopy) and cytokine release were quantified in the pouch exudates. Additionally, the acute and subacute toxic and genotoxic effects of the extract were evaluated. RESULTS: In vitro, the extract impaired neutrophil chemotaxis and its ability to produce and/or release cytokines (TNFα, IL-1ß, and IL-6) and NO upon LPS stimuli (p < 0.01). LPS-treated neutrophils incubated with the extract presented increased CD62L expression (p < 0.01), indicating a reduced activation. An enhanced efferocytosis of apoptotic neutrophils by macrophages was observed and accompanied by higher IL-10 and decreased TNFα secretion (p < 0.01). In vivo, similar results were noted, including reduction of neutrophil migration, protein exudation, and cytokine release (p < 0.01). Also, the extract increased fibroblast proliferation and promoted skin wound healing (p < 0.01). No signs of toxicity or genotoxicity were observed for the extract. CONCLUSION: S. diploconos fruit extract is anti-inflammatory by modulating neutrophil migration/activation as well macrophage-dependent efferocytosis and inflammatory mediator release. It also indicates its potential use as a healing agent. Finally, the absence of acute toxic and genotoxic effects reinforces its possible use as medicinal product.

Germacranolides from Carpesium divaricatum: Some New Data on Cytotoxic and Anti-Inflammatory Activity.

Carpesium divaricatum Sieb. & Zucc., a traditional medicinal plant used as an inflammation-relieving remedy, is a rich source of terpenoids. At least 40 germacrane-type sesquiterpene lactones, representatives of four different structural groups, were isolated from the plant. Cytotoxicity against cancer cells in vitro is the most frequently described biological activity of the compounds. However, little is known about the selectivity of the cytotoxic effect. The anti-inflammatory activity of the germacranolides is also poorly documented. The objective of the present study was to assess the cytotoxic activity of selected C. divaricatum germacranolides-derivatives of 4,5,8,9-tetrahydroxy-3-oxo-germacran-6,12-olide towards cancer and normal cell lines (including cells of different p53 status). Moreover, to assess the anti-inflammatory effect of the compounds, the release of four proinflammatory cytokines/chemokines (IL-1ß, IL-8, TNF-α and CCL2) by lipopolysaccharide-stimulated human neutrophils was measured by ELISA. The investigated sesquiterpene lactones demonstrated nonselective activity towards prostate cancer (Du145 and PC3) and normal prostate epithelial cells (PNT2) as well as against melanoma cells (A375 and HTB140) and keratinocytes (HaCaT). Cytotoxic activity against osteosarcoma cells was independent of their p53 status. In sub-cytotoxic concentrations (0.5-2.5 µM) the studied compounds significantly decreased cytokine/chemokine release by lipopolysaccharide-stimulated human leukocytes.

Disrupted Resolution Mechanisms Favor Altered Phagocyte Responses in COVID-19.

[Figure: see text].

Tea tree oil extract causes mitochondrial superoxide production and apoptosis as an anticancer agent, promoting tumor infiltrating neutrophils cytotoxic for breast cancer to induce tumor regression.

The antitumor activity of the tea tree oil (TTO) derived product, Melaleuca Alternifolia Concentrate (MAC) was characterized mechanistically at the molecular and cellular level. MAC was analyzed for its anticancer activity against human prostate (LNCaP) and breast (MCF-7) cancer cell lines growing in vitro. MAC (0.02-0.06% v/v) dose-dependently induced the intrinsic (mitochondrial) apoptotic pathway in both the LNCaP and MCF-7 cell lines, involving increased mitochondrial superoxide production, loss of mitochondrial membrane potential (MMP), caspase 3/7 activation, as well as the presence of TUNEL+ and cleaved-PARP+ cell populations. At concentrations of 0.01-0.04% v/v, MAC caused cell cycle arrest in the G0/1-phase, as well as autophagy. The in vivo anticancer actions of MAC were examined as a treatment in the FVB/N c-Neu murine model for spontaneously arising breast cancers. Intratumoral MAC injections (1-4% v/v) significantly suppressed tumor progression in a dose-dependent manner and was associated with greater levels of tumor infiltrating neutrophils exhibiting anticancer cytotoxic activity. Induction of breast cancer cell death by MAC via the mitochondrial apoptotic pathway was also replicated occurring in tumors treated in vivo. In conclusion, our data highlights the potential for the Melaleuca-derived MAC product inducing anticancer neutrophil influx, supporting its application as a novel therapeutic agent.

