Anti-inflammatory and anti-hyperalgesic effects induced by an aqueous aged black garlic extract in rodent models of ulcerative colitis and colitis-associated visceral pain.

Inflammatory bowel disease (IBD) is a morbid condition characterized by relapsing-remitting inflammation of the colon, accompanied by persistent gut dysmotility and abdominal pain. Different reports demonstrated biological activities of aged black garlic (ABG), including anti-inflammatory and antioxidant effects. We aimed to investigate beneficial effects exerted by ABGE on colon inflammation by using ex vivo and in vivo experimental models. We investigated the anti-inflammatory effects of an ABG water extract (ABGE) on rat colon specimens exposed to E. coli lipopolysaccharide (LPS), a known ex vivo experimental model of ulcerative colitis. We determined gene expression of various biomarkers involved in inflammation, including interleukin (IL)-1ß, IL-6, nuclear factor-kB (NF-kB), tumor necrosis factor (TNF)-α. Moreover, we studied the acute effects of ABGE on visceral pain associated with colitis induced by 2,4-di-nitrobenzene sulfonic acid (DNBS) injection in rats. ABGE suppressed LPS-induced gene expression of IL-1ß, IL-6, NF-kB, and TNF-α. In addition, the acute administration of ABGE (0.03-1 g kg-1) dose-dependently relieved post-inflammatory visceral pain, with the higher dose (1 g kg-1) able to significantly reduce both the behavioral nociceptive response and the entity of abdominal contraction (assessed by electromyography) in response to colorectal distension after the acute administration in DNBS-treated rats. Present findings showed that ABGE could represent a potential strategy for treatment of colitis-associated inflammatory process and visceral pain. The beneficial effects induced by the extract could be related to the pattern of polyphenolic composition, with particular regard to gallic acid and catechin.

Combining the Pharmaceutical and Toxicological Properties of Selected Essential Oils with their Chemical Components by GC-MS Analysis.

In the current investigation, a comprehensive analysis was carried out on essential oils (EOs) extracted from six aromatic plant species, namely Rosmarinus officinalis, Pelargonium graveolens, Thymus vulgaris, Origanum vulgare, Laurus nobilis, and Aloysia citrodora. An exploration was conducted into the chemical composition using Gas Chromatography-Mass Spectrometry (GC/MS), antioxidant properties assessed through DPPH, ABTS, CUPRAC, FRAP, MCA, and PBD assays, ecotoxicological impacts evaluated via allelopathy and the Daphnia magna heartbeat test, as well as bio-pharmacological effects including anticancer activity and gene expression analysis. Results revealed strong antioxidant activity in all essential oils, with T. vulgaris EO (2748.00 mg TE/g) and O. vulgare EO (2609.29 mg TE/g) leading in CUPRAC assay. R. officinalis EO showed the highest α-amylase inhibition at 1.58 mmol ACAE/g, while O. vulgare EO excelled in α-glucosidase inhibition at 1.57 mmol ACAE/g. Additionally, cytotoxic effects were evaluated on human colorectal cancer (HCT116) cells. A. citrodora, O. vulgare, and R. officinalis EOs were found the most potent anticancer, as also witnessed by their higher modulatory effects on the gene expression of BAX and Bcl-2. Collectively, the present data highlight the importance to implement the knowledge and to valorize the supply chain of aromatic plants.

Neuromodulatory Effects Induced by the Association of Moringa oleifera Lam., Tribulus terrestris L., Rhodiola rosea Lam., and Undaria pinnatidifida Extracts in the Hypothalamus.

