Ultra-high dose rate radiation has been reported to produce a more favorable toxicity and tumor control profile compared to conventional dose rates that are used for patient treatment. So far, the so-called FLASH effect has been validated for electron, photon and scattered proton beam, but not yet for proton pencil beam scanning (PBS). Because PBS is the state-of-the-art delivery modality for proton therapy and constitutes a wide and growing installation base, we determined the benefit of FLASH PBS on skin and soft tissue toxicity. Using a pencil beam scanning nozzle and the plateau region of a 250 MeV proton beam, a uniform physical dose of 35 Gy (toxicity study) or 15 Gy (tumor control study) was delivered to the right hind leg of mice at various dose rates: Sham, Conventional (Conv, 1 Gy/s), Flash60 (57 Gy/s) and Flash115 (115 Gy/s). Acute radiation effects were quantified by measurements of plasma and skin levels of TGF-β1 and skin toxicity scoring. Delayed irradiation response was defined by hind leg contracture as a surrogate of irradiation-induced skin and soft tissue toxicity and by plasma levels of 13 different cytokines (CXCL1, CXCL10, Eotaxin, IL1-beta, IL-6, MCP-1, Mip1alpha, TNF-alpha, TNF-beta, VEGF, G-CSF, GM-CSF and TGF- β1). Plasma and skin levels of TGF-β1, skin toxicity and leg contracture were all significantly decreased in FLASH compared to Conv groups of mice. FLASH and Conv PBS had similar efficacy with regards to growth control of MOC1 and MOC2 head and neck cancer cells transplanted into syngeneic, immunocompetent mice. These results demonstrate consistent delivery of FLASH PBS radiation from 1 to 115 Gy/s in a clinical gantry. Radiation response following delivery of 35 Gy indicates potential benefits of FLASH versus conventional PBS that are related to skin and soft tissue toxicity.

Keywords: FLASH; normal tissue toxicity; proton beam scanning; proton therapy; skin and soft tissue; ultra-high dose rate.

Background: On June 30, 2020, the WHO reported over 10 millions of COVID-19 cases worldwide with over half a million deaths. In severe cases the disease progresses into an Acute Respiratory Distress Syndrome (ARDS), which in turn depends on an overproduction of cytokines (IL-6, TNFα, IL-12, IL-8, CCL-2 and IL1) that causes alveolar and vascular lung damage. Clearly, it is essential to find an immunological treatment that controls the "cytokine storm". In the meantime, however, it is essential to have effective antiviral and anti-inflammatory drugs available immediately.

Pharmacologic therapy for covid-19: Hydroxychloroquine or chloroquine have been widely adopted worldwide for the treatment of SARS-CoV-2 pneumonia. However, the choice of this treatment was based on low quality of evidence, i.e. retrospective, non-randomized controlled studies. Recently, four large Randomized Controlled Trials (RCTs) have been performed in record time delivering reliable data: (1) the National Institutes of Health (NIH) RCT included 60 hospitals participating all over the world and showed the efficacy of remdesivir in reducing the recovery time in hospitalized adults with COVID-19 pneumonia; (2) three large RCTs already completed, for hydroxychloroquine, dexamethasone and Lopinavir and Ritonavir respectively. These trials were done under the umbrella of the 'Recovery' project, headed by the University of Oxford. The project includes 176 participating hospitals in the UK and was set up to verify the efficacy of some of the treatments used for COVID-19. These three 'Recovery' RCTs concluded definitely: (a) that treatment with hydroxychloroquine provides no benefits in patients hospitalized with COVID-19; (b) that treatment with dexamethasone reduced deaths by one-third in COVID-19 patients that were mechanically ventilated, and by one-fifth in patients receiving oxygen only; (c) that the combination of Lopinavir and Ritonavir is not effective in reducing mortality in COVID-19 hospitalized patients.

Conclusions: The results of these four large RCTs have provided sound indications to doctors for the treatment of patients with COVID-19 and prompted the correction of many institutional provisions and guidelines on COVID-19 treatments (i.e. FDA, NIH, UK Health Service, etc.). Even though a definitive treatment for COVID-19 has not yet been found, large RCTs stand as the Gold Standards for COVID-19 therapy and offer a solid scientific base on which to base treatment decisions.

Keywords: ARDS; COVID-19; Cytokine storm syndrome; Dexamethasone; Evidence based medicine; Hydroxychloroquine; Lopinavir—ritonavir; Remdesivir; SARS-CoV-2.

