Comparative Philosophy

Zhang, Xianglong 張祥龍, Family and Filial Reverence: A Cross-Cultural Perspective 家與孝: 從中西間視野看

Ethics, Politics, and the Recognition of Agency in Early Confucianism: A Commentary on Loubna El Amine's Classical Confucian Political Thought: A New Interpretation

Material Conditions, Hierarchy, and Order in Early Confucian Political Thought: A Response to Reviewers

Ivanhoe, Philip J., Oneness: East Asian Conceptions of Virtue, Happiness, and How We Are All Connected

Chai, David, Zhuangzi and the Becoming of Nothingness

Chen, Bo 陳波, and Jiang, Yi 江怡, eds., Analytic Philosophy: Review and Reflection 分析哲學 —— 回顧與反省

Zeng, Zhaoshi 曾昭式, A New Theory of the Historical Development of Chinese Logic in the Pre-Qin Era 先秦邏輯新論

The Ethical Stance of the "Qiwulun (Discourse on Corresponding Things)" Abstract This essay analyses the second chapter of the Zhuangzi 莊子, the "Qiwulun 齊物論." After a brief examination of its main ideas, it will be argued that the "Qiwulun" needs to be considered not as an equalization that makes everything indistinguishable but as a discourse on corresponding things. A more attentive analysis of this correspondence among the myriad things will lead to the consideration of their mutual transformation. The conclusion is that, contrary to the ontotheological nature of Western metaphysics that imposes a single logic for its hierarchical structure Being-beings, the correspondence of myriad things allows the differentiation and assimilation of values in a comprehensive and harmonious fashion. Thus, the essay offers an ethical reading of the "Qiwulun" and considers its significance in the contemporary world.

The Politics of Writing Chinese Philosophy: X iong Shili's New Treatise on the Uniqueness of Consciousness and the "Crystallization of Oriental Philosophy" Abstract This article situates Xiong Shili's 熊十力 classic work New Treatise on the Uniqueness of Consciousness (Xin Weishi Lun 新唯識論, 1932) within the central dilemma of post-May Fourth China surrounding the concerns with so-called modern universalism and Chinese particularism. I look at the way the text portrays its author as situated both within particular traditions and outside of them (in a realm of universality) in order to show how the figure of the author is presented as a site wherein Chinese/Asian particularism and universalism can be fused. My central aim, in doing so, is to argue that within the text's discourse on the positioning of its author resides an implicit argument for the universality of Chinese or Asian philosophy—as interpreted and subsumed by the text—and against the hegemonic intentions of Western philosophy. Yet I also suggest that the text reiterates a conception of universality and an intention to monopolize the universal which are characteristic of hegemonic discourses.

The Indispensability of Moral Cultivation in Confucian Politics

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Acta Biotheoretica

A Word of Welcome to Our New Editorial Board Members, and Presentation of the New Editorial Board

An Organisational Approach to Biological Communication Abstract This paper aims to provide a philosophical and theoretical account of biological communication grounded in the notion of organisation. The organisational approach characterises living systems as organised in such a way that they are capable to self-produce and self-maintain while in constant interaction with the environment. To apply this theoretical framework to the study of biological communication, we focus on a specific approach, based on the notion of influence, according to which communication takes place when a signal emitted by a sender triggers a change in the behaviour of the receiver that is functional for the sender itself. We critically analyse the current formulations of this account, that interpret what is functional for the sender in terms of evolutionary adaptations. Specifically, the adoption of this etiological functional framework may lead to the exclusion of several phenomena usually studied as instances of communication, and possibly even of entire fields of investigation such as synthetic biology. As an alternative, we reframe the influence approach in organisational terms, characterising functions in terms of contributions to the current organisation of a biological system. We develop a theoretical account of biological communication in which communicative functions are distinguished from other types of biological functions described by the organisational account (e.g. metabolic, ecological, etc.). The resulting organisational-influence approach allows to carry out causal analyses of current instances of phenomena of communication, without the need to provide etiological explanations. In such a way it makes it possible to understand in terms of communication those phenomena which realise interactive patterns typical of signalling interactions—and are usually studied as such in scientific practice—despite not being the result of evolutionary adaptations. Moreover, this approach provides operational tools to design and study communicative interactions in experimental fields such as synthetic biology.

Mathematical Modeling of Substrates Fluxes and Tumor Growth in the Brain Abstract The aim of this article is to show how a tumor can modify energy substrates fluxes in the brain to support its own growth. To address this question we use a modeling approach to explain brain nutrient kinetics. In particular we set up a system of 17 equations for oxygen, lactate, glucose concentrations and cells number in the brain. We prove the existence and uniqueness of nonnegative solutions and give bounds on the solutions. We also provide numerical simulations.

Multidrug Therapy for HIV Infection: Dynamics of Immune System Abstract A mathematical model of the dynamics of the immune system is considered to illustrate the effect of its response to HIV infection, i.e. on viral growth and on T-cell dynamics. The specific immune response is measured by the levels of cytotoxic lymphocytes in a human body. The existence and stability analyses are performed for infected steady state and uninfected steady state. In order to keep infection under control, roles of drug therapies are analyzed in the presence of efficient immune response. Numerical simulations are computed and exhibited to illustrate the support of the immune system to drug therapies, so as to ensure the decay of infection and to maintain the level of healthy cells.

The Coherence of Evolutionary Theory with Its Neighboring Theories Abstract Evolutionary theory coheres with its neighboring theories, such as the theory of plate tectonics, molecular biology, electromagnetic theory, and the germ theory of disease. These neighboring theories were previously unconceived, but they were later conceived, and then they cohered with evolutionary theory. Since evolutionary theory has been strengthened by its several neighboring theories that were previously unconceived, it will be strengthened by infinitely many hitherto unconceived neighboring theories. This argument for evolutionary theory echoes the problem of unconceived alternatives. Ironically, however, the former recommends that we take the realist attitude toward evolutionary theory, while the latter recommends that we take the antirealist attitude toward it.

Robust Model Selection and Estimation for Censored Survival Data with High Dimensional Genomic Covariates Abstract When relating genomic data to survival outcomes, there are three main challenges that are the censored survival outcomes, the high-dimensionality of the genomic data, and the non-normality of data. We propose a method to tackle these challenges simultaneously and obtain a robust estimation of detecting significant genes related to survival outcomes based on Accelerated Failure Time (AFT) model. Specifically, we include a general loss function to the AFT model, adopt model regularization and shrinkage technique, cope with parameters tuning and model selection, and develop an algorithm based on unified Expectation–Maximization approach for easy implementation. Simulation results demonstrate the advantages of the proposed method compared with existing methods when the data has heavy-tailed errors and correlated covariates. Two real case studies on patients are provided to illustrate the application of the proposed method.

Clearing New Ground

Walking the Line: A Tempered View of Contingency and Convergence in Life's History

Causally Modeling Adaptation to the Environment Abstract Brandon claims that to explain adaptation one must specify fitnesses in each selective environment and specify the distribution of individuals across selective environments. Glymour claims, using an example of the adaptive evolution of costly plasticity in a symmetric environment, that there are some predictive or explanatory tasks for which Brandon's claim is limited. In this paper, I provide necessary conditions for carrying out Brandon's task, produce a new version of the argument for his claim, and show that Glymour's reasons for making his claim are problematic. I provide a few interpretations of Glymour's argument but ultimately raise worries for what I take to be the key premises.

Control by Viability in a Chemotherapy Cancer Model Abstract The aim of this study is to provide a feedback control, called the Chemotherapy Protocol Law, with the purpose to keep the density of tumor cells that are treated by chemotherapy below a "tolerance level" \(L_c\) , while retaining the density of normal cells above a "healthy level" \(N_c\) . The mathematical model is a controlled dynamical system involving three nonlinear differential equations, based on a Gompertzian law of cell growth. By evoking viability and set-valued theories, we derive sufficient conditions for the existence of a Chemotherapy Protocol Law. Thereafter, on a suitable viability domain, we build a multifunction whose selections are the required Chemotherapy Protocol Laws. Finally, we propose a design of selection that generates a Chemotherapy Protocol Law.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Gynecology and Obstetrics

Reply to comments on "Risk factors, microbiology and management of infected lymphocyst after lymphadenectomy for gynecologic malignancies"

New associate editor: Prof. Dr. rer.nat. Udo Jeschke

Reply to the comments on "Modified hysterectomy for placenta increta and percreta: modifications of what?"

Modified hysterectomy for placenta increta and percreta: modifications of what?