The present study investigated the role of a commercial formulation constituted by herbal extracts from Rhodiola rosea, Undaria pinnatifida, Tribulus terrestris, and Moringa oleifera. The formulation was analysed for determining the content in total phenols and flavonoids and scavenging/reducing properties. The formulation was also tested on isolated mouse hypothalamus in order to investigate effects on serotonin, dopamine, neuropeptide Y (NPY), and orexin A. The gene expression of gonadrotopin releasing hormone (GnRH) was also assayed. The formulation was able to reduce dopamine and serotonin turnover, and this could be related, albeit partially, to the capability of different phytochemicals, among which hyperoside and catechin to inhibit monoaminooxidases activity. In parallel, the formulation was effective in reducing the gene expression of NPY and orexin-A and to improve the gene expression of GnRH. In this context, the increased GnRH gene expression induced by the formulation may contribute not only to improve the resistance towards the stress related to ageing, but also to prevent the reduction of libido that could be related with a stimulation of the serotoninergic pathway. According to the in silico analysis, hyperoside could play a pivotal role in modulating the gene expression of GnRH. Regarding NPY and orexin A gene expression, no direct interactions between the formulation phytochemicals and these neuropeptides were anticipated; thus, suggesting that the pattern of gene expression observed following exposure of the hypothalamus to the formulation may be secondary to inhibitory effects of dopamine and serotonin turnover. Concluding, the present study demonstrated the efficacy of the formulation in exerting neuromodulatory effects at the hypothalamic level; thus, suggesting the potential to contrast stress and fatigue.

Chemical Characterization of Different Extracts from Artemisia annua and Their Antioxidant, Enzyme Inhibitory and Anti-Inflammatory Properties.

Artemisia annua L. (Asteraceae Family) is an important plant in Asia that has been used for treating different diseases, including fever due to malaria, wounds, tubercolisis, scabues, pain, convulsions, diabetes, and inflammation. In this study we aimed to evaluate the effects of different polarity extracts (hexane, dichloromethane, ethyl acetate, ethanol, ethanol/water (70 %) and water) from A. annua against the burden of inflammation and oxidative stress occurring in colon tissue exposed to LPS. In parallel, chemical composition, antiradical, and enzyme inhibition effects against α-amylase, α-glucosidase, tyrosinase, and cholinesterases were evaluated. The water extract contained the highest content of the total phenolic with 34.59 mg gallic acid equivalent (GAE)/g extract, while the hexane had the highest content of the total flavonoid (20.06 mg rutin equivalent (RE)/g extract). In antioxidant assays, the polar extracts (ethanol, ethanol/water and water) exhibited stronger radical scavenging and reducing power abilities when compared to non-polar extracts. The hexane extract showed the best AChE, tyrosinase and glucosidase inhibitory effects. All extracts revealed effective anti-inflammatory agents, as demonstrated by the blunting effects on COX-2 and TNFα gene expression. These effects seemed to be not related to the only phenolic content. However, it is worthy of interest to highlight how the higher potency against LPS-induced gene expression was shown by the water extract ; thus suggesting a potential phytotherapy application in the management of clinical symptoms related to inflammatory colon diseases, although future in vivo studies are needed to confirm such in vitro and ex vivo observations.

Contrast-Enhanced Mammography Versus MRI in the Evaluation of Neoadjuvant Therapy Response in Patients With Breast Cancer: A Prospective Study.