OBJECTIVE

Spinal cord injury (SCI) is a devastating condition of the central nervous system. There is no proven therapeutic agent for the treatment of this complex disorder. Asiatic acid (AA) has been used as an anti-inflammatory and anti-oxidant agent in Eastern countries for many years. The aim of this study was to investigate the effectiveness of AA on the treatment of traumatic SCI in rats.

METHODS

Thirty-two adult male Sprague-Dawley rats were divided into 4 groups as laminectomy, laminectomy+trauma, vehicle, and AA treatment groups. SCI was created by the modified Allen"s weight-drop technique. After the injury, the levels of pro-inflammatory cytokines (IL-6, IL1-Β, TNF-α) and lipid peroxidation products (MDA) were measured. Tarlov functional recovery scores were also determined for each rat. The One-way ANOVA test was used for the analysis of difference between 4 experimental groups and the groups were compared individually by Tukey-LSD post hoc analysis test (p=0.001).

RESULTS

AA administration just after SCI attenuated the levels of lipid peroxidation products (MDA) and pro-inflammatory cytokines (TNF-α, IL1Β). It also increased the Tarlov functional recovery scores of the rats.

CONCLUSIONS

AA administration could attenuate a number of deleterious reactions after traumatic SCI. Further studies are needed to elucidate the pathways of neuroprotective effects of AA after spinal trauma.

Nootkatone (NTK) is a sesquiterpenoid found in essential oils of many species of Citrus (Rutaceae). Considering previous reports demonstrating that NTK inhibited inflammatory signaling pathways, this study aimed to investigate the effects of this compound in mice models of acute and chronic inflammation. Murine models of paw edema induced by carrageenan, dextran, histamine, and arachidonic acid, as well as carrageenan-induced peritonitis and pleurisy, were used to evaluate the effects of NTK on acute inflammation. A murine model of granuloma induced by cotton pellets was used to access the impact of NTK treatment on chronic inflammation. In the acute inflammation models, NTK demonstrated antiedematogenic effects and inhibited leukocyte recruitment, which was associated with decreased vascular permeability, inhibition of myeloperoxidase (MPO), interleukin (IL)1-β, and tumor necrosis factor (TNF)-α production. In silico analysis suggest that NTZ anti-inflammatory effects may also occur due to inhibition of cyclooxygenase (COX)-2 activity and antagonism of the histamine receptor type 1 (H₁). These mechanisms might have contributed to the reduction of granuloma weight and protein concentration in the homogenates, observed in the chronic inflammation model. In conclusion, NTK exerted anti-inflammatory effects that are associated with inhibition of IL1-β and TNF-α production, possibly due to inhibition of COX-2 activity and antagonism of the H1 receptor. However, further studies are required to characterize the effects of this compound on chronic inflammation.

Traumatic brain injury is a leading cause of death and disability around the world. So far, drugs are not available to repair brain damage. Human mesenchymal stem cell (hMSC) transplantation therapy is a promising approach, although the inflammatory microenvironment of the injured brain affects the efficacy of transplanted hMSCs. We hypothesize that reducing the inflammation in the cerebral microenvironment by reducing pro-inflammatory chemokines prior to hMSC administration will improve the efficacy of hMSC therapy. In a rat model of lateral fluid percussion injury, combined pioglitazone (PG) and hMSC (combination) treatment showed less anxiety-like behavior and improved sensorimotor responses to a noxious cold stimulus. Significant reduction in brain lesion volume, neurodegeneration, microgliosis and astrogliosis were observed after combination treatment. TBI induced expression of inflammatory chemokine CCL20 and IL1-β were significantly decreased in the combination treatment group. Combination treatment significantly increased brain-derived neurotrophic factor (BDNF) level and subventricular zone (SVZ) neurogenesis. Taken together, reducing proinflammatory cytokine expression in the cerebral tissues after TBI by PG administration and prior to hMSC therapy improves the outcome of the therapy in which BDNF could have a role.

