TLR9 Agonist Protects Mice from Radiation-Induced Gastrointestinal Syndrome

Radiation-induced gastrointestinal syndrome (RIGS) is due to the clonogenic loss of crypt cells and villi depopulation, resulting in disruption of mucosal barrier, bacterial invasion, inflammation and sepsis. Intestinal macrophages could recognize invading bacterial DNA via TLR9 receptors and transmit regenerative signals to the neighboring crypt. We therefore investigated whether systemic administration of designer TLR9 agonist could ameliorate RIGS by activating TLR9.Male C57Bl6 mice were distributed in four experimental cohorts, whole body irradiation (WBI) (8.4-10.4 Gy), TLR9 agonist (1 mg/kg s.c.), 1 h pre- or post-WBI and TLR9 agonist+WBI+iMyd88 (pretreatment with inhibitory peptide against Myd88). Animals were observed for survival and intestine was harvested for histological analysis. BALB/c mice with CT26 colon tumors in abdominal wall were irradiated with 14 Gy single dose of whole abdominal irradiation (AIR) for tumor growth study.Mice receiving pre-WBI TLR9 agonist demonstrated improvement of survival after 10.4 Gy (p<0.03), 9.4 Gy (p<0.008) and 8.4 Gy (p<0.002) of WBI, compared to untreated or iMyd88-treated controls. Post-WBI TLR9 agonist mitigates up to 8.4 Gy WBI (p<0.01). Histological analysis and xylose absorption test demonstrated significant structural and functional restitution of the intestine in WBI+TLR9 agonist cohorts. Although, AIR reduced tumor growth, all animals died within 12 days from RIGS. TLR9 agonist improved the survival of mice beyond 28 days post-AIR (p<0.008) with significant reduction of tumor growth (p<0.0001).TLR9 agonist treatment could serve both as a prophylactic or mitigating agent against acute radiation syndrome and also as an adjuvant therapy to increase the therapeutic ratio of abdominal Radiation Therapy for Gastro Intestinal malignancies

An In Situ Autologous Tumor Vaccination with Combined Radiation Therapy and TLR9 Agonist Therapy

PURPOSE:Recent studies have shown that a new generation of synthetic agonist of Toll-like receptor (TLR) 9 consisting a 3'-3'-attached structure and a dCp7-deaza-dG dinucultodie shows more potent immunostimulatory effects in both mouse and human than conventional CpG oligonucleotides. Radiation therapy (RT) provides a source of tumor antigens that are released from dying, irradiated, tumor cells without causing systemic immunosuppression. We, therefore, examined effect of combining RT with a designer synthetic agonist of TLR9 on anti-tumoral immunity, primary tumor growth retardation and metastases in a murine model of lung cancer. METHODS:Grouped C57BL/6 and congenic B cell deficient mice (B(-/-)) bearing footpad 3LL tumors were treated with PBS, TLR9 agonist, control oligonucelotide, RT or the combination of RT and TLR9 agonist. Immune phenotype of splenocytes and serum IFN-γ and IL-10 levels were analyzed by FACS and ELISA, 24 h after treatment. Tumor growth, lung metastases and survival rate were monitored and tumor specific antibodies in serum and deposition in tumor tissue were measured by ELISA and immunofluorescence. RESULTS:TLR9 agonist expanded and activated B cells and plasmacytoid dendritic cells in wild-type mice and natural killer DCs (NKDCs) in B cell-deficient (B(-/-)) mice bearing ectopic Lewis lung adenocarcinoma (3LL). Combined RT with TLR9 agonist treatment inhibited 3LL tumor growth in both wild type and B(-/-) mice. A strong tumor-specific humoral immune response (titer: 1/3200) with deposition of mouse IgG auto-antibodies in tumor tissue were found in wildtype mice, whereas the number of tumor infiltrating NKDCs increased in B(-/-) mice following RT+ TLR9 agonist therapy. Furthermore, mice receiving combination therapy had fewer lung metastases and a higher survival than single treatment cohorts. CONCLUSIONS:Combination therapy with TLR9 agonist and RT induces systemic anti-tumoral humoral response, augments tumoral infiltration of NKDCs, reduces pulmonary metastases and improves survival in a murine model of 3LL cancer