Distinct contribution of hyperbaric oxygen therapy to human neutrophil function and antibiotic efficacy against Staphylococcus aureus.

Staphylococcus aureus (SA) causes superficial and severe endovascular infections. The present in vitro study investigates the anti-SA mechanisms of hyperbaric oxygen therapy (HBOT) on direct bacterial killing, antibiotic potentiation, and polymorphonuclear leukocyte (PMN) enhancement. SA was exposed to isolated human PMNs, tobramycin, ciprofloxacin, or benzylpenicillin. HBOT was used as one 90-min session. Bacterial survival was evaluated after 4 h by quantitative bacteriology. PMN functionality as reactive oxygen species (ROS) production was measured by means of dihydrorhodamine 123 analysis. We showed that HBOT exhibits significant direct anti-SA effects. HBOT increased the anti-SA effects of PMNs by 18% after PMA stimulation (p = 0.0004) and by 15% in response to SA (p = 0.36). HBOT showed an additive effect as growth reductions of 26% to sub-MICs of tobramycin (p = 0.0057), 44% to sub-MICs of ciprofloxacin (p = 0.0001), and 26% to sub-MICs of penicillin (p = 0.038). The present in vitro study provides evidence that HBOT has differential mechanisms mediating its anti-SA effects. Our observation supports the clinical possibility for adjunctive HBOT to augment the host immune response and optimize the efficacy of antibiotic treatments.

Maternal selenium status plays a crucial role on clinical outcomes of pregnant women with COVID-19 infection.

Adequate maternal selenium level is essential for immune response and healthy pregnancy. This study aimed to shed light on the selenium status of pregnant women with COVID-19 and the effects of potential deficiency in serum selenium levels. Totally 141 pregnant women, 71 of them were COVID-19 patients, in different trimesters were included in the study. Maternal serum selenium levels, demographic and clinical parameters were determined. Serum selenium levels of pregnant women in the second (p: .0003) and third (p: .001) trimesters with COVID-19 were significantly lower than in the healthy group. Maternal selenium level was found to be negatively correlated with gestational week (p < .0001, r: -.541), D-dimer (p: .0002, r: -.363) and interleukin-6 (IL-6) level (p: .02, r: -.243). In the second trimester, serum selenium level positively correlated with white blood cell (p: .002, r: .424), neutrophil (p: .006, r: .39), lymphocyte (p: .004, r: .410) count and hemoglobin (p: .02, r: .323), hematocrit (p: .008, r: .38) status. In the third trimester, it was found that maternal selenium level positively correlated with monocyte (p: .04, r: .353) and negatively correlated with C-reactive protein level (p: .03, r: -.384). Serum selenium level was gradually decreased during the pregnancy period, however, this natural decrease was enhanced together with COVID-19 infection. The reason might be increased selenium needs depended on the immune response against infection. The decrease in maternal selenium level was found to be related to IL-6 and D-dimer levels, which indicate selenium's role in disease progression.

Ozone-Induced Oxidative Stress, Neutrophilic Airway Inflammation, and Glucocorticoid Resistance in Asthma.

Despite recent advances in using biologicals that target Th2 pathways, glucocorticoids form the mainstay of asthma treatment. Asthma morbidity and mortality remain high due to the wide variability of treatment responsiveness and complex clinical phenotypes driven by distinct underlying mechanisms. Emerging evidence suggests that inhalation of the toxic air pollutant, ozone, worsens asthma by impairing glucocorticoid responsiveness. This review discusses the role of oxidative stress in glucocorticoid resistance in asthma. The underlying mechanisms point to a central role of oxidative stress pathways. The primary data source for this review consisted of peer-reviewed publications on the impact of ozone on airway inflammation and glucocorticoid responsiveness indexed in PubMed. Our main search strategy focused on cross-referencing "asthma and glucocorticoid resistance" against "ozone, oxidative stress, alarmins, innate lymphoid, NK and γδ T cells, dendritic cells and alveolar type II epithelial cells, glucocorticoid receptor and transcription factors". Recent work was placed in the context from articles in the last 10 years and older seminal research papers and comprehensive reviews. We excluded papers that did not focus on respiratory injury in the setting of oxidative stress. The pathways discussed here have however wide clinical implications to pathologies associated with inflammation and oxidative stress and in which glucocorticoid treatment is essential.