Management of rheumatoid arthritis (RA) has radically changed over the last decade. Diagnostic methods have improved with availability of highly specific tests such as antibody to cyclic citrullinated peptide (specificity approximately 96%), and introduction of advanced imaging modalities such as ultrasound and magnetic resonance imaging to facilitate earlier diagnosis. The current aim of management is to achieve remission and prevent joint damage. In order to achieve this goal, inflammation is suppressed as much as possible during the early phase of the disease before onset of joint damage. Aggressive treatments with combinations of disease modifying anti-rheumatic drugs are commenced earlier in the course of disease, and tight control maintained with regular objective monitoring of disease activity. Early use of anti-TNFalpha (tumour necrosis factor alpha) therapy in combination with methotrexate helps to achieve better clinical and radiographic outcomes, which can be maintained for up to 5 years after withdrawal of anti-TNFalpha therapy. Apart from anti-TNFalpha, several other biological treatments are now available, including those that target CD20 on B cells (rituximab), cytokines such as IL1 (anakinra) and IL6 (tocilizumab), and molecules that cause interaction between antigen presenting cells and T cells (abatacept). There is better awareness and understanding of RA associated complications such as cardiovascular disease and osteoporosis. Use of non-steroidal anti-inflammatory drugs is on the decline in light of recent concerns about cardiovascular safety. Evidence is emerging in support of statins and bisphosphonates for improving RA disease activity and preventing erosions, respectively. In the coming years, further improvements in therapeutic strategies are likely with the pace at which research is currently progressing.

OBJECTIVE

To evaluate the effect of orally administered plecanatide on colorectal dysplasia in Apc+/Min-FCCC mice with dextran sodium sulfate (DSS)-induced inflammation.

METHODS

Inflammation driven colorectal carcinogenesis was induced in Apc+/Min-FCCC mice by administering DSS in their drinking water. Mice were fed a diet supplemented with plecanatide (0-20 ppm) and its effect on the multiplicity of histopathologically confirmed polypoid, flat and indeterminate dysplasia was evaluated. Plecanatide-mediated activation of guanylate cyclase-C (GC-C) signaling was assessed in colon tissues by measuring cyclic guanosine monophosphate (cGMP) by ELISA, protein kinase G-II and vasodilator stimulated phosphoprotein by immunoblotting. Ki-67, c-myc and cyclin D1 were used as markers of proliferation. Cellular levels and localization of β-catenin in colon tissues were assessed by immunoblotting and immunohistochemistry, respectively. Uroguanylin (UG) and GC-C transcript levels were measured by quantitative reverse transcription polymerase chain reaction (RT-PCR). A mouse cytokine array panel was used to detect cytokines in the supernatant of colon explant cultures.

RESULTS

Oral treatment of Apc+/MinFCCC mice with plecanatide produced a statistically significant reduction in the formation of inflammation-driven polypoid, flat and indeterminate dysplasias. This anti-carcinogenic activity of plecanatide was accompanied by activation of cGMP/GC-C signaling mediated inhibition of Wnt/β-catenin signaling and reduced proliferation. Plecanatide also decreased secretion of pro-inflammatory cytokines (IL-6, IL1 TNF), chemokines (MIP-1, IP-10) and growth factors (GCSF and GMCSF) from colon explants derived from mice with acute DSS-induced inflammation. The effect of plecanatide-mediated inhibition of inflammation/dysplasia on endogenous expression of UG and GC-C transcripts was measured in intestinal tissues. Although GC-C expression was not altered appreciably, a statistically significant increase in the level of UG transcripts was detected in the proximal small intestine and colon, potentially due to a reduction in intestinal inflammation and/or neoplasia. Taken together, these results suggest that reductions in endogenous UG, accompanied by dysregulation in GC-C signaling, may be an early event in inflammation-promoted colorectal neoplasia; an event that can potentially be ameliorated by prophylactic intervention with plecanatide.

CONCLUSIONS

This study provides the first evidence that orally administered plecanatide reduces the multiplicity of inflammation-driven colonic dysplasia in mice, demonstrating the utility for developing GC-C agonists as chemopreventive agents.