A Rapid Method to Regenerate Piezoelectric Microcantilever Sensors (PEMS)

Piezoelectric microcantilever sensors (PEMS) can be sensitive tools for the detection of proteins and cells in biological fluids. However, currently available PEMS can only be used a single time or must be completely stripped and refunctionalized prior to subsequent uses. Here we report the successful use of an alternative regeneration protocol employing high salt concentrations to remove the target, leaving the functional probe immobilized on the microcantilever surface. Our model system employed the extracellular domain (ECD) of recombinant human Epidermal Growth Factor Receptor (EGFR) as the probe and anti-human EGFR polyclonal antibodies as the target. We report that high concentrations of MgCl2 dissociated polyclonal antibodies specifically bound to EGFR ECD immobilized on the sensor surface without affecting its bioactivity. This simple regeneration protocol both minimized the time required to re-conjugate the probe and preserved the density of probe immobilized on PEMS surface, yielding identical biosensor sensitivity over a series of assays

COVID-19 and Rheumatoid Arthritis Crosstalk: Emerging Association, Therapeutic Options and Challenges

Hyperactivation of immune responses resulting in excessive release of pro-inflammatory mediators in alveoli/lung structures is the principal pathological feature of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The cytokine hyperactivation in COVID-19 appears to be similar to those seen in rheumatoid arthritis (RA), an autoimmune disease. Emerging evidence conferred the severity and risk of COVID-19 to RA patients. Amid the evidence of musculoskeletal manifestations involving immune-inflammation-dependent mechanisms and cases of arthralgia and/or myalgia in COVID-19, crosstalk between COVID-19 and RA is often debated. The present article sheds light on the pathological crosstalk between COVID-19 and RA, the risk of RA patients in acquiring SARS-CoV-2 infection, and the aspects of SARS-CoV-2 infection in RA development. We also conferred whether RA can exacerbate COVID-19 outcomes based on available clinical readouts. The mechanistic overlapping in immune-inflammatory features in both COVID-19 and RA was discussed. We showed the emerging links of angiotensin-converting enzyme (ACE)-dependent and macrophage-mediated pathways in both diseases. Moreover, a detailed review of immediate challenges and key recommendations for anti-rheumatic drugs in the COVID-19 setting was presented for better clinical monitoring and management of RA patients. Taken together, the present article summarizes available knowledge on the emerging COVID-19 and RA crosstalk and their mechanistic overlaps, challenges, and therapeutic options

The Importance of Research on the Origin of SARS-CoV-2

The origin of the severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) virus causing the COVID-19 pandemic has not yet been fully determined. Despite the consensus about the SARS-CoV-2 origin from bat CoV RaTG13, discrepancy to host tropism to other human Coronaviruses exist. SARS-CoV-2 also possesses some differences in its S protein receptor-binding domain, glycan-binding N-terminal domain and the surface of the sialic acid-binding domain. Despite similarities based on cryo-EM and biochemical studies, the SARS-CoV-2 shows higher stability and binding affinity to the ACE2 receptor. The SARS-CoV-2 does not appear to present a mutational “hot spot” as only the D614G mutation has been identified from clinical isolates. As laboratory manipulation is highly unlikely for the origin of SARS-CoV-2, the current possibilities comprise either natural selection in animal host before zoonotic transfer or natural selection in humans following zoonotic transfer. In the former case, despite SARS-CoV-2 and bat RaTG13 showing 96% identity some pangolin Coronaviruses exhibit very high similarity to particularly the receptor-binding domain of SARS-CoV-2. In the latter case, it can be hypothesized that the SARS-CoV-2 genome has adapted during human-to-human transmission and based on available data, the isolated SARS-CoV-2 genomes derive from a common origin. Before the origin of SARS-CoV-2 can be confirmed additional research is required

A New Minimal-Stress Freely-Moving Rat Model for Preclinical Studies on Intranasal Administration of CNS Drugs