Risk factors, microbiology, and management of infected lymphocyst after lymphadenectomy for gynecologic malignancies: letter to the editor

Calprotectin in pregnancy and pregnancy-associated diseases: a systematic review and prospective cohort study Abstract Purpose Calprotectin, a marker of acute and chronic inflammation, may play a role in pregnancy-associated disorders. We aimed to summarize available clinical data on calprotectin in pregnancy and to establish normal values of calprotectin during the course of pregnancy. Methods We performed a systematic review of the databases PubMed and Cochrane Central Register of Controlled Trials to identify experimental and clinical evidence assessing the role of calprotectin in pregnancy. In addition, we performed a prospective cohort study assessing serum and urine calprotectin throughout pregnancy. Results We identified 17 studies investigating 1638 pregnant women, 151 newborns, and 99 non-pregnant controls, measuring calprotectin in different compartments. Calprotectin was present in meconium and elevated in fecal samples of pregnant women with active inflammatory bowel disease. In women with pregnancy-induced hypertension, mild and severe preeclampsia (PE), calprotectin was significantly elevated in maternal plasma and serum, but not in fetal serum, amniotic fluid, and umbilical cord blood. For the cohort study, we recruited 196 pregnant women. PE and concomitant renal disease were present in 6/196 (3%) and 11/196 (5.6%) of women, respectively. Throughout pregnancy, median serum and urine levels of calprotectin largely exceed reported concentrations of the healthy non-pregnant population, but showed no significant variations between trimesters 1–3 and post-partum. Calprotectin in serum was correlated with systolic blood pressure and in urine with leukocytes and total protein. No significant differences were found in subgroup analyses of smokers vs. non-smokers, PE vs. none, and renal disease (kidney stones, reflux) vs. none. Conclusion Calprotectin concentrations in amnion fluid and stools serve as potential indicators of inflammatory states during pregnancy. Urinary calprotectin concentrations are continuously high during pregnancy and show no significant variations between trimesters 1–3 and post-partum.

The security of radical trachelectomy in the treatment of IA–IIA cervical carcinoma requires further evaluation: updated meta-analysis and trial sequential analysis Abstract Purpose The aim of this study was to assess the security of radical trachelectomy (RT) in the treatment of IA–IIA cervical carcinoma and conducted a new survey based upon the results of previous researches. Methods The PMC, PubMed, Web of Science, Cochrane and EMBASE databases were retrieved to collect prospective clinical controlled trials (CCTs) published from 1984 to 2018. The oncologic outcomes were evaluated by meta-analysis, trial sequence analysis (TSA) and statistical analysis. Results Five prospective CCTs were collected in this study. The recurrence rate and mortality of RT was similar to that of radical hysterectomy (RH), which was consistent with the oncologic outcomes of meta-analysis and TSA. Patients with tumors 2–4 cm in diameter were more likely to receive RH, which may be a potential factor in the higher rate of adjuvant chemotherapy in the this group, and RH was significantly associated with the risk of intraoperative blood transfusion. It is notable that considerable negative margin was achieved by radical abdominal trachelectomy (RAT), and the clinical effect of RAT was slightly better than that of radical vaginal trachelectomy (RVT). However, the TSA results showed that the cumulative cases were not up to the required sample size to obtain the true negative or positive results. Conclusions It is safe and effective for early-stage patients with cervical cancer whose lesions are less than 2 cm to receive RVT. For those patients with lesions 2–4 cm who desire fertility preservation and without any evidence of infertility, RAT can be a feasible alternative to RH under fully informed consent. However, more CCTs with larger sample size are still required for further validation.

A new approach to prevent ischemia/reperfusion injury in a rat model: remote ischemic conditioning Abstract Purpose To evaluate the effect of remote ischemic conditioning (RIC) on ovarian ischemia/reperfusion injury in a rat model. Methods A total of 36 Wistar albino rats with a body weight of 220–250 g were used for this study. Right adnexal torsion was performed for 180 min, and at the end of the period, the adnex was released and the abdomen was reclosed for 180 min for reperfusion. Torsion and detorsion procedures were applied to all rats except group 1 (sham, control). The right lower extremity was tied to perform remote tissue ischemia in groups 3, 4, 5, and 6. The goal of the procedure, which was purplish discoloration and pulselessness of the extremity, was maintained. After 5 min of ischemia, reperfusion was achieved for 5 min. Repeating this procedure 3 times was defined as hypoxia attacks (RIC). Retrieved ovaries were examined for tissue injury with biochemical, histopathologic, and immunohistochemical analysis. Results Unlike the control group, vascular congestion, hemorrhage, edema, and inflammatory cell infiltration were observed in group 2 (only I/R [ischemia/reperfusion]). In groups 3 (I/R + RIC), 4 (I/R + RIC), 5 (I/R + RIC), and 6 (I/R + RIC), edema and inflammatory cell infiltration were not observed. However, vascular congestion and hemorrhage that were detected in these groups were higher than in group 1 (Control) and less than in group 2 (I/R). The Caspase-3 Index was found to be increased in all groups compared to group 1 (P < .001). However, the increase in the RIC-performed groups was significantly less than in group 2. The apoptotic index, which was determined by the TUNEL, was also found to be increased in all groups compared to group 1 (P < .001). When the comparison was made in relation to group 2, the decrease of AI in RIC-performed groups was statistically significant, except the decrease in group 6 (P = .29). Conclusions It is not clinically conceivable to prepare the tissue for pre-ischemia in ovarian torsion. However, the RIC application, which will be initiated if torsion is suspected when arrangements are made for surgery, might be a simple, effective, and inexpensive approach to prevent I/R injury in the clinic.

Efficacy of pegylated liposomal doxorubicin maintenance therapy in platinum-sensitive recurrent epithelial ovarian cancer: a retrospective study Abstract Objective To examine the effectiveness of pegylated liposomal doxorubicin (PLD) maintenance therapy (intravenous administration at dose 40 mg/m2 on day 1, repeated every 4 weeks) after first-line salvage chemotherapy for platinum-sensitive recurrent epithelial ovarian cancer. Methods This retrospective cohort study examined women with a first recurrence of platinum-sensitive epithelial ovarian cancer diagnosed between 2005 and 2015. Eligible cases had PLD maintenance following the first-line salvage chemotherapy (n = 28). Outcomes of interest included adverse events related to PLD maintenance therapy and survival outcome after the first recurrence. Results The median number of PLD maintenance cycles was 7.5 (range 2–26), and 11 (40%) women received ≥ 12 cycles. The median cumulative dose of PLD was 432.5 mg/m2 (range 120–1200 mg/m2). No women developed cardiotoxicity or secondary malignancies. There were 16 (57%) women who developed any grade of adverse events, including 3 (11%) women who developed grade 3 adverse events. There were no grade 4 adverse events. The most common adverse event was mucositis (n = 7, 25%). Dose reduction due to adverse events occurred in 14 (50%) women including 3 (11%) women with discontinuation due to toxicity. Median progression-free survival and overall survival after the initiation of PLD maintenance was 14.5 months (2-year rate 21.1%) and 51.2 months (5-year rate 43.4%), respectively. Conclusion Our study suggests that PLD maintenance therapy for platinum-sensitive recurrent ovarian cancer is relatively well tolerated with the use of dose reduction to manage toxicity. Our study suggests that PLD maintenance therapy may be effective for women with platinum-sensitive recurrent epithelial ovarian cancer.

Current practice and physicians' opinion about preoperative hair removal as a part of ERAS pathway implementation in gynecology and gynecology–oncology: a NOGGO-AGO survey of 148 gynecological departments in Germany Abstract Purpose To gather standardized information about current practices and doctors' opinions on preoperative hair removal (PHR) from the surgical site and to evaluate the extent of PHR as one of the elements of enhanced recovery after surgery (ERAS) pathways that is established in the clinical routine in gynecology and gynecology–oncology departments in Germany. Methods We performed a nationwide survey among 638 primary, secondary and tertiary health care gynecological departments in Germany. Data were obtained by sending a multiple-choice questionnaire about preoperative management of hair removal. The authors also evaluated the awareness of doctors regarding PHR as well as the method and time frames of PHR. The results were compared to the existing standard of procedure (SOP) and guidelines. Results 148 units (23.2%) took part in the survey; participants in the survey were mostly chief physicians in 47.3% of the cases. Half (50.7%) of all the responses came from certified gynecological cancer centers. A SOP regarding PHR was reported as present in 113 clinics (76.4%). 83.8% of all units are performing PHR for midline laparotomy, 52.7% in laparoscopic operations, and 45.3% in vaginal operations. 48% used a clipper, while 43.2% utilized a single-use razor. 56.1% shaved instantly before the operation, whereas 35.8% did it the day before and earlier. 40.3% of chief physicians believe that PHR causes more surgical site infections (SSI) compared to only 11.5% of junior doctors. Conclusion PHR in gynecological departments in Germany is performed very heterogeneously and SOPs are often not based on guidelines and ERAS principles. Around one-third of the German gynecological clinics keep strictly to the guidelines. The awareness on PHR and SSI among junior doctors is very low.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Clinical & Experimental Metastasis

