Targeting of a Fixed Bacterial Immunogen to Fc Receptors Reverses the Anti-Inflammatory Properties of the Gram-Negative Bacterium,Francisella tularensis,during the Early Stages of Infection

Production of pro-inflammatory cytokines by innate immune cells at the early stages of bacterial infection is important for host protection against the pathogen. Many intracellular bacteria, including Francisella tularensis, the agent of tularemia, utilize the anti-inflammatory cytokine IL-10, to evade the host immune response. It is well established that IL-10 has the ability to inhibit robust antigen presentation by dendritic cells and macrophages, thus suppressing the generation of protective immunity. The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date. In the current study, we hypothesized that F. tularensis polarizes antigen presenting cells during the early stages of infection towards an anti-inflammatory status characterized by increased synthesis of IL-10 and decreased production of IL-12p70 and TNF-α in an IFN-ɣ-dependent fashion. In addition, F. tularensis drives an alternative activation of alveolar macrophages within the first 48 hours post-infection, thus allowing the bacterium to avoid protective immunity. Furthermore, we demonstrate that targeting inactivated F. tularensis (iFt) to Fcγ receptors (FcɣRs) via intranasal immunization with mAb-iFt complexes, a proven vaccine strategy in our laboratories, reverses the anti-inflammatory effects of the bacterium on macrophages by down-regulating production of IL-10. More specifically, we observed that targeting of iFt to FcγRs enhances the classical activation of macrophages not only within the respiratory mucosa, but also systemically, at the early stages of infection. These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.     

Predicting Outcome on Admission and Post-Admission for Acetaminophen-Induced Acute Liver Failure Using Classification and Regression Tree Models

Background/Aim

Assessing prognosis for acetaminophen-induced acute liver failure (APAP-ALF) patients often presents significant challenges. King’s College (KCC) has been validated on hospital admission, but little has been published on later phases of illness. We aimed to improve determinations of prognosis both at the time of and following admission for APAP-ALF using Classification and Regression Tree (CART) models.

Methods

CART models were applied to US ALFSG registry data to predict 21-day death or liver transplant early (on admission) and post-admission (days 3-7) for 803 APAP-ALF patients enrolled 01/1998–09/2013. Accuracy in prediction of outcome (AC), sensitivity (SN), specificity (SP), and area under receiver-operating curve (AUROC) were compared between 3 models: KCC (INR, creatinine, coma grade, pH), CART analysis using only KCC variables (KCC-CART) and a CART model using new variables (NEW-CART).

Results

Traditional KCC yielded 69% AC, 90% SP, 27% SN, and 0.58 AUROC on admission, with similar performance post-admission. KCC-CART at admission offered predictive 66% AC, 65% SP, 67% SN, and 0.74 AUROC. Post-admission, KCC-CART had predictive 82% AC, 86% SP, 46% SN and 0.81 AUROC. NEW-CART models using MELD (Model for end stage liver disease), lactate and mechanical ventilation on admission yielded predictive 72% AC, 71% SP, 77% SN and AUROC 0.79. For later stages, NEW-CART (MELD, lactate, coma grade) offered predictive AC 86%, SP 91%, SN 46%, AUROC 0.73.

Conclusion

CARTs offer simple prognostic models for APAP-ALF patients, which have higher AUROC and SN than KCC, with similar AC and negligibly worse SP. Admission and post-admission predictions were developed.

Key Points

• Prognostication in acetaminophen-induced acute liver failure (APAP-ALF) is challenging beyond admission• Little has been published regarding the use of King’s College Criteria (KCC) beyond admission and KCC has shown limited sensitivity in subsequent studies• Classification and Regression Tree (CART) methodology allows the development of predictive models using binary splits and offers an intuitive method for predicting outcome, using processes familiar to clinicians• Data from the ALFSG registry suggested that CART prognosis models for the APAP population offer improved sensitivity and model performance over traditional regression-based KCC, while maintaining similar accuracy and negligibly worse specificity• KCC-CART models offered modest improvement over traditional KCC, with NEW-CART models performing better than KCC-CART particularly at late time points     

The power of proteasome inhibition in multiple myeloma

ABSTRACT

Introduction: Proteasome inhibitors (PIs) are therapeutic backbones of multiple myeloma treatment, with PI-based therapies being standards of care throughout the treatment algorithm. Proteasome inhibition affects multiple critical signaling pathways in myeloma cells and interacts synergistically with mechanisms of action of other conventional and novel agents, resulting in substantial anti-myeloma activity and at least additive effects.