BACKGROUND. Contrast-enhanced mammography (CEM) is rapidly expanding as a credible alternative to MRI in various clinical settings. OBJECTIVE. The purpose of this study was to compare CEM and MRI for neoadjuvant therapy (NAT) response assessment in patients with breast cancer. METHODS. This prospective study included 51 patients (mean age, 46 ± 11 [SD] years) with biopsy-proven breast cancer who were candidates for NAT from May 2015 to April 2018. Patients underwent both CEM and MRI before, during, and after NAT (pre-NAT, mid-NAT, and post-NAT, respectively). Post-NAT CEM included a 6-minute delayed acquisition. One breast radiologist with experience in CEM reviewed CEM examinations; one breast radiologist with experience in MRI reviewed MRI examinations. The radiologists assessed for the presence of an enhancing lesion; if an enhancing lesion was detected, its size was measured. RECIST version 1.1 response assessment categories were derived. Pathologic complete response (pCR) was defined as absence of both invasive cancer and ductal carcinoma in situ (DCIS). RESULTS. Of 51 patients, 16 achieved pCR. CEM yielded systematically lower size measurements compared with MRI (mean difference, -0.2 mm for pre-NAT, -0.7 mm for mid-NAT, and -0.3 mm for post-NAT). All post-NAT imaging tests yielded systematically larger size measurements compared with pathology (mean difference, 0.8 mm for CEM, 1.2 mm for MRI, and 1.9 mm for delayed CEM). Of 12 patients with residual DCIS, an enhancing lesion was detected in seven on post-NAT CEM, eight on post-NAT MRI, and nine on post-NAT delayed CEM. Agreement of RECIST response categories between CEM and MRI, expressed as kappa coefficient, was 0.791 at mid-NAT and 0.871 at post-NAT. For detecting pCR by post-NAT imaging, sensitivity and specificity were 81% and 83% for CEM, 100% and 86% for MRI, and 81% and 89% for delayed CEM. Sensitivity was significantly higher for MRI than CEM (p = .001) and delayed CEM (p = .002); remaining comparisons were not significant (p > .05). CONCLUSION. After NAT for breast cancer, CEM and MRI yielded comparable assessments of lesion size (both slightly overestimated vs pathology) and RECIST categories and showed no significant difference in specificity for pCR. MRI had higher sensitivity for pCR. Delayed CEM acquisition may help detect residual DCIS. CLINICAL IMPACT. Although MRI remains the preferred test for NAT response monitoring, the findings support CEM as a useful alternative when MRI is contraindicated or not tolerated.

A grape (Vitis vinifera L.) pomace water extract modulates inflammatory and immune response in SW-480 cells and isolated mouse colon.

Grape (Vitis vinifera L.) pomace is a residue derived from the winemaking process, which contains bioactive compounds displaying noteworthy health-promoting properties. The aim of the present study was to investigate the phenolic composition and protective effects of a water extract of grape pomace (WEGP) in colorectal cancer cell line SW480 and in isolated mouse colon exposed to Escherichia coli lipopolysaccharide (LPS). The extract decreased SW-480 cell viability, as well as vascular endothelial factor A (VEGFA), hypoxia-induced factor 1α (HIF1α), and transient receptor potential M8 (TRPM8) LPS-induced gene expression. Moreover, the extract inhibited mRNA levels of nuclear factor kB (NFkB), cyclooxygenase (COX)-2, tumor necrosis factor (TNF)α, interleukin (IL)-6, IL-1ß, IL-10, inducible nitric oxide synthase (iNOS), and interferon (IFN)γ, in isolated colon. Conversely, WEGP increased the gene expression of antioxidant catalase (CAT) and superoxide dismutase (SOD), in the same model. The modulatory effects exerted by WEGP could be related, at least in part, to the phenolic composition, with particular regards to the catechin level. Docking calculations also predicted the interactions of catechin toward TRPM8 receptor, deeply involved in colon cancer; thus further suggesting the grape pomace as a valuable source of bioactive extracts and phytochemicals with protective effects in the colon.

Protective Effects of PollenAid Plus Soft Gel Capsules' Hydroalcoholic Extract in Isolated Prostates and Ovaries Exposed to Lipopolysaccharide.

Pollen extract represents an innovative approach for the management of the clinical symptoms related to prostatitis and pelvic inflammatory disease (PID). In this context, the aims of the present work were to analyze the phenolic composition of a hydroalcoholic extract of PollenAid Plus soft gel capsules, and to evaluate the extract's cytotoxic effects, in human prostate cancer PC3 cells and human ovary cancer OVCAR-3 cells. Additionally, protective effects were investigated in isolated prostate and ovary specimens exposed to lipopolysaccharide (LPS). The phytochemical investigation identified catechin, chlorogenic acid, gentisic acid, and 3-hydroxytyrosol as the prominent phenolics. The extract did not exert a relevant cytotoxic effect on PC3 and OVCAR-3 cells. However, the extract showed a dose-dependent inhibition of pro-inflammatory IL-6 and TNF-α gene expression in prostate and ovary specimens, and the extract was effective in preventing the LPS-induced upregulation of CAT and SOD gene expression, which are deeply involved in tissue antioxidant defense systems. Finally, a docking approach suggested the capability of catechin and chlorogenic acid to interact with the TRPV1 receptor, playing a master role in prostate inflammation. Overall, the present findings demonstrated anti-inflammatory and antioxidant effects of this formulation; thus, suggesting its capability in the management of the clinical symptoms related to prostatitis and PID.