Effect of bisphosphonates on overall survival in subgroups of patients with prostate cancer Abstract Adjuvant therapy with bisphosphonates in prostate cancer is effective in improving bone mineral density and thus reducing fractures and skeletal-related events. We analyzed the effect of bisphosphonates on overall survival (OS) in subgroups of patients with prostate cancer. A systematic literature search was conducted of the PubMed database and the bibliographies of related studies. The long-term OS rates were extracted from every eligible trial. The hazard ratio (HR) was pooled with the fixed effects model, and preplanned subgroup analyses were performed. The search yielded 112 articles, of which 10 articles with 13 patient subgroups met the eligibility criteria. The meta-analysis of all 13 subgroups showed that adjuvant bisphosphonate therapy did not significantly improve OS versus the control group (HR = 0.961, 95% CI 0.899–1.026, p = 0.233) with low heterogeneity (I2 = 13.47%, degrees of freedom = 12, p = 0.336). There was no significant improvement in OS with the addition of bisphosphonates in the major subgroup analyses (metastatic (M1) versus non-metastatic, clodronate versus zoledronic acid, castration-sensitive prostate cancer (CSPC) versus castration-refractory prostate cancer). When the subgroups were further divided, adjuvant bisphosphonate therapy significantly improved OS in patients with CSPC + M1 (HR = 0.874, 95% CI 0.778–0.982, p = 0.023; I2 = 0.0%, degrees of freedom = 3, p = 0.579). Our study demonstrated that bisphosphonates do not significantly improve long-term OS in patients with prostate cancer. However, adjuvant bisphosphonate therapy significantly improves OS in the subgroup of patients with CSPC + M1.

Quantitative assessment of breast cancer liver metastasis expansion with patient-derived xenografts Abstract Advanced breast cancer often spreads to the bone, brain, liver, and lungs. The survival time of a patient with breast cancer liver metastasis is often less than 9 months without treatment. Experimental model systems often focus on the lung as a site of metastatic relapse, and therefore, there is less of an understanding of the biological processes that occur during expansive liver metastasis growth. In these studies, 14 genetically distinct breast cancer patient-derived xenografts (PDXs) were characterized for growth in the liver after portal vein injection of cancer cells. Growth in the liver occurred in 12 of 14 models, and the relative growth rate across the PDXs was overall similar to growth in the mammary gland. Pathological and immunohistochemical analyses revealed that the proliferation rates of metastases were relatively similar as the metastases expanded until the tumors became necrotic, and then slightly lower proliferation rates were observed. There were influxes of macrophages and neutrophils as the metastases increased in size, suggesting these innate immune cells may result in differential responses to therapeutics in micrometastases compared to macrometastases. The development and characterization of these models is important as future studies can utilize this information to determine if targeted therapies can slow the progression of metastatic disease at different stages in the liver.

The extracellular matrix in tumor progression and metastasis Abstract The extracellular matrix (ECM) constitutes the scaffold of tissues and organs. It is a complex network of extracellular proteins, proteoglycans and glycoproteins, which form supramolecular aggregates, such as fibrils and sheet-like networks. In addition to its biochemical composition, including the covalent intermolecular cross-linkages, the ECM is also characterized by its biophysical parameters, such as topography, molecular density, stiffness/rigidity and tension. Taking these biochemical and biophysical parameters into consideration, the ECM is very versatile and undergoes constant remodeling. This review focusses on this remodeling of the ECM under the influence of a primary solid tumor mass. Within this tumor stroma, not only the cancer cells but also the resident fibroblasts, which differentiate into cancer-associated fibroblasts (CAFs), modify the ECM. Growth factors and chemokines, which are tethered to and released from the ECM, as well as metabolic changes of the cells within the tumor bulk, add to the tumor-supporting tumor microenvironment. Metastasizing cancer cells from a primary tumor mass infiltrate into the ECM, which variably may facilitate cancer cell migration or act as barrier, which has to be proteolytically breached by the infiltrating tumor cell. The biochemical and biophysical properties therefore determine the rates and routes of metastatic dissemination. Moreover, primed by soluble factors of the primary tumor, the ECM of distant organs may be remodeled in a way to facilitate the engraftment of metastasizing cancer cells. Such premetastatic niches are responsible for the organotropic preference of certain cancer entities to colonize at certain sites in distant organs and to establish a metastasis. Translational application of our knowledge about the cancer-primed ECM is sparse with respect to therapeutic approaches, whereas tumor-induced ECM alterations such as increased tissue stiffness and desmoplasia, as well as breaching the basement membrane are hallmark of malignancy and diagnostically and histologically harnessed.

Circulating microRNAs in head and neck cancer: a scoping review of methods Abstract Circulating microRNAs have been described as head and neck cancer biomarkers in multiple anatomical subsites including the oral cavity, nasopharynx, larynx, salivary glands and the skin. While there is an expanding volume of published literature showing the significance of individual or panels of microRNAs, the clinical validation of candidate biomarkers is lacking. The various methods used to collect, store, process and interpret these microRNAs are likely introducing bias and contributing to the inconsistent results. A systematic scoping review was conducted using PRISMA standards to identify published English literature between 2007 and 2018. Pubmed and EMBASE databases were searched using specific keyword combinations related to head and neck cancer, circulating samples (whole blood, plasma or serum) and microRNA. Following the title and abstract review, two primary authors appraised the articles for their suitability to include in the review based on the detail of methodological descriptions. Thirty suitable articles were identified relating to nasopharyngeal carcinoma, oral cavity, oropharyngeal and laryngeal squamous cell carcinoma as well as primary salivary gland malignancies. Comprehensive methodological analysis identified poor reporting of detailed methodology, variations in collection, storage, pre-processing, RNA isolation and relative quantification including normalisation method. We recommend standardising the pre-processing, RNA isolation, normalisation and relative quantitation steps at biomarker discovery phase. Such standardisation would allow for bias minimisation and effective progression into clinical validation phases.

Promoter hypermethylation-mediated downregulation of tumor suppressor gene SEMA3B and lncRNA SEMA3B-AS1 correlates with progression and prognosis of esophageal squamous cell carcinoma Abstract Frequent deletions of tumor-suppressor genes at chromosome 3p21.3 have been detected in esophageal squamous cell carcinoma (ESCC). As a candidate tumor suppressor gene, semaphorin 3B (SEMA3B) is located at 3p21.3 and is frequently inactivated in several tumors. However, the role and inactivation mechanisms of SEMA3B and its antisense long non-coding RNA (lncRNA) SEMA3B-AS1 in the carcinogenesis of ESCC have not been fully elucidated. The present study was conducted to investigate the role, epigenetic inactivation mechanisms, and prognostic value of SEMA3B and SEMA3B-AS1 in ESCC tumorigenesis and prognosis. Frequent downregulation of SEMA3B and SEMA3B-AS1 was detected in esophageal cancer cells and ESCC tissues, and the expression level of SEMA3B and SEMA3B-AS1 in ESCC tissues was correlated with TNM stage and lymph node metastasis. SEMA3B and SEMA3B-AS1 shared the same CpG island in the promoter region and the expression of both genes might be regulated by the promoter methylation status. Furthermore, transcription factor Sp1 activated SEMA3B or SEMA3B-AS1 transcription and the promoter hypermethylation of SEMA3B and SEMA3B-AS1 influenced Sp1 binding ability. Moreover, over-expression of SEMA3B and SEMA3B-AS1 suppressed the viability and invasion of esophageal cancer cells in vitro. SEMA3B-AS1 influenced the protein expression of SEMA3B. SEMA3B or SEMA3B-AS1 expression and promoter methylation status were correlated with ESCC patients' survival. Thus, these findings suggest that SEMA3B and SEMA3B-AS1 may act as tumor suppressors and may serve as potential targets for antitumor therapy.