Areas covered: This review summarizes the biologic effects of proteasome inhibition in myeloma and provides an overview of the importance of proteasome inhibition to the current treatment algorithm. It reviews key clinical data on three PIs, specifically bortezomib, carfilzomib, and ixazomib; assesses ongoing phase 3 trials with these agents; and looks ahead to the increasingly broad role of both approved PIs and PIs under investigation in the frontline and relapsed settings.

Expert commentary: Progress to date with PIs in multiple myeloma has been impressive, but there remain unmet needs and challenges, as well as increasing opportunities to optimize the use of these agents. Understanding discrepancies between PIs in terms of efficacy and safety profile is a key goal of ongoing research, along with proteomics-based efforts to identify potential biomarkers of sensitivity and resistance, thereby enabling increasingly personalized treatment approaches in the future.     

A dynamic view of trauma/hemorrhage-induced inflammation in mice: principal drivers and networks

Background

Complex biological processes such as acute inflammation induced by trauma/hemorrhagic shock/ (T/HS) are dynamic and multi-dimensional. We utilized multiplexing cytokine analysis coupled with data-driven modeling to gain a systems perspective into T/HS.

Methodology/Principal Findings

Mice were subjected to surgical cannulation trauma (ST) ± hemorrhagic shock (HS; 25 mmHg), and followed for 1, 2, 3, or 4 h in each case. Serum was assayed for 20 cytokines and NO₂ ⁻/NO₃ ⁻. These data were analyzed using four data-driven methods (Hierarchical Clustering Analysis [HCA], multivariate analysis [MA], Principal Component Analysis [PCA], and Dynamic Network Analysis [DyNA]). Using HCA, animals subjected to ST vs. ST + HS could be partially segregated based on inflammatory mediator profiles, despite a large overlap. Based on MA, interleukin [IL]-12p40/p70 (IL-12.total), monokine induced by interferon-γ (CXCL-9) [MIG], and IP-10 were the best discriminators between ST and ST/HS. PCA suggested that the inflammatory mediators found in the three main principal components in animals subjected to ST were IL-6, IL-10, and IL-13, while the three principal components in ST + HS included a large number of cytokines including IL-6, IL-10, keratinocyte-derived cytokine (CXCL-1) [KC], and tumor necrosis factor-α [TNF-α]. DyNA suggested that the circulating mediators produced in response to ST were characterized by a high degree of interconnection/complexity at all time points; the response to ST + HS consisted of different central nodes, and exhibited zero network density over the first 2 h with lesser connectivity vs. ST at all time points. DyNA also helped link the conclusions from MA and PCA, in that central nodes consisting of IP-10 and IL-12 were seen in ST, while MIG and IL-6 were central nodes in ST + HS.

Conclusions/Significance

These studies help elucidate the dynamics of T/HS-induced inflammation, complementing other forms of dynamic mechanistic modeling. These methods should be applicable to the analysis of other complex biological processes.     

Decline in local abundance of bottlenose dolphins (Tursiops truncatus) in the Bay of Islands,New Zealand

Regional populations of bottlenose dolphins (Tursiops truncatus) around New Zealand are genetically isolated from each other and the species was recently classified as nationally endangered based on relatively small population sizes and reports of high calf mortality. Here, we estimate the abundance and trends in one of these regional populations, the Bay of Islands, using a photo‐identification database collected from 1997 to 1999 and from 2003 to 2006, containing a total of 3,841 records of 317 individual dolphins. Estimates of abundance obtained with the robust design fluctuated widely but showed a significant decline in the number of dolphins present in the bay over time (7.5% annual rate of decline). Temporary emigration was random and fluctuated considerably (γ  =  0.18, SE = 0.07 to γ  =  0.84, SE = 0.06). Apparent survival was estimated at 0.928 (CI = 0.911–0.942). Seasonal estimates (26 seasons) obtained in POPAN also showed a significant decline in abundance (5.8% annual rate of decline). Despite the decline observed in local abundance, dolphins continue to be found regularly in the Bay of Islands, suggesting that fewer dolphins use the bay on regular basis. Consequently, it seems that a change in habitat use, mortality and possibly low recruitment could underlie the apparent local decline.     