New Biological and Chemical Evidences of Two Lamiaceae Species (Thymbra capitata and Thymus sipyleus subsp. rosulans): In Vitro, In Silico and Ex Vivo Approaches.

In this study, the methanolic and infusion extracts of two species, Thymbra capitata and Thymus sipyleus subsp. rosulans, were tested for their chemical composition and biological abilities (antioxidant, enzyme inhibitory and anti-inflammatory effects). The extracts yielded total phenolic and flavonoid contents in the range of 83.43-127.52 mg GAE/g and 9.41-46.34 mg RE/g, respectively. HPLC analysis revealed rosmarinic acid to be a major component of the studied extracts (15.85-26.43%). The best ABTS radical scavenging ability was observed in the methanol extract of T. capitata with 379.11 mg TE/g, followed by in the methanol extract of T. sipylus (360.93 mg TE/g). In the CUPRAC assay, the highest reducing ability was also found in the methanol extract of T. capitata with 802.22 mg TE/g. The phosphomolybdenum ability ranged from 2.39 to 3.61 mmol TE/g. In terms of tyrosinase inhibitory effects, the tested methanol extracts (83.18-89.66 mg KAE/g) were higher than the tested water extracts (18.74-19.11 mg KAE/g). Regarding the BChE inhibitory effects, the methanol extracts were active on the enzyme while the water extracts showed no inhibitory effect on it. Overall, the methanolic extracts showed better enzyme inhibition compared to the infusion extracts. Molecular docking also showed the selected exhibited potential binding affinities with all enzymes, with a preference for cholinesterases. Additionally, the extracts were effective in attenuating the LPS-induced increase in COX-2 and IL-6 gene expression in isolated colon, thus indicating promising anti-inflammatory effects. The preliminary results of this study suggest that these species are good natural sources of antioxidants and also provide some scope as enzyme inhibitors, most likely due to their bioactive contents such as phenolic acids, and thus can be exploited for different applications related to health promotion and disease prevention.

Novel Simple Conjugation Chemistries for Decoration of GMMA with Heterologous Antigens.

Outer Membrane Vesicles (OMV) constitute a promising platform for the development of efficient vaccines. OMV can be decorated with heterologous antigens (proteins or polysaccharides), becoming attractive novel carriers for the development of multicomponent vaccines. Chemical conjugation represents a tool for linking antigens, also from phylogenetically distant pathogens, to OMV. Here we develop two simple and widely applicable conjugation chemistries targeting proteins or lipopolysaccharides on the surface of Generalized Modules for Membrane Antigens (GMMA), OMV spontaneously released from Gram-negative bacteria mutated to increase vesicle yield and reduce potential reactogenicity. A Design of Experiment approach was used to identify optimal conditions for GMMA activation before conjugation, resulting in consistent processes and ensuring conjugation efficiency. Conjugates produced by both chemistries induced strong humoral response against the heterologous antigen and GMMA. Additionally, the use of the two orthogonal chemistries allowed to control the linkage of two different antigens on the same GMMA particle. This work supports the further advancement of this novel platform with great potential for the design of effective vaccines.

Phenolic Characterization and Neuroprotective Properties of Grape Pomace Extracts.