Identification of canonical NFκB (C-NFκB) pathway in uveal melanoma and their relation with patient outcome Abstract Inflammation in uveal melanoma (UM) is linked to a bad prognosis. It is rare type of cancer, of which the metastases are usually fatal within a year. Infiltration with an inflammatory infiltrate increases with disease progression but does not seem to inhibit metastasis. The Canonical NFκB (C-NFκB) pathway is known to play a crucial role in tumor inflammation. We therefore, studied the expression of canonical NFκB proteins and their prognostic relevance in UM. Our study evaluated the expression of C-NFκB proteins (p65, p50, and c-Rel) by using immunohistochemistry on sections from 75 formalin-fixed UM. Activation of the NFκB subunit was determined on fresh tumor specimens by measuring the DNA-binding activity in nuclei using an NFκB ELISA assay. Real-time PCR was performed on frozen material on 58 tumors. The presence of native C-NFκB heterodimers (p65/p50 and c-Rel/p50) was confirmed by co-immunoprecipitation followed by Western blotting. We observed a high nuclear immunoreactivity of p65, p50, and c-Rel proteins in 54, 60 and 41% UM cases, respectively. Expression of C-NFκB proteins significantly correlated with parameters which are related to the inflammatory environment of UM. Nuclear immunoreactivity of p65 and p50 was associated with lower patient survival (p = 0.041; p = 0.048) while c-Rel was not. Our finding reveals that C-NFκB proteins expressed are more often in UM with inflammation than those without inflammation. Activation of the canonical NFκB pathway is more frequent in high risk UM patients. These observations might help to understand the behaviour of high risk tumors, with upregulation of C-NFκB proteins contributing to tumor aggressiveness.

Non-canonical roles for metabolic enzymes and intermediates in malignant progression and metastasis Abstract Metabolic alterations are established as a hallmark of cancer. Such hallmark changes in cancer metabolism are characterized by reprogramming of energy-producing pathways and increases in the generation of biosynthetic intermediates to meet the needs of rapidly proliferating tumor cells. Various metabolic phenotypes such as aerobic glycolysis, increased glutamine consumption, and lipolysis have also been associated with the process of metastasis. However, in addition to the energy and biosynthetic alterations, a number of secondary functions of enzymes and metabolites are emerging that specifically contribute to metastasis. Here, we describe atypical intracellular roles of metabolic enzymes, extracellular functions of metabolic enzymes, roles of metabolites as signaling molecules, and epigenetic regulation mediated by altered metabolism, all of which can affect metastatic progression. We highlight how some of these mechanisms are already being exploited for therapeutic purposes, and discuss how others show similar potential.

Discovery of CCL18 antagonist blocking breast cancer metastasis Abstract Our previous studies have proved that CCL18 is the most secreted chemokine in breast cancer microenvironment by tumor associated macrophages (TAMs). CCL18 promotes breast cancer invasiveness by binding to its cognate receptor PITPNM3 and activating the downstream signaling pathways. The high level of CCL18 in serum or tumor stroma is associated with tumor metastasis and poor patients overall survival. In this study, we identify an effective small molecular compound (SMC) to antagonize the effect of CCL18. We screen more than 1000 SMCs from Sun Yat-sen University SMC library and select 15 top scored SMCs by using computer-aided virtual screening based on the structure of CCL18. Then in vitro cell migration assay narrows down the selected 15 SMCs to the most effective SMC-21598. We find 10 µM SMC-21598 significantly inhibits CCL18-induced breast cancer cells adherence, invasiveness, and migration. Our further surface plasmon resonance (SPR), fluorescence spectroscopy and isothermal titration calorimetry (ITC) assays reveal that SMC-21598 binds tightly to CCL18, which blocks the binding of CCL18 with its receptor PITPNM3. The in vivo animal experiments show that SMC-21598 doesn't significantly affect xenografts growth, but inhibits lung metastasis. Our study provides a potential lead compound to antagonize CCL18 function. It would be of great significance to develop SMC drugs to ameliorate breast cancer metastasis and prolong patients' survival.

Histopathological growth patterns of colorectal liver metastasis exhibit little heterogeneity and can be determined with a high diagnostic accuracy Abstract Colorectal liver metastases (CRLM) exhibit distinct histopathological growth patterns (HGPs) that are indicative of prognosis following surgical treatment. This study aims to assess the reliability and replicability of this histological biomarker. Within and between metastasis HGP concordance was analysed in patients who underwent surgery for CRLM. An independent cohort was used for external validation. Within metastasis concordance was assessed in CRLM with ≥ 2 tissue blocks. Similarly, concordance amongst multiple metastases was determined in patients with ≥ 2 resected CRLM. Diagnostic accuracy [expressed in area under the curve (AUC)] was compared by number of blocks and number of metastases scored. Interobserver agreement (Cohen's k) compared to the gold standard was determined for a pathologist and a PhD candidate without experience in HGP assessment after one and two training sessions. Both the within (95%, n = 825) and the between metastasis (90%, n = 363) HGP concordance was high. These results could be replicated in the external validation cohort with a within and between metastasis concordance of 97% and 94%, respectively. Diagnostic accuracy improved when scoring 2 versus 1 blocks(s) or CRLM (AUC = 95.9 vs. 97.7 [p = 0.039] and AUC = 96.5 vs. 93.3 [p = 0.026], respectively), but not when scoring 3 versus 2 blocks or CRLM (both p > 0.2). After two training sessions the interobserver agreement for both the pathologist and the PhD candidate were excellent (k = 0.953 and k = 0.951, respectively). The histopathological growth patterns of colorectal liver metastasis exhibit little heterogeneity and can be determined with a high diagnostic accuracy, making them a reliable and replicable histological biomarker.

Metastasis is impaired by endothelial-specific Dll4 loss-of-function through inhibition of epithelial-to-mesenchymal transition and reduction of cancer stem cells and circulating tumor cells Abstract Systemic inhibition of Dll4 has been shown to thoroughly reduce cancer metastasis. The exact cause of this effect and whether it is endothelial mediated remains to be clarified. Therefore, we proposed to analyze the impact of endothelial Dll4 loss-of-function on metastasis induction on three early steps of the metastatic process, regulation of epithelial-to-mesenchymal transition (EMT), cancer stem cell (CSC) frequency and circulating tumor cell (CTC) number. For this, Lewis Lung Carcinoma (LLC) cells were used to model mouse tumor metastasis in vivo, by subcutaneous transplantation into endothelial-specific Dll4 loss-of-function mice. We observed that endothelial-specific Dll4 loss-of-function is responsible for the tumor vascular regression that leads to the reduction of tumor burden. It induces an increase in tumoral blood vessel density, but the neovessels are poorly perfused, with increased leakage and reduced perivascular maturation. Unexpectedly, although hypoxia was increased in the tumor, the number and burden of macro-metastasis was significantly reduced. This is likely to be a consequence of the observed reduction in both EMT and CSC numbers caused by the endothelial-specific Dll4 loss-of-function. This multifactorial context may explain the concomitantly observed reduction of the circulating tumor cell count. Furthermore, our results suggest that endothelial Dll4/Notch-function mediates tumor hypoxia-driven increase of EMT. Therefore, it appears that endothelial Dll4 may constitute a promising target to prevent metastasis.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Cancer Microenvironment

Immunosuppressive Tumor Microenvironment Status and Histological Grading of Endometrial Carcinoma Abstract The recent successes of new cancer immunotherapy approaches have led to investigate their relevance in the context of the Endometrial Carcinoma (EC). These therapies, that take the tumor-induced immunosuppressive microenvironment into account, target the tumor immune escape, in particular the inhibitory receptors involved in the regulation of the effector T cells' activity (immune checkpoints). The aim of this study was to identify, in ECs, differences in intergrades immune status that could contribute to the differences in tumor aggressiveness, and could also be used as theranostic tools. The immune status of tumors was assessed by quantitative real-time PCR. We analyzed the expression of specific genes associated to specific leukocytes subpopulations and the expression of reporting genes associated with the tumor escape/resistance. This study highlights significant differences in the EC intergrades immune status especially the tumor-infiltrating cell types and their activation status as well as in the molecular factors produced by the environment. The immune microenvironment of grade 1 ECs hints at a robust tumoricidal milieu while that of higher grades is more evocative of a tolerogenic milieu. This genes-based immunological monitoring of tumors that easily highlights significant intergrade differences relating to the density, composition and functional state of the leukocyte infiltrate, could give solid arguments for choosing the best therapeutic options, especially those targeting immune checkpoints. Moreover it could enable an easy adaptation of individual treatment approaches for each patient.

Editorial Comment: Deciphering the "Metastasome" Leads to Novel Hypotheses on Understanding the Evolution of Metastasis and Implicates Consequences for Metastasis Therapy and Prevention

Targeting Hyaluronan Interactions for Glioblastoma Stem Cell Therapy Abstract Even with rigorous treatments, glioblastoma multiforme (GBM) has an abysmal median survival rate, greatly due to the drug-resistant glioblastoma stem cell (GSC) population. GSCs are known to remodel their microenvironment, but the precise role of extracellular matrix components hyaluronic acid (HA) and hyaluronidases (HAases) on the GSC population is still largely unknown. Our objective was to determine how HAase can sensitize GSCs to chemotherapy drugs by disrupting the HA-CD44 signaling. GBM cell line U87-MG and patient-derived D456 cells were grown in GSC-enriching media and treated with HA or HAase. Expressions of GSC markers, HA-related genes, and drug resistance genes were measured via flow cytometry, confocal microscopy, and qRT-PCR. Proliferation after combined HAase and temozolomide (TMZ) treatment was measured via WST-8. HA supplementation promoted the expression of GSC markers and CD44 in GBM cells cultured in serum-free media. Conversely, HAase addition inhibited GSC gene expression while promoting CD44 expression. Finally, HAase sensitized GBM cells to TMZ. We propose a combined treatment of HAase and chemotherapy drugs by disrupting the stemness-promoting HA to target GSCs. This combination therapy shows promise even when temozolomide treatment alone causes resistance.