BRAF Mutation Testing of Thyroid Fine-Needle Aspiration Specimens Enhances the Predictability of Malignancy in Thyroid Follicular Lesions of Undetermined Significance

Background/Objective: The Bethesda 2007 Thyroid Cytology Classification defines atypia of undetermined significance/follicular lesion of undetermined significance (AUS/FLUS) as a heterogeneous category of cases that are neither convincingly benign nor sufficiently atypical for a diagnosis of follicular neoplasm or suspicious for malignancy. At our institution, we refer to these cases as 'indeterminate' and they are further subclassified into two categories. BRAF mutation occurs in 40-60% of papillary thyroid carcinoma (PTC). In this study, we examined cases in the AUS/FLUS category in correlation with BRAF mutation analysis and surgical pathology outcome. Study Design: Thyroid fine-needle aspiration (FNA) cytology specimens interpreted as 'indeterminate' were selected from our files, and available remnants of thin-layer processed specimens were used for BRAF mutation analysis. Surgical pathology reports were reviewed for the final outcomes in these patients. Results: Of the 84 indeterminate cases with BRAF mutation analysis, only 49 had follow-up with surgical intervention. Sixteen cases had BRAF mutation. All of the BRAF-positive cases had a final diagnosis of PTC. Conclusions: The sensitivity and specificity of BRAF mutation in detecting PTC in FNA specimens with indeterminate diagnosis was 59.3 and 100%, respectively, while the positive and negative predictive values were 100 and 65.6%, respectively. The limited data supports the use of BRAF mutation analysis to predict the risk of malignancy in patients with indeterminate thyroid FNAs.     

Evaluation of 2′-α-fluorine modified nucleoside phosphonates as potential inhibitors of HCV polymerase

Ribonucleoside phosphonate analogues containing 2′-α-fluoro modifications were synthesized and their potency evaluated against HCV RNA polymerase. The diphosphophosphonate (triphosphate equivalent) adenine and cytidine analogues displayed potent inhibition of the HCV polymerase in the range of 1.9–2.1 μM, but only modest cell-based activity in the HCV replicon. Pro-drugs of the parent nucleoside phosphonates improved the cell-based activity.     

Changes of free, soluble conjugated and bound polyamine titers of jojoba explants under sodium chloride salinity in vitro

Jojoba (Simmondsia chinensis L.) single node explants were cultured in a basal medium supplemented with 17.8 microM 6-benzyladenine and four levels of sodium chloride concentration (0, 56.41, 112.82 and 169.23 mM). The free, the soluble conjugated and the insoluble bound forms of polyamines (PAs) (putrescine (Put), spermidine (Spd) and spermine (Spm)) were determined monthly during a 3-month proliferation stage. Free Put and Spd were found in higher levels in the control treatment, while Spm content was higher in the salt treatments. All soluble conjugated PAs were found to be in lower concentrations in explants growing on medium supplemented with salt, while the opposite was true for the insoluble bound PAs. It appeared that certain PAs and PAs forms could play a significant role in the adaptation mechanism of jojoba under saline conditions.     

Klebsiella pneumoniae disassembles host microtubules in lung epithelial cells

Klebsiella pneumoniae raises significant concerns to the health care industry as these microbes are the source of widespread contamination of medical equipment, cause pneumonia as well as other multiorgan metastatic infections and have gained multidrug resistance. Despite soaring mortality rates, the host cell alterations occurring during these infections remain poorly understood. Here, we show that during in vitro and in vivo K. pneumoniae infections of lung epithelia, microtubules are severed and then eliminated. This destruction does not require direct association of K. pneumoniae with the host cells, as microtubules are disassembled in cells that are distant from the infecting bacteria. This microtubule dismantling is dependent on the K. pneumoniae (Kp) gene ytfL as non‐pathogenic Escherichia coli expressing Kp ytfL disassemble microtubules in the absence of K. pneumoniae itself. Our data points to the host katanin catalytic subunit A like 1 protein (KATNAL1) and the katanin regulatory subunit B1 protein (KATNB1) as the gatekeepers to the microtubule severing event as both proteins localise specifically to microtubule cut sites. Infected cells that had either of these proteins knocked out maintained intact microtubules. Taken together, we have identified a novel mechanism that a bacterial pathogen has exploited to cause microtubule destruction within the host epithelia.