Vitis vinifera (grape) contains various compounds with acknowledged phytochemical and pharmacological properties. Among the different parts of the plant, pomace is of particular interest as a winemaking industry by-product. A characterization of the water extract from grape pomace from Montepulciano d'Abruzzo variety (Villamagna doc) was conducted, and the bioactive phenolic compounds were quantified through HPLC-DAD-MS analysis. HypoE22, a hypothalamic cell line, was challenged with an oxidative stimulus and exposed to different concentrations (1 µg/mL-1 mg/mL) of the pomace extract for 24, 48, and 72 h. In the same conditions, cells were exposed to the sole catechin, in a concentration range (5-500 ng/mL) consistent with the catechin level in the extract. Cell proliferation was investigated by MTT assay, dopamine release through HPLC-EC method, PGE2 amount by an ELISA kit, and expressions of neurotrophin brain-derived neurotrophic factor (BDNF) and of cyclooxygenase-2 (COX-2) by RT-PCR. The extract reverted the cytotoxicity exerted by the oxidative stimulus at all the experimental times in a dose-dependent manner, whereas the catechin was able to revert the oxidative stress-induced depletion of dopamine 48 h and 72 h after the stimulus. The extract and the catechin were also effective in preventing the downregulation of BDNF and the concomitant upregulation of COX-2 gene expression. In accordance, PGE2 release was augmented by the oxidative stress conditions and reverted by the administration of the water extract from grace pomace and catechin, which were equally effective. These results suggest that the neuroprotection induced by the extract could be ascribed, albeit partially, to its catechin content.

Unravelling the Phytochemical Composition and the Pharmacological Properties of an Optimized Extract from the Fruit from Prunus mahaleb L.: From Traditional Liqueur Market to the Pharmacy Shelf.

Prunus mahaleb L. fruit has long been used in the production of traditional liqueurs. The fruit also displayed scavenging and reducing activity, in vitro. The present study focused on unravelling peripheral and central protective effects, antimicrobial but also anti-COVID-19 properties exerted by the water extract of P. mahaleb. Anti-inflammatory effects were studied in isolated mouse colons exposed to lipopolysaccharide. Neuroprotection, measured as a blunting effect on hydrogen-peroxide-induced dopamine turnover, was investigated in hypothalamic HypoE22 cells. Antimicrobial effects were tested against different Gram+ and Gram- bacterial strains. Whereas anti-COVID-19 activity was studied in lung adenocarcinoma H1299 cells, where the gene expression of ACE2 and TMPRSS2 was measured after extract treatment. The bacteriostatic effects induced on Gram+ and Gram- strains, together with the inhibition of COX-2, TNFα, HIF1α, and VEGFA in the colon, suggest the potential of P. mahaleb water extract in contrasting the clinical symptoms related to ulcerative colitis. The inhibition of the hydrogen peroxide-induced DOPAC/DA ratio indicates promising neuroprotective effects. Finally, the downregulation of the gene expression of ACE2 and TMPRSS2 in H1299 cells, suggests the potential to inhibit SARS-CoV-2 virus entry in the human host. Overall, the results support the valorization of the local cultivation of P. mahaleb.

Anti-Inflammatory and Neuromodulatory Effects Induced by Tanacetum parthenium Water Extract: Results from In Silico, In Vitro and Ex Vivo Studies.

Tanacetum parthenium (feverfew) has traditionally been employed as a phytotherapeutic remedy in the treatment of migraine. In this study, a commercial T. parthenium water extract was investigated to explore its anti-inflammatory and neuromodulatory effects. Isolated mouse cortexes were exposed to a K+ 60 mM Krebs-Ringer buffer and treated with T. parthenium water extract. The prostaglandin E2 (PGE2) level, brain-derived neurotrophic factor (BDNF), interleukin-10 (IL-10), and IL-1ß gene expression were evaluated in the cortex. The effects on dopamine (DA) release and dopamine transporter (DAT) gene expression were assayed in hypothalamic HypoE22 cells. A bioinformatics analysis was conducted to further investigate the mechanism of action. The extract was effective in reducing cortex PGE2 release and IL-1ß gene expression. In the same experimental system, IL-10 and BDNF gene expressions increased, and in HypoE22 cells, the extract decreased the extracellular dopamine level and increased the DAT gene expression due to the direct interaction of parthenolide with the DAT. Overall, the present findings highlight the efficacy of T. parthenium water extract in controlling the inflammatory pathways that occur during cortical-spreading depression. Additionally, the inhibition of the hypothalamic DA release observed in this study further supports the role of dopaminergic pathways as key targets for novel pharmacological approaches in the management of migraine attacks.