Peripheral Blood Cell Interactions of Cancer-Derived Exosomes Affect Immune Function Abstract Cancer-derived exosomes are constitutively produced and secreted into the blood and biofluids of their host patients providing a liquid biopsy for early detection and diagnosis. Given their ubiquitous nature, cancer exosomes influence biological mechanisms that are beneficial to the tumor cells where they are produced and the microenvironment in which these tumors exist. Accumulating evidence suggests that exosomes transport proteins, lipids, DNA, mRNA, miRNA and long non coding RNA (lncRNA) for the purpose of cell-cell and cell-extracellular communication. These exosomes consistently reflect the status as well as identity of their cell of origin and as such may conceivably be affecting the ability of a functional immune system to recognize and eliminate cancer cells. Recognizing and mapping the pathways in which immune suppression is garnered through these tumor derived exosome (TEX) may lead to treatment strategies in which specific cell membrane proteins or receptors may be targeted, allowing for immune surveillance to once again help with the treatment of cancer. This Review focuses on how cancer exosomes interact with immune cells in the blood.

The Tumor Microenvironment in Post-Transplant Lymphoproliferative Disorders Abstract Post-transplant lymphoproliferative disorders (PTLDs) cover a broad spectrum of lymphoproliferative lesions arising after solid organ or allogeneic hematopoietic stem cell transplantation. The composition and function of the tumor microenvironment (TME), consisting of all non-malignant constituents of a tumor, is greatly impacted in PTLD through a complex interplay between 4 factors: 1) the graft organ causes immune stimulation through chronic antigen presentation; 2) the therapy to prevent organ rejection interferes with the immune system; 3) the oncogenic Epstein-Barr virus (EBV), present in 80% of PTLDs, has a causative role in the oncogenic transformation of lymphocytes and influences immune responses; 4) interaction with the donor-derived immune cells accompanying the graft. These factors make PTLDs an interesting model to look at cancer-microenvironment interactions and current findings can be of interest for other malignancies including solid tumors. Here we will review the current knowledge of the TME composition in PTLD with a focus on the different factors involved in PTLD development.

Mechanobiology of Cancer Stem Cells and Their Niche Abstract Though the existence of cancer stem cells remained enigmatic initially, over the time their participation in tumorigenesis and tumor progression has become highly evident. Today, they are also appreciated as the causal element for tumor heterogeneity and drug-resistance. Cancer stem cells activate a set of molecular pathways some of which are triggered by the unique mechanical properties of the tumor tissue stroma. A relatively new field called mechanobiology has emerged, which aims to critically evaluate the mechanical properties associated with biological events like tissue morphogenesis, cell-cell or cell-matrix interactions, cellular migration and also the development and progression of cancer. Development of more realistic model systems and biophysical instrumentation for observation and manipulation of cell-dynamics in real-time has invoked a hope for some novel therapeutic modalities against cancer in the future. This review discusses the fundamental concepts of cancer stem cells from an intriguing viewpoint of mechanobiology and some important breakthroughs to date.

Altered T Cell Migratory Capacity in the Progression from Barrett Oesophagus to Oesophageal Adenocarcinoma Abstract Oesophageal adenocarcinoma (OAC) is an inflammation-driven cancer with poor prognosis and incidence is increasing rapidly. OAC arises from gastro-oesophageal reflux disease (GORD) and reflux-induced Barrett oesophagus (BO). The role of T cells in this disease progression is not yet fully understood. We have previously demonstrated higher proportions of pro-tumour Th2 cells in BO tissue, implicating them in its pathogenesis. While a Th2 immune profile is thought to underlie the metaplastic transformation in BO and promote OAC development, our studies suggest that the abundance of Th2 cells in BO tissue is likely to occur through altered T cell recruitment. This study examined the chemokine networks governing T cell migration to oesophageal tissue during disease progression. Here, we have identified that circulating T cells in OAC patients, exhibit impaired migratory capacity with decreased frequencies of Th1-associated CXCR3+ and Th17-associated CCR6+ cells. Despite the abundance of Th1 chemokines RANTES (CCL5) and MIP-1α (CCL3) in OAC tumour, enrichments of intratumoural T cells expressing corresponding receptors were not observed. These data suggest that T cell infiltration of oesophageal tissue is compromised in OAC and suggest that future therapies targeting T cell trafficking should occur at the pre-neoplastic stage. This is supported by the finding that antagonism of Th2-biased CCR4 significantly reduces T cell migration in BO but not OAC patients. Since we have previously reported a predominant Th2 immune profile in BO, we suggest that chemokine receptor antagonism may be a viable treatment option to alleviate Th2-predominance in BO and interrupt progression to OAC.

Cancer-Associated Fibroblasts Enhance Survival and Progression of the Aggressive Pancreatic Tumor Via FGF-2 and CXCL8 Abstract Pancreatic ductal adenocarcinoma remains one of the most challenging human cancers. Desmoplasia is predominant in this disease exhibiting a strong stromal reaction with an abundance of the cancer-associated fibroblasts (CAFs). We aimed in this study to investigate the reciprocal interaction between the tumor cells and the CAFs and its effect on tumor cells survival. We hypothesized that the survival of pancreatic cancer cell with aggressive phenotype is modulated by the Interactions between malignant pancreatic tumor cells and surrounding CAFs. To examine this, we utilized co-culture methods where tumor cells with different malignant potentials, HPAF (low) HPAF-CD11 (moderate/high) co-cultured with CAFs. CAFs-conditioned media increased the growth of HPAF-CD11 but not HPAF cells and increased CXCL8 levels highly in HPAF-CD11 and slightly in HPAF. The growth stimulatory effect and elevated CXCL8 level caused by CAFs-conditioned media were diminished by neutralizing the fibroblast growth factor-2 (FGF-2). In addition, conditioned media of HPAF-CD11 increased CAFs cell number whereas that of HPAF did not, and these effects were suppressed by neutralizing CXCL8. Furthermore, data from gene expression microarray study exhibited different expression profiles between HPAF and HPAF-CD11 when co-culture with CAFs. A significant increase in CXCL8 and FGF-2 expression was observed with HPAF-CD11/CAFs co-culture and to a lower extent with HPAF/CAFs co-culture. Together, these data demonstrate a paracrine bi-directional interaction between pancreatic tumor cells and the CAFs through CXCL8 and FGF-2 that helps the tumor growth. Future in-depth study of these pathways will assist in obtaining diagnostic and therapeutic tools for pancreatic ductal adenocarcinoma.

Tumor-Associated T-Lymphocytes and Macrophages are Decreased in Endometrioid Endometrial Carcinoma with MELF-Pattern Stromal Changes Abstract Microcystic, elongated, fragmented (MELF)-pattern is an unusual morphology of myometrial invasive front in endometrioid endometrial carcinoma (EA). The aim of the study was to investigate potential correlation between MELF-pattern and peritumoral inflammatory immune response. A total of 96 out of 368 patients were included in this study. CD3, CD20, CD57. CD68 and S100 markers were used for the detection of tumor-associated T-lymphocytes (TAT), tumor-associated B-lymphocytes (TAB), tumor-associated NK-lymphocytes (NK), tumor-associated macrophages and dendritic cells respectively. Mann-Whitney tests, receiver operating characteristic (ROC) curve analysis, and Spearman correlation were used as methods for statistical analyses. Odds ratio with 95% confidence interval (95% CI) was determined with the use of a logistic regression model. A p < 0.05 was considered statistically significant. Our results suggested that the number of CD3 and CD68 cells were significantly lower (p < 0.001) in cases of endometrioid carcinoma with MELF-pattern. A significant correlation between the presence of MELF-pattern and decrease of CD3 positive T-lymphocytes (r = 0.691; p < 0.001) was also observed. Additionally, we found an inverse correlation between the presence of MELF-pattern and TAM (r = 0.568; p = 0.001). Therefore, our data suggest that MELF-pattern may be associated with EA stroma fibrosis that contains immune cells infiltration and demonstrated a decrease in the number of TAT and TAM cells. This may indicate the poor clinical prognosis of this disease.