Purpose. To develop a new minimal-stress model for intranasal administration in freely moving rats and to evaluate in this model the brain distribution of acetaminophen following intranasal versus intravenous administration. Methods. Male Wistar rats received one intranasal cannula, an intra-cerebral microdialysis probe, and two blood cannulas for drug administration and serial blood sampling respectively. To evaluate this novel model, the following experiments were conducted. 1) Evans Blue was administered to verify the selectivity of intranasal exposure. 2) During a 1 min infusion 10, 20, or 40 μl saline was administered intranasally or 250 µl intravenously. Corticosterone plasma concentrations over time were compared as biomarkers for stress. 3) 200 µg of the model drug acetaminophen was given in identical setup and plasma, and brain pharmacokinetics were determined. Results. In 96 % of the rats, only the targeted nasal cavity was deeply colored. Corticosterone plasma concentrations were not influenced, neither by route nor volume of administration. Pharmacokinetics of acetaminophen were identical after intravenous and intranasal administration, although the Cmax in microdialysates was reached a little earlier following intravenous administration. Conclusion. A new minimal-stress model for intranasal administration in freely moving rats has been successfully developed and allows direct comparison with intravenous administration. KEY WORDS: acetaminophen; brain; intranasal infusion; microdialysis; pharmacokinetics

Modeling Personalized Adjuvant TreaTment in EaRly stage coloN cancer (PATTERN)

Aim To develop a decision model for the population-level evaluation of strategies to improve the selection of stage II colon cancer (CC) patients who benefit from adjuvant chemotherapy. Methods A Markov cohort model with a one-month cycle length and a lifelong time horizon was developed. Five health states were included; diagnosis, 90-day mortality, death other causes, recurrence and CC death. Data from the Netherlands Cancer Registry were used to parameterize the model. Transition probabilities were estimated using parametric survival models including relevant clinical and pathological covariates. Subsequently, biomarker status was implemented using external data. Treatment effect was incorporated using pooled trial data. Model development, data sources used, parameter estimation, and internal and external validation are described in detail. To illustrate the use of the model, three example strategies were evaluated in which allocation of treatment was based on (A) 100% adherence to the Dutch guidelines, (B) observed adherence to guideline recommendations and (C) a biomarker-driven strategy. Results Overall, the model showed good internal and external validity. Age, tumor growth, tumor sidedness, evaluated lymph nodes, and biomarker status were included as covariates. For the example strategies, the model predicted 83, 87 and 77 CC deaths after 5 years in a cohort of 1000 patients for strategies A, B and C, respectively. Conclusion This model can be used to evaluate strategies for the allocation of adjuvant chemotherapy in stage II CC patients. In future studies, the model will be used to estimate population-level long-term health gain and cost-effectiveness of biomarker-based selection strategies.Financial support for this study was provided by a grant from ZonMw (Grant number: 848015007). ZonMw had no role in designing the study, interpreting the data, writing the manuscript, and publishing the report

Urgent Need for Field Surveys of Coronaviruses in Southeast Asia to Understand the SARS-CoV-2 Phylogeny and Risk Assessment for Future Outbreaks

Phylogenetic analysis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is focused on a single isolate of bat coronaviruses (bat CoVs) which does not adequately represent genetically related coronaviruses (CoVs) [...

PORTAL: Pilot study on the safety and tolerance of preoperative melatonin application in patients undergoing major liver resection: a double-blind randomized placebo-controlled trial

<p>Abstract</p> <p>Background</p> <p>Major surgical procedures facilitate systemic endotoxinemia and formation of free radicals with subsequent inflammatory changes that can influence the postoperative course. Experimental data suggest that preoperative supraphysiological doses of melatonin, a potent immuno-modulator and antioxidant, would decrease postoperative infectious and non-infectious complications induced by major abdominal surgery.</p> <p>Methods/Design</p> <p>A randomized controlled double blind single center clinical trial with two study arms comprising a total of 40 patients has been designed to assess the effects of a single preoperative dose of melatonin before major liver resection. Primary endpoints include the determination of safety and tolerance of the regimen as well as clinical parameters reflecting pathophysiological functions of the liver. Furthermore, data on clinical outcome (infectious and non-infectious complications) will be collected as secondary endpoints to allow a power calculation for a randomized clinical trial aiming at clinical efficacy.</p> <p>Discussion</p> <p>Based on experimental data, this ongoing clinical trial represents an advanced element of the research chain from bench to bedside in order to reach the highest level of evidence-based clinical facts to determine if melatonin can improve the general outcome after liver resection.</p> <p>Trial Registration</p> <p>EudraCT200600530815</p