Atypical Presentation of Ewing's Sarcoma with a Single Left Orbital Metastasis.

BACKGROUND: We present an uncommon case of Ewing's sarcoma in a 16-year-old boy. CASE REPORT: This case can be considered unique because of the atypical presentation, normal laboratory tests and absence of the typical symptoms such as pain, masses or swelling, fatigue or weight loss, breathing problems linked to lung metastases or pathologic fractures. The only event that brought the patient to our attention was the sudden onset of left proptosis. CONCLUSIONS: The final histopathology together with CT and PET-CT findings led to the diagnosis of a multi-metastatic Ewing's sarcoma involving the orbit, skeleton, bone marrow and lymph nodes.

Putative correlates of protection against shigellosis assessing immunomarkers across responses to S. sonnei investigational vaccine.

Shigella spp. are a leading bacterial cause of diarrhea. No widely licensed vaccines are available and there is no generally accepted correlate of protection. We tested a S. sonnei Generalized Modules for Membrane Antigen (GMMA)-based vaccine (1790GAHB) in a phase 2b, placebo-controlled, randomized, controlled human infection model study (NCT03527173) enrolling healthy United States adults aged 18-50 years. We report analyses evaluating immune responses to vaccination, with the aim to identify correlates of risk for shigellosis among assessed immunomarkers. We found that 1790GAHB elicited S. sonnei lipopolysaccharide specific α4ß7+ immunoglobulin (Ig) G and IgA secreting B cells which are likely homing to the gut, indicating the ability to induce a mucosal in addition to a systemic response, despite parenteral delivery. We were unable to establish or confirm threshold levels that predict vaccine efficacy facilitating the evaluation of vaccine candidates. However, serum anti-lipopolysaccharide IgG and bactericidal activity were identified as potential correlates of risk for shigellosis.

Comprehensive metabolite and biological profile of "Sulmona Red Garlic" ecotype's aerial bulbils.

"Sulmona Red Garlic" is a well-known Italian traditional product. Bulbs, used for culinary purposes, have been largely investigated for their medicinal properties whereas aerial bulbils are usually removed as waste material. Here, for the first time, chemical composition and biological properties of the hydroalcoholic extract from aerial bulbils were investigated. Complementary information on metabolite composition were obtained using both NMR based untargeted and HPLC-DAD targeted methodologies. The NMR analysis revealed the presence of sugars, organic acids, amino acids, organosulphur compounds (methiin, alliin, allicin and cycloalliin), and other secondary metabolites. In particular, methiin and alliin were identified for the first time in the NMR spectra of aerial bulbil garlic extracts. Polyphenol content was determined by HPLC-DAD analysis: catechin, chlorogenic acid, and gallic acid turned out to be the most abundant phenolics. Hydroalcoholic extract blocked cell proliferation of colon cancer cell line HCT116 with an IC50 of 352.07 µg/mL, while it was non-toxic to myoblast cell line C2C12. In addition, it caused seedling germination reduction of two edible and herbaceous dicotyledon species, namely Cichorium intybus and C. endivia. Moreover, the same extract reduced the gene expression of TNF-α (tumor necrosis factor), HIF1-α (hypoxia-inducible factor), VEGFA (vascular endothelial growth factor), and transient receptor potential (TRP) M8 (TRPM8) indicating the ability to contrast cancer development through the angiogenic pathway. Final, in silico experiments were also carried out supporting the biological effects of organosulphur compounds, particularly alliin, which may directly interact with TRPM8. The results here reported suggest the potential use of garlic aerial bulbils often considered a waste product as a source in phytotherapeutic remedies.

Age-dependent acquisition of IgG antibodies to Shigella serotypes-a retrospective analysis of seroprevalence in Kenyan children with implications for infant vaccination.