T-Cell Clustering in Neoplastic Follicles of Follicular Lymphoma Abstract The nonneoplastic microenvironment is abundant in follicular lymphoma. Its composition has been reported to be associated with the course of the disease. Lack of animal models hampers studies of interaction between lymphoma and bystander cells. We aimed to identify indicators of cellular interaction exemplified by nonrandom distribution of cell types within neoplastic follicles. Physiological germinal centers and follicles in follicular lymphoma were stained to identify macrophages, all T, follicular T-helper, dendritic and B cells. Density of cell types and cell distribution (spatial point pattern) were analyzed by digital image analysis. The density of all T, follicular T-helper and dendritic cells was higher in the dark zone than in the light zone of physiological germinal centers. Densities of cell types in follicular lymphoma were intermediate between the light and the dark zone. All cell types analyzed showed a completely random spatial distribution pattern within the dark and the light zone, respectively. In follicular lymphoma B cells and macrophages displayed complete spatial randomness. In contrast, all T cells, follicular T-helper cells and dendritic cells showed clustering of each individual cell type within a radius of 6–10 μm in the lymphoma. We conclude that the distribution of nonneoplastic cells within follicles of follicular lymphoma is not random. T cells and dendritic cells form clusters within the follicles, suggestive of sites of interaction between microenvironment and lymphoma cells. These clusters might help to understand the interaction of lymphoma cells with the microenvironment and might provide a structure for therapeutic intervention.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Folia Microbiologica

Infection caused by Klebsiella pneumoniae ST11 in a patient after craniectomy Abstract Klebsiella pneumoniae infections have always been an important problem in public health, but today, the increasing resistance of these bacteria to antibiotics due to β-lactamases production has renewed interest in K. pneumoniae infections. The aim of the study was to present a case of a neurosurgical patient with multidrug-resistant K. pneumoniae ST11 infection after craniectomy. Four K. pneumoniae isolates from various clinical materials of the patient undergone identification and susceptibility testing with the Vitek2 system. Tests for β-lactamases production were performed according to EUCAST guidelines. Strains were analyzed for bla genes responsible for β-lactamase production (blaTEM, blaSHV, blaCTX-M, blaVIM, blaIMP, blaNDM, blaKPC, blaOXA-48) using PCR. Moreover, the genetic relatedness of these isolates was determined by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). All tested strain presented multidrug resistance. The highest susceptibility was observed for imipenem, meropenem, and ertapenem. The strain isolated from the nervous system was ESBL-positive with blaSHV-11, blaTEM-1, and blaCTX-M-15 genes. Additionally, the strain from urine was blaKPC-3-positive. Molecular typing revealed that all strains belonged to the same clone and identified two PFGE profiles. The analysis of MLST allelic profile showed that tested K. pneumoniae strains belonged to ST11. Identification of ST11 K. pneumoniae as etiological factor of infection unfavorably impacts on prognosis among neurosurgical patient after craniectomy.

Effect of silver nanoparticles conjugated to thiosemicarbazide on biofilm formation and expression of intercellular adhesion molecule genes, icaAD , in Staphylococcus aureus Abstract Biofilm formation is regarded as an important factor in the establishment of infections caused by Staphylococcus aureus. In the present study, phenotypic and molecular assays were used to evaluate antibiofilm potential of thiosemicarbazide (Tsc) conjugated with silver nanoparticles (Ag NPs) and functionalized by glutamic acid (Ag@Glu/Tsc NPs) against methicillin-resistant S. aureus (MRSA). Ag NPs were synthesized using precipitation method and conjugated to Tsc using glutamic acid. The NPs were characterized using SEM and FTIR spectroscopy analyses. Then, antibiofilm potential of the prepared NPs against MRSA strains was evaluated using phenotypic method and their effects on the expression of biofilm-associated genes icaA and icaD. Finally, the genes involved with the synthesis of intercellular adhesion molecules were determined. According to the results, Ag@Glu/Tsc NPs inhibited biofilm formation of MRSA strains up to 76.7% compared with the control. In addition, expression of the biofilm-associated genes icaA and icaD reduced by 66.7% and 60.3%, respectively in the presence of sub-inhibitory concentration of Ag@Glu/Tsc NPs. In conclusion, Ag@Glu/Tsc NPs could be considered as a potent antibacterial agent to inhibit bacterial biofilms.

Comparative evaluation of structure and characteristic of peptidyl-prolyl cis-trans isomerase proteins and their function in Salmonella Typhimurium stress responses and virulence Abstract Peptidyl-prolyl cis-trans isomerases (PPIase) exhibit chaperone activity and assist in protein folding by increasing the rate of cis-trans transition on proline-peptide bonds. The current study aimed to identify and characterize three genes, ppiA, ppiB, and ppiC, which encode proteins of the PPIase family in the bacterium Salmonella enterica serovar Typhimurium. Salmonella Typhimurium is a facultative intracellular zoonotic pathogen that causes food- and water-borne gastroenteritis in humans (leading to bacteremia in immune-compromised subjects). Recombinant clones for the three genes were constructed and sequenced and the sequences submitted to NCBI GenBank. Three-dimensional structures for the corresponding proteins were predicted by comparative modeling. A maximum-likelihood phylogenetic gene tree constructed for the three genes showed a low evolutionary mean diversity, indicating strong evolutionary conservation. Further, single-gene deletion mutant strains, generated for the respective genes, were observed to be more susceptible to the stationary phase of growth and heat stress conditions and showed reduced survival within macrophage cells line. The present study thus indicates that ppiA, ppiB, and ppiC genes are conserved among Salmonella genome, are critical for the growth of Salmonella Typhimurium in the examined stress conditions, and may play a role in its responses and virulence.

Analysis of inflammatory cytokine expression in the urinary tract of BALB/c mice infected with Proteus ( P .) mirabilis and enteroaggregative Escherichia ( E .) coli (EAEC) strains Abstract This study aimed to analyze the proinflammatory cytokine mRNA expression in the urinary tract of BALB/c mice infected with bacterial strains with uropathogenic potential. Groups of four 6-week-old female BALB/c mice were intraurethrally inoculated with 5 × 107 colony-forming units (CFU) of P. mirabilis ATCC29906, EAEC O42, P. mirabilis RTX339, or sterile saline (control group) and then sacrificed at 0, 2, 4, 7, or 10 days post-infection (p.i.). Samples were cultured to determine the CFU/mL in urine or CFU/g in the bladders and kidneys. Cytokine expression (tumor necrosis factor (TNF)-α and interleukin (IL)-1β, -6, and -8) was evaluated in the target organs using real-time PCR and immunohistochemistry; histology was examined with hematoxylin and eosin staining. The results are presented as the means and standard deviations and were compared using one-way ANOVA, with p < 0.05 indicating significant differences. Bacteriuria was not detected in the infected groups; bacterial colonization occurred in the target organs at all time points, but was higher in mice infected with EAEC O42 or P. mirabilis RTX339 at 7 days p.i. The expression of all cytokine mRNAs was seen, but only the levels of the IL-8 protein increased in situ at 7 days p.i. in the P. mirabilis RTX339 and EAEC O42 groups in both organs. Morphological alterations, observed in all of the infected groups, were more prominent in the EAEC O42 and P. mirabilis RTX339 groups. The findings provide insights into the uropathogenicity and inflammatory cytokine expression in the urinary tract of mice infected with three previously untested bacterial strains.

Cellulolytic thermophilic microorganisms in white biotechnology: a review Abstract Enzymes of microbial origin are of immense importance for organic material decomposition leading to bioremediation of organic waste, bioenergy generation, large-scale industrial bioprocesses, etc. The market demand for microbial cellulase enzyme is growing more rapidly which ultimately becomes the driving force towards research on this biocatalyst, widely used in various industrial activities. The use of novel cellulase genes obtained from various thermophiles through metagenomics and genetic engineering as well as following metabolic engineering pathways would be able to enhance the production of thermophilic cellulase at industrial scale. The present review is mainly focused on thermophilic cellulolytic bacteria, discoveries on cellulase gene, genetically modified cellulase, metabolic engineering, and their various industrial applications. A lot of lacunae are yet to overcome for thermophiles such as metagenome analysis, metabolic pathway modification study, search of heterologous hosts in gene expression system, and improved recombinant strain for better cellulase yield as well as value-added product formation.