The Food and Drug Administration recently warned of the fatal cardiovascular risks of azithromycin in humans. In addition, a recently published study documented azithromycin-induced cardiotoxicity in rats. This study aimed to justify the exact cardiovascular events accompanying azithromycin administration in rats, focusing on electrocardiographic, biochemical, and histopathological changes. In addition, the underlying mechanisms were studied regarding reactive oxygen species production, cytokine release, and apoptotic cell-death. Finally, the supposed protective effects of both carvedilol and vitamin C were assessed. Four groups of rats were used: (1) control, (2) azithromycin, (3) azithromycin + carvedilol, and (4) azithromycin + vitamin C. Azithromycin resulted in marked atrophy of cardiac muscle fibers and electrocardiographic segment alteration. It increased the heart rate, lactate dehydrogenase, creatine phosphokinase, malondialdehyde, nitric oxide, interleukin-1 beta (IL1-β), tumor necrosis factor alpha (TNF-α), nuclear factor kappa beta (NF-κB), and caspase-3. It decreased reduced glutathione, glutathione peroxidase, and superoxide dismutase. Carvedilol and vitamin C prevented most of the azithromycin-induced electrocardiographic and histopathological changes. Carvedilol and vitamin C decreased lactate dehydrogenase, malondialdehyde, IL1-β, TNF-α, NF-κB, and caspase-3. Both agents increased glutathione peroxidase. This study shows that both carvedilol and vitamin C protect against azithromycin-induced cardiotoxicity through antioxidant, immunomodulatory, and antiapoptotic mechanisms.

In this study, we observed that mice lacking the IL-1 receptor (IL-1R) (IL1r-/-) or deficient in IL1-β developed multiple epidermal cysts after chronic UVB exposure. Cysts that developed in IL1r-/- mice were characterized by the presence of the hair follicle marker Sox 9, keratins 10 and 14, and normal melanocyte distribution and retinoid X receptor-α expression. The increased incidence of cysts in IL1r-/- mice was associated with less skin inflammation as characterized by decreased recruitment of macrophages, and their skin also maintained epidermal barrier function compared with wild-type mice. Transcriptional analysis of the skin of IL1r-/- mice after UVB exposure showed decreased gene expression of proinflammatory cytokines such as tumor necrosis factor-α and IL-6. In vitro, primary keratinocytes derived from IL1r-/- mice were more resistant to UVB-triggered cell death compared with wild-type cells, and tumor necrosis factor-α release was completely blocked in the absence of IL-1R. These observations illustrate an unexpected yet prominent phenotype associated with the lack of IL-1R signaling in mice and support further investigation into the role of IL-1 ligands in epidermal repair and innate immune response after damaging UVB exposure.

Interleukin 4 (IL4) is a cytokine produced by T cells and mast cells/basophils. IL4 has a key role in IgE production and in the pathogenesis of atopic diseases. Disorders such as rheumatoid arthritis are characterized by increased production of proinflammatory cytokines and reduced production of IL4. Furthermore, rheumatoid T cells are of the TH 1 type, not producing IL4. In contrast, circulating IL4 has been detected in scleroderma patients, whose T cells are of the TH 2 type, producing IL4. Since IL4 inhibits the production of proinflammatory cytokines such as IL1, IL6, TNF alpha and IL8, we hypothesized that a deficit in IL4 production might be related to the pathogenesis of rheumatoid arthritis. Addition of IL4 strongly reduced the production of proinflammatory cytokines and expression of corresponding mRNA by synovial membrane pieces from RA patients. RA synovitis is also associated with marked proliferation of synoviocytes. Studies of synoviocytes showed that IL4 inhibited the growth promoting effect of PDGF and IL1 beta. IL4 also inhibited production of IL6 by juxta-articular bone pieces. IL4 was found to reduce disease activity and progression in various arthritis models. These anti-inflammatory and anti-proliferative properties suggest that IL4 may be of potential clinical value in RA. Conversely, as IL4 induces dermal fibroblasts to secrete collagen, IL4 might be involved in the pathogenesis of scleroderma.