Gut microbiota metabolism and the permeability of natural products contained in infusions from herb of European goldenrod Solidago virgaurea L.

ETHNOPHARMACOLOGICAL RELEVANCE: Solidago virgaurea L. (also known as European goldenrod) is a pharmacopoeial plant material popularly used by patients in the form of an infusion. It was traditionally used in Europe and North America for the treatment of urinary tract conditions. It is also reported as a topical agent for skin disorders. AIM OF THE STUDY: Gut microbiota metabolism plays a crucial role in the bioavailability of natural products contained in plant extracts taken orally. The aim of the current study was to establish the biotransformation of compounds contained in an infusion from goldenrod using human and piglet fecal microbiota in vitro. The permeability of unmetabolized natural products and gut microbiota metabolites was evaluated using a Caco-2 cell model. Preliminary anti-inflammatory assays of raw extract using human neutrophils were also established. MATERIAL AND METHODS: An infusion was prepared from Solidaginis virgaureae herba commercially available on the market. The characterization of the raw extract was performed by UHPLC-DAD-MS method. The infusion was incubated with human or swine fecal samples in anaerobic conditions. Metabolism products were analyzed and identified by UHPLC-DAD-MS technique. The permeability of the natural products contained in the raw infusion and after metabolism was checked by UHPLC method. The influence of raw extracts on proinflammatory functions of human neutrophils after LPS stimulation was established by flow cytometry and ELISA. RESULTS: The experiments showed that goldenrod infusion contains mainly caffeoylquinic acid derivatives, flavonoids, and some phenylpropanoids. Natural products present in the extract were transformed by human and swine microbiota to smaller molecules mainly phenylpropanoid acid derivatives. The permeability assays showed that most of the parental compound present in the infusion cannot cross the gut epithelial barrier. In contrast, metabolites were able to cross the Caco-2 monolayer. Depending on the structure, different possible mechanisms of transport were observed. The infusion did not significantly influence the proinflammatory functions of human neutrophils. CONCLUSIONS: Following oral administration of goldenrod infusion, phytochemicals are prone to undergoing metabolism by gut microbiota to smaller phenylpropionic acid derivatives that can be bioavailable after crossing the gut epithelial barrier to be further metabolized and distributed. Detected metabolites should be considered as potentially active compounds responsible for the bioactivity of the raw plant material in vivo.

Evaluation of In Vitro Immunomodulatory Activity of Withania somnifera Roots on Human Neutrophils.

The present study encompasses in vitro immunomodulatory activity of Withania somnifera WS (Solanaceae) examined on human neutrophils. The isolated compound of Withania somnifera roots was screened for its possible immunomodulatory activity by nitro blue tetrazolium test, phagocytosis of killed Candida albicans, neutrophil locomotion and chemotaxis. The extracted compound was examined at concentrations, viz. 10 µg/ml, 20 µg/ml, 40 µg/ml, 100 µg/ml and 1000 µg/ml. The activity expressed by the compound on human neutrophils was found to be significant for examined parameters.

High molecular pyrogens present in plant extracts interfere with examinations of their immunomodulatory properties in vitro.

The widely accepted strategy to justify the use of medicinal plant extracts in diseases with inflammatory background is their examination on in vitro models using immune cells. It is also a key initial step of research for active principles, which could be then isolated and tested on more advanced models, becoming new pharmacologically active lead molecules. The crucial aspect which has not been so far addressed in this context, is the presence of pyrogens in plant preparations. The aim of this study was the examination of pyrogens interference with in vitro evaluation of anti-inflammatory activity of plant extracts using human primary neutrophils model together with introduction of effective method of interfering factors elimination. The obtained results showed that chosen plant extracts contained pyrogens, which were responsible for concentration-dependent stimulation of pro-inflammatory cytokines production by human neutrophils in vitro in the same extent as LPS did. The ultrafiltration method was successfully applied for pyrogens elimination, which effectiveness was confirmed using LAL test. The determined interference of pyrogens implies the necessity of their consideration and removal when in vitro studies include direct addition of plant extracts to the cell culture, what can be obtained by ultrafiltration, which does not affect extract composition.