Phospholipid biosynthesis disruption renders the yeast cells sensitive to antifungals Abstract To understand the role of phospholipids on Cdr1p (drug exporter)-mediated drug resistance in yeast, the phospholipids biosynthesis genes PSD1, PSD2, CHO2, and OPI3 were deleted in a strain of Saccharomyces cerevisiae already overexpressing Cdr1-GFP of Candida albicans as a heterologous system. The effect of phospholipids biosynthesis gene deletion was analyzed on Cdr1p-GFP-mediated drug resistance as well as its localization. The results indicate that phospholipids biosynthesis disruption makes the cell sensitive to several drugs including fluconazole (FLC), with Δpsd1/Cdr1-GFP being worst affected. Interestingly, unlike sterols and sphingolipids, the localization of Cdr1p was unaffected by phospholipid biosynthesis gene disruption. Concomitantly, phospholipids mutants also showed an increase in reactive oxygen species (ROS) generation, as verified by fluorescence probe 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA) method. In addition, the sensitivity of phospholipid mutants with FLC was found to be synergistic to ROS generation, resulting in further reduction of growth. Thus, this study proposes phospholipid biosynthesis as a novel target for antifungal therapy.

BAL fluid analysis in the identification of infectious agents in patients with hematological malignancies and pulmonary infiltrates Abstract The present study aims to evaluate the diagnostic yield of bronchoalveolar lavage (BAL) fluid in patients with hematological malignancies and describe the most common pathogens detected in BAL fluid (BALF.) An analysis of 480 BALF samples was performed in patients with hematological malignancies over a period of 7 years. The results of culture methods, PCR, and immunoenzymatic sandwich microplate assays for Aspergillusgalactomannan (GM) in BALF were analyzed. Further, the diagnostic thresholds for Aspergillus GM and Pneumocystis jiroveci were also calculated. Microbiological findings were present in 87% of BALF samples. Possible infectious pathogens were detected in 55% of cases; 32% were classified as colonizing. No significant difference in diagnostic yield or pathogen spectrum was found between non-neutropenic and neutropenic patients. There was one significant difference in BALF findings among intensive care units (ICU) versus non-ICU patients for Aspergillus spp. (22% versus 9%, p = 0.03). The most common pathogens were Aspergillus spp. (n = 86, 33% of BAL with causative pathogens) and Streptococcus pneumoniae (n = 46, 18%); polymicrobial etiology was documented in 20% of cases. A quantitative PCR value of > 1860 cp/mL for Pneumocystis jirovecii was set as a diagnostic threshold for pneumocystis pneumonia. The absorbance index of GM in BALF of 0.5 was set as a diagnostic threshold for aspergillosis. The examination of BAL fluid revealed the presence of pathogen in more than 50% of cases and is, therefore, highly useful in this regard when concerning pulmonary infiltrates.

Glutamine synthetase type I ( gln AI) represents a rewarding molecular marker in the classification of bifidobacteria and related genera Abstract The family Bifidobacteriaceae constitutes an important phylogenetic group that particularly includes bifidobacterial taxa demonstrating proven or debated positive effects on host health. The increasingly widespread application of probiotic cultures in the twenty-first century requires detailed classification to the level of particular strains. This study aimed to apply the glutamine synthetase class I (glnAI) gene region (717 bp representing approximately 50% of the entire gene sequence) using specific PCR primers for the classification, typing, and phylogenetic analysis of bifidobacteria and closely related scardovial genera. In the family Bifidobacteriaceae, this is the first report on the use of this gene for such purposes. To achieve high-value results, almost all valid Bifidobacteriaceae type strains (75) and 15 strains isolated from various environments were evaluated. The threshold value of the glnAI gene identity among Bifidobacterium species (86.9%) was comparable to that of other phylogenetic/identification markers proposed for bifidobacteria and was much lower compared to the 16S rRNA gene. Further statistical and phylogenetic analyses suggest that the glnAI gene can be applied as a novel genetic marker in the classification, genotyping, and phylogenetic analysis of isolates belonging to the family Bifidobacteriaceae.

Development of a sequence-characterized amplified region marker for detection of Ascochyta rabiei causing Ascochyta blight in chickpea Abstract Ascochyta blight of chickpea is caused by Ascochyta rabiei (Pass.) Labr. which is primarily seedborne. For rapid detection and precise identification of A. rabiei, a sequence-characterized amplified region (SCAR) marker was developed for detection of genomic DNA and infected plant DNA. An SSR primer amplified monomorphic band was cloned in pGEM®-T easy vector and sequenced. The best primer pair was selected and validated on A. rabiei. The specificity and sensitivity of the SCAR-based marker designated as MBAR was evaluated using conventional PCR and real-time PCR. The marker produced consistently an amplicon size of 196 bp in all A. rabiei isolates tested. The sensitivity of the marker was 0.1 ng of genomic fungal DNA and 0.5 ng of plant DNA by conventional PCR and 0.5 pg of A. rabiei DNA and 1.0 pg of plant DNA by real-time PCR. This is the first SCAR marker having high specificity and sensitivity towards A. rabiei. The marker may be useful in detecting the pathogen before the disease appearance and in plant quarantine program to detect the pathogen in seed lots.

Recommendations for cerebrospinal fluid analysis Abstract Diseases of the central nervous system (CNS) mean for the human organism a potentially dangerous situation. An investigation of cerebrospinal fluid (CSF) provides important information about a character of CNS impairment in the decision-making diagnostic and therapeutic algorithm. The authors present a brief overview of available cerebrospinal fluid assays, shortened indication criteria, a recommended algorithm of CSF assessment in different suspected diseases, and a view of the external quality system. The whole portfolio of obtainable CSF methodology is further subdivided according to the adequate choice into the first and inevitable basic routine panel, and following complicated analyses of highly specialized character. The basic panel is considered for standard laboratories, the complete specialized assessment should be provided by a super-consulting laboratory.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com

Food Analytical Methods

Quantitation of Selenomethionine in Multivitamins and Selenium Supplements by High Performance Liquid Chromatography Inductively-Coupled Plasma Mass Spectrometry Abstract VITA-1 and VITB-1 multivitamin and mineral supplement candidate reference materials from the National Research Council Canada were analyzed for their total selenium and selenomethionine contents. Following a methanesulfonic acid reflux to extract selenomethionine from the selenized yeast in the multivitamins, analysis by high-performance liquid chromatography-inductively coupled plasma triple quadrupole mass spectrometry resulted in concentrations of 7.4 ± 3.0 μg SeMet g–1 and 16.4 ± 6.5 μg SeMet g−1 in VITA-1 and VITB-1, respectively. Twelve commercially available multivitamins and selenium supplements were analyzed following the same protocol. Seven of these were noted to contain 85–115% of the selenium stated on the label; the others ranged from 5 to 147% of the claimed amount. Only one multivitamin contained selenomethionine at a concentration above the detection limit, but the amount found in the selenium supplements matched the label claims within a reasonable level of uncertainty. For comparison, two certified reference materials—wheat gluten and egg powder, both certified for total selenium only—were also examined and it was determined that selenomethionine accounted for 58% and 25% of the selenium in these food products, respectively.

Development of a Sensitive Monoclonal Antibody–Based Indirect Competitive Enzyme-Linked Immunosorbent Assay for the Determination of Monensin in Edible Chicken Tissues Abstract To monitor monensin (MON) residues in chicken tissues, a monoclonal antibody (mAb)–based indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) was developed in this study. The haptenMON was obtained by adding hydrochloric acid to a MON salt, and then was coupled with carrier protein. After the inoculation of female Balb/c mice and cell fusions, one cell line, MON/4C9, was obtained. The MON/4C9 antibody exhibited the ability to specifically recognize MON with IC50 1.28 μg L−1.Based on this mAb, an optimized ic-ELISA protocol was performed using only methanol-water (8:2, v/v) in chicken tissue samples. The limits of detection and the limits of quantification of MON in various sample matrices varied from 0.38 to 1.01 μg kg−1. The recoveries ranged from 71.9 to 116.9% in chicken tissues, and the intra- and inter-assay CVs were all less than 18%. Moreover, the developed method also exhibited a positive correlation with the results of HPLC-MS conducted on the samples. These results suggest that the prepared mAb and the developed ic-ELISA method will be a useful tool for detecting MON in chicken tissues.

Aflatoxin B 1 Determination in Peanuts by Magnetic Nanoparticle–Based Immunofluorescence Assay Abstract A sensitive immunofluorescence assay for rapid AFB1 determination was developed. Polyclonal anti-AFB1 antibody was immobilized onto magnetic nanoparticles (MNPs). The optimal amount of immobilized antibody was found—40 μg antibody per 1 mg MNPs. The conjugate of AFB1 with bovine serum albumin (BSA) and fluorescein isothiocyanate was prepared. UV-Vis and fluorescence spectrophotometric analyses of the obtained conjugate proved the successful conjugate formation. Optimal amount of MNPs with immobilized antibody (0.25 mg) and the optimal concentration of fluorescent conjugate of AFB1 (28 μg/mL) were established. A competitive assay of AFB1 was performed. The toxin solubility and stability were optimal for the assay when using buffer solution with 10% methanol. It was found that the sensitivity of the developed AFB1 assay was significantly higher with 0.1% BSA stabilizing AFB1 solutions. The linear range of the proposed method was 2–100 pg/mL. The immunoassay was applied for determination of AFB1 in spiked peanuts.