(<em>b</em>)Background:</<em>b</em>) The auto-inflammation and phospholipase Cγ2 (PLCγ2)-associated anti<em>b</em>ody deficiency and immune dysregulation (APLAID) syndrome is a rare primary immunodeficiency caused <em>b</em>y a gain-of-function mutation S707Y in the <i>PLCG2</i> gene previously descri<em>b</em>ed in two patients from one family. The APLAID patients presented with early-onset <em>b</em>listering skin lesions, posterior uveitis, inflammatory <em>b</em>owel disease (IBD) and recurrent sinopulmonary infections caused <em>b</em>y a humoral defect, <em>b</em>ut lacked circulating autoanti<em>b</em>odies and had no cold-induced urticaria, contrary to the patients with the related PLAID syndrome. (<em>b</em>)Case:</<em>b</em>) We descri<em>b</em>e a new APLAID patient who presented with vesiculopustular rash in the 1st weeks of life, followed <em>b</em>y IBD, posterior uveitis, recurrent chest infections, interstitial pneumonitis, and also had sensorineural deafness and cutis laxa. Her disease has <em>b</em>een refractory to most treatments, including <em>IL1</em> <em>b</em>lockers and a trial with ruxolitini<em>b</em> has <em>b</em>een attempted. (<em>b</em>)Results:</<em>b</em>) In this patient, we found a unique <i>de novo</i> heterozygous missense L848P mutation in the <i>PLCG2</i> gene, predicted to affect the PLCγ2 structure. Similarly to S707Y, the L848P mutation led to the increased <em>b</em>asal and EGF-stimulated PLCγ2 activity <i>in vitro</i>. Whole <em>b</em>lood assays showed reduced production of IFN-γ and IL-17 in response to polyclonal T-cell stimulation and reduced production of IL-10 and IL-1β after LPS stimulation. Reduced IL-1β levels and the lack of clinical response to treatment with IL-1 <em>b</em>lockers argue against NLRP3 inflammasome hyperactivation <em>b</em>eing the main mechanism mediating the APLAID pathogenesis. (<em>b</em>)Conclusion:</<em>b</em>) Our findings indicate that L848P is novel a gain-of-function mutation that leads to PLCγ2 activation and suggest cutis laxa as a possi<em>b</em>le clinical manifestations of the APLAID syndrome.

Sepsis-associated encephalopathy causes brain dysfunction that can result in cognitive impairments in sepsis survivor patients. In previous work, we showed that simvastatin attenuated oxidative stress in brain structures related to memory in septic rats. However, there is still a need to evaluate the long-term impact of simvastatin administration on brain neurodegenerative processes and cognitive damage in sepsis survivors. Here, we investigated the possible neuroprotective role of simvastatin in neuroinflammation, and neurodegeneration conditions of brain structures related to memory in rats at 10 days after sepsis survival. Male Wistar rats (250-300 g) were submitted to cecal ligation and puncture (CLP, n = 42) or remained as non-manipulated (naïve, n = 30). Both groups were treated (before and after the surgery) by gavage with simvastatin (20 mg/kg) or an equivalent volume of saline and observed for 10 days. Simvastatin-treated rats that survived to sepsis showed a reduction in the levels of nitrate, IL1-β, and IL-6 and an increase in Bcl-2 protein expression in the prefrontal cortex and hippocampus, and synaptophysin only in the hippocampus. Immunofluorescence revealed a reduction of glial activation, neurodegeneration, apoptosis, and amyloid aggregates confirmed by quantification of GFAP, Iba-1, phospho Ser₃₉₆-tau, total tau, cleaved caspase-3, and thioflavin-S in the prefrontal cortex and hippocampus. In addition, treated animals presented better performance in tasks involving habituation memory, discriminative, and aversive memory. These results suggest that statins exert a neuroprotective role by upregulation of the Bcl-2 and gliosis reduction, which may prevent the cognitive deficit observed in sepsis survivor animals.

Keywords: Astrocytes; Encephalopathy; Hippocampus; Microglia; Neurodegeneration; Prefrontal cortex.

Non-tuberculous mycobacteria (NTM), also known as an environmental and atypical mycobacteria, can cause the chronic pulmonary infectious diseases. Macrophages have been suggested as the main host cell to initiate the innate immune responses to NTM infection. However, the molecular mechanism to regulate the antimicrobial immune responses to NTM is still largely unknown. Current study showed that the NTM clinical groups, Mycobacterium abscessus and Mycobacterium smegmatis, significantly induced the M1 macrophage polarization with the characteristic production of nitric oxide (NO) and marker gene expression of iNOS, IFNγ, TNF-α, IL1-β and IL-6. Interestingly, a non-histone nuclear protein, HMGN2 (high-mobility group N2), was found to be spontaneously induced during NTM-activated M1 macrophage polarization. Functional studies revealed that HMGN2 deficiency in NTM-infected macrophage promotes the expression of M1 markers and the production of NO via the enhanced activation of NF-κB and MAPK signalling. Further studies exhibited that HMGN2 knock-down also enhanced IFNγ-induced M1 macrophage polarization. Finally, we observed that silencing HMGN2 affected the survival of NTM in macrophage, which might largely relevant to enhanced macrophage polarization into M1 phenotype under the NTM infection. Collectively, current studies thus suggested a novel function of HMGN2 in regulating the anti-non-tuberculous mycobacteria innate immunity of macrophage.