Background: Shigellosis mainly affects children under 5 years of age living in low- and middle-income countries, who are the target population for vaccination. There are, however, limited data available to define the appropriate timing for vaccine administration in this age group. Information on antibody responses following natural infection, proxy for exposure, could help guide vaccination strategies. Methods: We undertook a retrospective analysis of antibodies to five of the most prevalent Shigella serotypes among children aged <5 years in Kenya. Serum samples from a cross-sectional serosurvey in three Kenyan sites (Nairobi, Siaya, and Kilifi) were analyzed by standardized ELISA to measure IgG against Shigella sonnei and Shigella flexneri 1b, 2a, 3a, and 6. We identified factors associated with seropositivity to each Shigella serotype, including seropositivity to other Shigella serotypes. Results: A total of 474 samples, one for each participant, were analyzed: Nairobi (n = 169), Siaya (n = 185), and Kilifi (n = 120). The median age of the participants was 13.4 months (IQR 7.0-35.6), and the male:female ratio was 1:1. Geometric mean concentrations (GMCs) for each serotype increased with age, mostly in the second year of life. The overall seroprevalence of IgG antibodies increased with age except for S. flexneri 6 which was high across all age subgroups. In the second year of life, there was a statistically significant increase of antibody GMCs against all five serotypes (p = 0.01-0.0001) and a significant increase of seroprevalence for S. flexneri 2a (p = 0.006), S. flexneri 3a (p = 0.006), and S. sonnei (p = 0.05) compared with the second part of the first year of life. Among all possible pairwise comparisons of antibody seropositivity, there was a significant association between S. flexneri 1b and 2a (OR = 6.75, 95% CI 3-14, p < 0.001) and between S. flexneri 1b and 3a (OR = 23.85, 95% CI 11-54, p < 0.001). Conclusion: Children living in low- and middle-income settings such as Kenya are exposed to Shigella infection starting from the first year of life and acquire serotype-specific antibodies against multiple serotypes. The data from this study suggest that Shigella vaccination should be targeted to infants, ideally at 6 or at least 9 months of age, to ensure children are protected in the second year of life when exposure significantly increases.

NRF2 activation is involved in ozonated human serum upregulation of HO-1 in endothelial cells.

During the last decade, it has been shown that the activation of NRF2 and the binding to electrophile-responsive element (EpREs), stimulates the expression of a great number of genes responsible for the synthesis of phase I and phase II proteins, including antioxidants enzymes and heme oxygenase-1 (HO-1). This critical cell response occurs in cardiovascular, degenerative and chronic infective diseases aggravated by a chronic oxidative stress. In our previous reports we have shown that ozonated plasma is able to up-regulate HO-1 expression in endothelial cells. In the present work we investigated a candidate mechanism involved in this process. After treatment with increasing doses of ozonated serum (20, 40 and 80 µg/mL O(3) per mL of serum), a clear dose dependent activation of NRF2 and the subsequent induction of HO-1 and NAD(P)H quinone oxidoreductase 1(NQO1) was observed. This effect was also present when cells were treated with serum and hydrogen peroxide (H(2)O(2)) or serum and 4-hydroxynonenal (4HNE). Moreover, the treatment with ozonated serum was associated with a dose-dependent activation of extracellular-signal-regulated kinases (ERK1/2) and p38 MAP kinases (p38), not directly involved in NRF2 activation. These data, provide a new insight on the mechanism responsible for the induction of HO-1 expression by ozonated serum in the endothelium, and have a practical importance as an expedient approach to the treatment of patients with both effective orthodox drugs and ozonated autohemotherapy, targeted to the restoration of redox homeostasis.

Downregulation of NOX4 expression by roflumilast N-oxide reduces markers of fibrosis in lung fibroblasts.