Simultaneous Determination of Isofetamid and Its Two Metabolites in Fruits and Vegetables Using Ultra-Performance Liquid Chromatography with Tandem Mass Spectrometry Abstract In this paper, an efficient analytical method for the simultaneous determination of a new fungicide (isofetamid) and its two metabolites in fruits and vegetables was developed using the QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) pretreatment method and ultra-performance liquid chromatography-tandem mass spectrometry. The three compounds were extracted with acetonitrile and cleaned up using primary secondary amine, octadecylsilane, and graphitized carbon black. The three analytes were well separated with good specificity using an electrospray ionization source in positive mode within 5.0 min. The mean recoveries in apple, strawberry, grape, tomato, cucumber, and lettuce for the three target compounds at different levels ranged from 74 to 107%, with relative standard deviations lower than 12.3%. The method showed excellent linearity (R2 ≥ 0.9946) for the three compounds. The limits of quantification of isofetamid and its metabolites were 1 μg kg−1 in apple, tomato, and cucumber and 100 μg kg−1 in strawberry, grape, and lettuce. The proposed method was successfully applied to a residue dynamic experiment, and it can be considered a strong alternative method to monitor the residues of isofetamid and its metabolites simultaneously in fruits and vegetables.

Fast Sequential Determination of Zn, Fe, Mg, Ca, Na, and K in Infant Formulas by High-Resolution Continuum Source Flame Atomic Absorption Spectrometry Using Ultrasound-Assisted Extraction Abstract In this work, an analytical procedure based on ultrasound-assisted extraction (UAE) was developed for the fast sequential determination of six elements (Zn, Fe, Mg, Ca, Na, and K) in infant formulas using high-resolution continuum source flame atomic absorption spectrometry (HR-CS F AAS). The analytes extraction was performed using HNO3 2.1 mol L−1 and 10 min of sonication time at 80 °C. The limits of detection obtained were 12, 13, 0.48, 96, 137, and 14 mg kg−1 for Zn, Fe, Mg, Ca, Na, and K, respectively. The analysis of a certified reference material of non-fat milk powder (SRM 1549) and infant formula samples after microwave digestion confirmed the trueness of the developed method. Thus, the UAE procedure was employed in order to determine these six elements and their concentration in six infant formulas and their content (in mg kg−1) ranged between 64 and 431 for Zn, 64–370 for Fe, 420–2421 for Mg, 1145–8547 for Na, 750–14,491 for K, and 2827–11,340 for Ca.

Front-Face Fluorescence Combined with Second-Order Multiway Classification, Based on Polyphenol and Chlorophyll Compounds, for Virgin Olive Oil Monitoring Under Different Photo- and Thermal-Oxidation Procedures Abstract Room temperature excitation-emission matrices (EEMs), recorded using front-face fluorescence spectroscopy, and combined with second-order multiway classification methods, were explored as potential tools for controlling the storage conditions of extra virgin olive oil samples (EVOO). Factors such as UV-irradiation, sunlight exposition and temperature up to 80 °C were studied. For each sample, EEMs were obtained in two spectral regions, corresponding to polyphenols, and chlorophyll and derivatives, respectively. The full fluorescence information of excitation-emission matrices was processed with parallel factor analysis (PARAFAC) and PARAFAC supervised by linear discriminant analysis (LDA). The models allowed the discrimination between non-irradiated and irradiated EVOO samples, in both spectral regions. With a temperature of 80 °C, and a heating time of 30 min, the formation of secondary oxidation products was appreciable. In these conditions, the first component of PARAFAC showed a remarkable modification in its profile and LDA-PARAFAC allowed the discrimination between non-heated and heated EVOO samples.

Magnetic Graphene Solid-Phase Extraction for the Determination of 47 Kinds of Non-steroidal Anti-inflammatory Drug Residues in Animal Food with Liquid Chromatography Tandem Mass Spectrometry Abstract Based on magnetic graphene (Fe3O4-G), a liquid chromatography with tandem mass spectrometric detection (LC-MS/MS) method coupled with magnetic solid-phase extraction was developed for the simultaneous detection of 47 kinds of nonsteroidal anti-inflammatory drug (NSAID) residues in meat of swine, chicken, and bovine. The samples were extracted with acetonitrile-phosphate, then the extracts were purified by dispersed solid-phase extraction based on magnetic graphene (Fe3O4-G). The linearity range is 0.1–50 μg L−1. Limits of detection were between 0.1 μg kg−1 and 0.5 μg kg−1, and limits of quantification were between 0.5 μg kg−1 and 1.0 μg kg−1. The average recoveries were 72.4–97.1% at spiking levels. The inter-day RSDs varied from 5.3 to 12.6%. Due to the advantages of easy collection of magnetic solid-phase extraction materials, the method was more rapid and efficient for the enrichment and separation without filling columns. The sensitivity, precision, and accuracy method was suitable for simultaneous detection of multi-component residues of nonsteroidal anti-inflammatory drugs.

Magnetically Modified Porous β-Cyclodextrin Polymers for Dispersive Solid-Phase Extraction High-Performance Liquid Chromatography Analysis of Sudan Dyes Abstract Two types of magnetically modified porous β-cyclodextrin polymers (PCDPs) were successfully synthesized. The first type, named MPCDPs(C), was fabricated by co-precipitating PCDPs with Fe3O4, resulting in a Brunauer–Emmett–Teller (BET) surface area of 106.68 m2 g−1. The second type, named MPCDPs(M), was fabricated by grafting PCDPs onto the surface of magnetic microspheres, resulting in a BET surface area of 34.63 m2 g−1. Both MPCDPs were found to enable effective extraction and separation of Sudan dyes from foodstuffs and environmental water. Coupled with high-performance liquid chromatography, the limits of detection for Sudan dyes were in the range of 0.013–0.054 ng mL−1 with MPCDPs(C), and 0.028–0.039 ng mL−1 with MPCDPs(M). Recoveries were between 85.8 and 102.8% for spiked food samples and 88.3 and 103.2% for spiked water samples, suggesting that the developed methods are promising for accurate quantification of Sudan dyes at trace levels in foodstuffs and environmental water.

Isotope Dilution LC-MS/MS Method for Glycine Betaine in Manila Clam ( Tapes philippinarum ) Abstract Glycine betaine (GB) is a natural compound with demonstrated metabolic functions in mammals and beneficial effects on the human body: it protects cells under osmotic stress, as an organic osmolyte, and acts as a methylating agent in several vital biochemical pathways. The main dietary sources of GB are vegetables, such as cereals, spinach, chard and beetroot, above all; shellfish is one of the richest animal sources. A simple method for the quantification of GB in the edible portion of Manila clams (Tapes philippinarum) was setup, using ultra-performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS). Isotope-labelled internal standards were used during sample extraction, in order to ensure an accurate quantification. The method was successfully validated following the Commission Decision 657/2002/EC as a guideline; since GB is an endogenous compound, present in very high concentrations in the target matrix, appropriate adjustments were taken. The method developed reported very satisfying performances in terms of linearity, trueness and precision; moreover, its applicability was demonstrated on commercial pools of Manila clams.

Optimization of Ultrasound-Assisted Extraction of Anthocyanins and Phenolic Compounds from Black Soybeans ( Glycine max L.) Abstract Black soybeans have been reported to contain high antioxidant content, including anthocyanins. Ultrasound-assisted water extraction of antioxidants from Korean black soybeans (Glycine max [L.] Merr. Cheongja4ho) was investigated for the first time. A three-level, three-factor Box-Behnken design using a response surface methodology (RSM) was employed to optimize the conditions for the maximum yields of three dependent variables, including total anthocyanin content, total phenol content, and 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging activity, based on 17 different experiments. The solid-liquid ratio (X1, 1/30–1/50 g mL−1), extraction time (X2, 1–9 min), and amplitude (X3, 20–100%) significantly affected three dependent variables. The optimized conditions were solid-liquid ratio of 49.1 g mL−1, extraction time of 8.59 min, and amplitude of 81.4%. At the optimum point, the total anthocyanin content, total phenol content, and ABTS•+ scavenging activity were 66.44, 940.96, and 242.45 mg 100 g−1. The difference (%) between the predicted and experimental values was less than 6.40%, indicating that the model fits well. The conditions optimized for ultrasound-assisted water extraction of antioxidants from black soybeans can be applied as an efficient technique for extracting anthocyanins in plants on a larger scale.

Alexandros Sfakianakis
Anapafseos 5 . Agios Nikolaos
Crete.Greece.72100
2841026182

6948891480

alsfakia@gmail.com