The purpose of this study was to inhibit endotoxin induced cytokines production and liver injury by liver non-parenchymal cell (NPC) selective delivery of nuclear factor kappaB (NFkappaB) decoy using mannosylated cationic liposomes (Man-liposomes). In this study, we examined the distribution, inhibitory effect on cytokines production and ALT/AST of intravenously injected Man-liposome/NFkappaB decoy complex. Man-liposome/[(32)P] NFkappaB decoy complexes mostly accumulated in the liver, preferentially in NPC. In a murine lipopolysaccharide-induced liver failure model, the production of tumor necrosis factor-alpha (TNFalpha), IFNgamma, IL1-beta, ALT and AST were effectively reduced by Man-liposome complexes. However, cationic or galactosylated cationic liposome complexes could not inhibit TNFalpha production.

The role of host genetic variation in the development of complicated Staphylococcus aureus bacteremia (SAB) is poorly understood. We used whole exome sequencing (WES) to examine the cumulative effect of coding variants in each gene on risk of complicated SAB in a discovery sample of 168 SAB cases (84 complicated and 84 uncomplicated, frequency matched by age, sex, and bacterial clonal complex [CC]), and then evaluated the most significantly associated genes in a replication sample of 240 SAB cases (122 complicated and 118 uncomplicated, frequency matched for age, sex, and CC) using targeted sequence capture. In the discovery sample, gene-based analysis using the SKAT-O program identified 334 genes associated with complicated SAB at p<3.5 x 10-3. These, along with eight biologically relevant candidate genes were examined in the replication sample. Gene-based analysis of the 342 genes in the replication sample using SKAT-O identified one gene, GLS2, significantly associated with complicated SAB (p = 1.2 x 10-4) after Bonferroni correction. In Firth-bias corrected logistic regression analysis of individual variants, the strongest association across all 10,931 variants in the replication sample was with rs2657878 in GLS2 (p = 5 x 10-4). This variant is strongly correlated with a missense variant (rs2657879, p = 4.4 x 10-3) in which the minor allele (associated here with complicated SAB) has been previously associated with lower plasma concentration of glutamine. In a microarray-based gene-expression analysis, individuals with SAB exhibited significantly lower expression levels of GLS2 than healthy controls. Similarly, Gls2 expression is lower in response to S. aureus exposure in mouse RAW 264.7 macrophage cells. Compared to wild-type cells, RAW 264.7 cells with Gls2 silenced by CRISPR-Cas9 genome editing have decreased IL1-β transcription and increased nitric oxide production after S. aureus exposure. GLS2 is an interesting candidate gene for complicated SAB due to its role in regulating glutamine metabolism, a key factor in leukocyte activation.

Group B Streptococcus (GBS) is an opportunistic pathogen that causes preterm birth and neonatal disease. Although GBS is known to exhibit vast diversity in virulence across strains, the mechanisms of GBS-associated pathogenesis are incompletely understood. We hypothesized that GBS strains of different genotypes would vary in their ability to elicit host inflammatory responses, and that strains associated with neonatal disease would induce different cytokine profiles than those associated with colonization. Using a multiplexed, antibody-based protein detection array, we found that production of a discrete number of inflammatory mediators by THP-1 macrophage-like cells was universally induced in response to challenge with each of five genetically distinct GBS isolates, while other responses appeared to be strain-specific. Key array responses were validated by ELISA using the initial five strains as well as ten additional strains with distinct genotypic and phenotypic characteristics. Interestingly, IL-6 was significantly elevated following infection with neonatal infection-associated sequence type (ST)-17 strains and among strains possessing capsule (cps) type III. Significant differences in production of IL1-β, IL-10 and MCP-2 were also identified across STs and cps types. These data support our hypothesis and suggest that unique host innate immune responses reflect strain-specific differences in virulence across GBS isolates. Such data might inform the development of improved diagnostic or prognostic strategies against invasive GBS infections.