The phosphodiesterase 4 inhibitor roflumilast prevents bleomycin- (BLM-) induced lung fibrosis in animal models. However, its mechanism of action remains unknown. We investigated whether roflumilast N-oxide (RNO), the active metabolite of roflumilast, can modulate in vitro the oxidative effects of BLM on human lung fibroblasts (HLF). In addition, since BLM increases the production of F2-isoprostanes that have per se fibrogenic activity, the effect of RNO on oxidative stress and fibrogenesis induced by the F2-isoprostane 8-epi-PGF2α was investigated. HLF were preincubated either with the vehicle or with RNO and exposed to either BLM or 8-epi-PGF2α. Proliferation and collagen synthesis were assessed as [(3)H]-thymidine and [(3)H]-proline incorporation. Reactive oxygen species (ROS) and F2-isoprostanes were measured. NADPH oxidase 4 (NOX4) protein and mRNA were also evaluated. BLM increased both cell proliferation and collagen synthesis and enhanced ROS and F2-isoprostane production. These effects were significantly prevented by RNO. Also, RNO significantly reduced the increase in both NOX4 mRNA and protein, induced by BLM. Finally, 8-epi-PGF2α per se stimulated HLF proliferation, collagen synthesis, and NOX4 expression and ROS generation, and RNO prevented these effects. Thus, the antifibrotic effect of RNO observed in vivo may be related to its ability to mitigate ROS generation via downregulation of NOX4.

From an in vivo to an in vitro relative potency (IVRP) assay to fully characterize a multicomponent O-antigen based vaccine against Shigella.

Outer membrane vesicles (OMV) represent an innovative platform for the design of polysaccharide based vaccines. Generalized Modules for Membrane Antigens (GMMA), OMV released from engineered Gram-negative bacteria, have been proposed for the delivery of the O-Antigen, key target for protective immunity against several pathogens including Shigella. altSonflex1-2-3 is a GMMA based vaccine, including S. sonnei and S. flexneri 1b, 2a and 3a O-Antigens, with the aim to elicit broad protection against the most prevalent Shigella serotypes, especially affecting children in low-middle income countries. Here we developed an In Vitro Relative Potency assay, based on recognition of O-Antigen by functional monoclonal antibodies selected to bind the key epitopes of the different O-Antigen active ingredients, directly applied to our Alhydrogel-formulated vaccine. Heat-stressed altSonflex1-2-3 formulations were generated and extensively characterized. The impact of detected biochemical changes in in vivo and in vitro potency assays was assessed. The overall results showed how the in vitro assay can replace the use of animals, overcoming the inherently high variability of in vivo potency studies. The entire panel of physico-chemical methods developed will contribute to detect suboptimal batches and will be valuable to perform stability studies. The work on Shigella vaccine candidate can be easily extended to other O-Antigen based vaccines.

ECHOPvir: A Mixture of Echinacea and Hop Extracts Endowed with Cytoprotective, Immunomodulatory and Antiviral Properties.

Respiratory viral infections continue to pose significant challenges, particularly for more susceptible and immunocompromised individuals. Nutraceutical strategies have been proposed as promising strategies to mitigate their impact and improve public health. In the present study, we developed a mixture of two hydroalcoholic extracts from the aerial parts of Echinacea purpurea (L.) Moench (ECP) and the cones of Humulus lupulus L. (HOP) that can be harnessed in the prevention and treatment of viral respiratory diseases. The ECP/HOP mixture (named ECHOPvir) was characterized for the antioxidant and cytoprotective properties in airway cells. Moreover, the immunomodulating properties of the mixture in murine macrophages against antioxidant and inflammatory stimuli and its antiviral efficacy against the PR8/H1N1 influenza virus were assayed. The modulation of the Nrf2 was also investigated as a mechanistic hypothesis. The ECP/HOP mixture showed a promising multitarget bioactivity profile, with combined cytoprotective, antioxidant, immunomodulating and antiviral activities, likely due to the peculiar phytocomplexes of both ECP and HOP, and often potentiated the effect of the single extracts. The Nrf2 activation seemed to trigger these cytoprotective properties and suggest a possible usefulness in counteracting the damage caused by different stressors, including viral infection. Further studies may strengthen the interest in this product and underpin its future nutraceutical applications.