The anti-tussive, anti-inflammatory effects and sub-chronic toxicological evaluation of perilla seed oil.

BACKGROUND: Perilla seed oil (PSO) is the main constituent of perilla seeds currently being used in the food industry, however it also has great clinical potential in the regulation of lung function as a nutrition supplement because of the high content of α-linolenic acid (ALA). In this study, the pharmacological activities including anti-tussive, expectorant and anti-inflammatory effect of PSO were performed. Furthermore, the 90-day sub-chronic oral toxicity with a 30 day recovery period was evaluated in Wistar rats. RESULTS: The pharmacological studies demonstrated that PSO inhibited cough frequency induced by capsaicine in mice. PSO also inhibited the leukotriene B4 (LTB4) release from the calcium ionophore A23187-induced polymorphonuclear neutrophils (PMNs) to some extent. In this sub-chronic toxicity study, mortality, clinical signs, body weight, food consumption, hematology, serum biochemistry, urinalysis, organ weight, necropsy, and histopathology were used to evaluate the toxicity of PSO. Lower body weight and various negative impacts on liver related parameters without histopathological lesion were observed in the 16 g kg-1 groups. No clinically significant changes were discovered in the 4 g kg-1 group during the test period. CONCLUSION: In summary, PSO exhibited anti-tussive and anti-inflammatory activities in vivo and in vitro. These sub-chronic toxicity studies inferred that the 'no-observed adverse effect level' (NOAEL) of PSO in Wistar rats was determined to be 4 g kg-1 . These results may provide a safety profile and a valuable reference for the use of PSO. © 2020 Society of Chemical Industry.

Treatment strategy of adding transcatheter arterial chemoembolization to sorafenib for advanced stage hepatocellular carcinoma.

BACKGROUND: Therapeutic effect and immunosuppressor cell alteration in adding transcatheter arterial chemoembolization (TACE) to sorafenib for advanced stage hepatocellular carcinoma (HCC) remain unclear. AIMS: To examine the therapeutic effect and immunosuppressor cell alteration in adding TACE to sorafenib. METHODS: Forty-four advanced stage HCC patients were divided into group A (n = 17) treated by sorafenib (400-600 mg/day) alone and group B patients (n = 27) treated by sorafenib and TACE. The frequency of regulatory T-cells and myeloid-derived suppressor cells (MDSC), and patients' outcomes were examined. Advanced HCC patients' survival was improved by adding TACE to sorafenib if N/L was reduced from ≥2.5 to <2.5 by TACE. RESULTS: The median (interquartile) follow-up for all patients was 8.5 (3.5 to 15.5) with a range from 1 to 71 months. The median (interquartile) survival was 5.0 (2.3-11.3) months for group A and 11.0 (5.0-19.0) months for group B patients (P = .024). In group A, the patients (n = 8) with neutrophil-to-lymphocytes ratio (N/L) < 2.5 had better survival than the patients (n = 9) with N/L ≥ 2.5 (P = .006). In group B, 6 of 13 patients with N/L ≥ 2.5 had N/L reduction to <2.5 after combination therapy of sorafenib and TACE, and their 6-month, 1-year and 2-year survival were improved (P = .013). For immune cell examination, the frequency of CD4+ and CD8+ T-lymphocytes, regulatory T-cell and MDSC were not altered by sorafenib treatment. However, actual number of lymphocytes had a tendency to increase (from 978.5 ± 319.4/mm3 prior to treatment to 1378.0 ± 403.3/mm3 , P = .086) for the patients with N/L reduction. CONCLUSION: Immunosuppressor cells were not altered by sorafeinb. Patients' survival was improved if N/L ≥ 2.5 was reduced to <2.5 by TACE.