To develop an alternative bio-control measure for multi-drug resistant pathogenic Aeromonas hydrophila, which causes motile Aeromonas septicemia in fish, novel virulent phage (AHP-1) was isolated from carp tissues. Morphological analysis by transmission electron microscopy revealed that AHP-1 belongs to Myoviridae family. AHP-1 displayed 81% of moderate adsorption by 25 min, and latent period of 40 min with burst size of 97 PFU mL^-1 at an optimal multiplicity of infection (MOI) 0.1. AHP-1 was stable over a broad range of pH (4-11), temperature (4-50 °C), and salinity (0.1-3.5%). Both time and MOI dependent in vitro A. hydrophila growth inhibition was observed with AHP-1. AHP-1 (10 MOI) showed higher growth inhibition against A. hydrophila than chloramphenicol (5 μg mL^-1), and combined treatment was more promising than individuals. Immune gene expression analysis of zebrafish upon continuous bath exposure to AHP-1 resulted significantly higher (il-6 and sod-1) or slight induction (tnf-α, il1-β, il-10, and cxcl-8a) than controls at beginning of the phage exposure, but those lowered to basal level by day 12 post-phage exposure. It suggests no adverse immune responses have occurred for the AHP-1 dose that used, and have potential for the phage therapy. Further detailed in vivo studies are needed to confirm the protective efficacy of newly isolated AHP-1 against A. hydrophila infection.

<A<em>b</em>stractText>A su<em>b</em>stantial proportion patients with inflammatory <em>b</em>owel disease (IBD) have a primary non-response to inflixima<em>b</em>; markers are needed to identify patients most likely to respond to treatment. We investigated whether production of tumor necrosis factor (TNF) <em>b</em>y peripheral <em>b</em>lood mononuclear cells (PBMCs) can <em>b</em>e used as a marker to predict response.</A<em>b</em>stractText><p><div>(<em>b</em>)METHODS</<em>b</em>)</div>We performed a prospective study of 41 adults with IBD (mean age, 38 years; 21 male; 21 with Crohn's disease and 20 with ulcerative colitis) not treated with a <em>b</em>iologic agent within the past 6 months; patients were given their first infusion of inflixima<em>b</em> at a hospital or clinic in Berlin, Germany. We collected data on clinical scores, levels of C-reactive protein, and ultrasound results (Lim<em>b</em>erg scores) at <em>b</em>aseline (<em>b</em>efore the first infusion) and after 6 weeks (3^rd inflixima<em>b</em> infusion). PMBCs were o<em>b</em>tained from patients at <em>b</em>aseline and 10 healthy individuals (controls) and incu<em>b</em>ated with lipopolysaccharide. We measured production of cytokines (TNF, interleukin 1 [<em>IL1</em>], IL6, IL8, <em>IL1</em>0, <em>IL1</em>2p70, and IL22) <em>b</em>y ELISA and performed cytometric <em>b</em>ead array and flow cytometry analyses. The primary endpoint was clinical response (decrease in Harvey Bradshaw Index scores of 2 or more or decrease in partial Mayo scores of 3 or more at week 6) in patients with PBMCs that produced high vs low levels of TNF.</p><A<em>b</em>stractText>Responders had a shorter median disease duration (P=.018) and higher median Lim<em>b</em>erg score (P=.021), than nonresponders. Baseline PBMCs from responders produced significantly more TNF (P=.049) and IL6 (P=.028) than from nonresponders; a level of 500 pg/ml TNF identified responders with 82% sensitivity and 78% specificity. In patients with Crohn's disease, this cutoff value (500 pg/ml TNF) identified responders with 100% sensitivity and 82% specificity; TNF levels a<em>b</em>ove this level were independently associated with response to inflixima<em>b</em> in multivariate analysis (odds ratio, 16.2; 9% CI, 1.8-148.7; P=.014). The percentage of TNF-positive cells was higher among CD14+ monocytes than lymphocytes after stimulation.</A<em>b</em>stractText><A<em>b</em>stractText>Production of a high level of TNF <em>b</em>y PBMCs (specifically CD14+ cells) from patients with IBD can identify those most likely to have a clinical response to inflixima<em>b</em> therapy. In patients with Crohn's disease, a cutoff value of 500 pg/ml TNF identified responders with 100% sensitivity and 82% specificity.</A<em>b</em>stractText>

BACKGROUND This study aimed to identify the pharmacological targets and mechanisms of action of the traditional Chinese medicine, formononetin, in the treatment of Alzheimer's disease (AD) using network pharmacological analysis. MATERIAL AND METHODS Targets of AD were obtained by using DisGeNET gene discovery platform, the herbal ingredients target (HIT) database, the SuperPred, and the SwissTargetPrediction compound target prediction platforms. Pathogenic and therapeutic targets were imported to the STRING biological database, and Cytoscape network integration software was used to construct component-target and disease-target interaction networks. Core targets were identified by topological analysis and were further tested to identify the biological processes and signaling pathways. RESULTS Seven key target genes for formononetin in the treatment of patients with AD were identified, including estrogen receptor alpha (ESR1), peroxisome proliferator-activated receptor gamma (PPARG), tumor protein p53 (TP53), sirtuin 1 (SIRT1), tumor necrosis factor (TNF), cytochrome P450 19A1 (CYP19A1), and nuclear factor (erythroid-derived 2)-like 2 (NFE2L2). The biological processes included hormone metabolism, regulation of nucleoside, nucleotide and nucleic acid metabolism, apoptosis, energy pathways, metabolism, cell communication, and signal transduction. The signaling pathways included histone acetylation and deacetylation (HDAC) class I, regulation of p38-alpha/beta, p38 mitogen-activated protein kinase (MAPK) signaling pathway, bone morphogenetic protein (BMP) receptor signaling, interleukin-1 (IL1) mediated signaling events, the tumor necrosis factor (TNF) receptor signaling pathway, and cytoplasmic and nuclear Smad2/3 signaling. CONCLUSIONS Pharmacological network analysis was used to identify the gene targets and mechanisms of formononetin treatment in patients with AD.

Immune thrombocytopenia (ITP) is an autoimmune disease which leads to accelerated platelet clearance. We investigated the plasma cytokine, chemokine and growth factor signatures and their clinical significance in pediatric ITP patients during acute, chronic and follow-up stages as well as the effects of intravenous immunoglobulin (IVIg) treatment, by using the Multiplex technology. In acute ITP before and/or after IVIg treatment we found significantly increased plasma levels of the pro- (tumour necrosis factor-α (TNF-α), interleukin IL-15) and anti- (IL-1 receptor antagonist (Ra), IL-10 and the growth factor interferon γ-induced protein (IP-10)) inflammatory cytokines, compared to healthy controls. Except for IL1-Ra, these cytokines decreased to normal levels in chronic patients. In contrast, growth-regulated α protein (GRO) and soluble CD40 ligand (sCD40L), known as platelet-derived molecules, were found to be significantly decreased in acute and increased in chronic ITP patients compared to healthy controls. GRO levels positively correlated with the platelet counts in the follow-up and chronic cohort. Monocyte counts showed a significant positive correlation only with IP-10 levels in acute ITP after IVIg treatment and follow-up patients. Expression levels of mRNAs for macrophage inflammatory protein MIP1-β, IL-1Ra and GRO determined in peripheral blood mononuclear cells (PBMCs) were significantly reduced in both acute and chronic ITP compared to controls. Our findings suggest that the different clinical presentation of acute and chronic pediatric ITP and to a lesser extent the IVIg treatment effects are characterized overall by a counterbalanced cytokine, chemokine and growth factor pattern response that might exert a pathogenic role in this disease.

The molecular mechanisms underlying the capability of pituitary tumours to avoid unregulated cell proliferation are still not well understood. However, the NF-κB transcription factor, which is able to modulate not only cellular senescence but also tumour progression, has emerged as a targeted candidate. This work was focused on the NF-κB role in cellular senescence during the progression of experimental pituitary tumours. Also, the contribution of the signalling pathways in senescence-associated NF-κB activation, and the senescence-associated secretory phenotype (SASP) and pro-survival-NF-κB target genes transcription were analysed. A robust NF-κB activation was seen at E20-E40 of tumour development accompanied by a marked SA-β-Gal co-reactivity in the tumour pituitary parenchyma. The induction of TNFα and IL1-β as specific SASP-related NF-κB target genes as well as bcl-2 and bcl-xl pro-survival genes was shown to be accompanied by increases in the p-p38 MAPK protein levels, starting at the E20 stage and strengthening from 40 to 60 days of tumour growth. It is noteworthy that, p-JNK displayed a similar pattern of activation during pituitary tumour development, while p-AKT and p-ERK1/2 were downregulated. By employing a pharmacological strategy to abrogate NF-κB activity, we demonstrated a marked reduction in SA-β-Gal activity and a slight decrease in Ki67 immunopositive cells after NF-κB blockade. These results suggest a central role for NF-κB in the regulation of the cellular senescence program, leading to the strikingly benign intrinsic nature of pituitary adenomas.