Role of serial fluorodeoxyglucose positron emission tomography-computed tomography (18FDG-PET-CT) in assessing treatment response in treatment naïve chronic pulmonary aspergillosis subjects.

BACKGROUND: The role of 2-deoxy-2-18(F) fluoro-D-glucose (FDG) positron emission tomography (PET)-computed tomography (CT) in assessing treatment response in chronic pulmonary aspergillosis (CPA) remains to be determined. OBJECTIVE: To compare changes in FDG-PET/CT parameters in CPA subjects with treatment success or failure. METHODS: We treated consecutive treatment-naïve CPA subjects with six months of oral itraconazole. We performed PET-CT at baseline and six months. A multi-disciplinary team categorized response as treatment success or failure. We recorded the maximum standardised uptake value (SUVmax), SUVpeak, and total glycolytic activity (TLG). After treatment, FDG uptake similar to the background uptake or ≥13 units decline in Z-score was considered a complete metabolic response (CMR). A >25%, >30%, and > 45% decline in SUVmax, SUVpeak, and TLG was labelled as a partial metabolic response (PMR). A >30%, >30%, or >75% increase in the SUVmax, SUVpeak, and TLG represented progressive metabolic disease. RESULTS: We included 94 CPA subjects (63 males) with a mean age of 46.2 years. A follow-up PET-CT was performed on 77 subjects. We recorded treatment success and failure in 43 and 34 subjects. The median SUVmax at baseline was 6.7, which significantly reduced with treatment. CMR was seen in 18.6% of those with treatment success and none with treatment failure. A higher proportion of subjects with treatment success achieved PMR. 19% of the subjects with treatment success had progressive metabolic disease. CONCLUSION: FGD-PET/CT demonstrated metabolic activity in all CPA subjects. Most PET-CT parameters improved with treatment; however, one-fifth of the subjects were misclassified on PET-CT.

A Point Mutation in Cassette Relieves the Repression Regulation of CcpA Resulting in an Increase in the Degradation of 2,3-Butanediol in Lactococcus lactis.

In lactic acid bacteria, the global transcriptional regulator CcpA regulates carbon metabolism by repressing and activating the central carbon metabolism pathway, thus decreasing or increasing the yield of certain metabolites to maximize carbon flow. However, there are no reports on the deregulation of the inhibitory effects of CcpA on the metabolism of secondary metabolites. In this study, we identified a single-base mutant strain of Lactococcus lactis N8-2 that is capable of metabolizing 2,3-butanediol. It has been established that CcpA dissociates from the catabolite responsive element (cre) site due to a mutation, leading to the activation of derepression and expression of the 2,3-butanediol dehydrogenase gene cluster (butB and butA). Transcriptome analysis and quantitative polymerase chain reaction (Q-PCR) results showed significant upregulation of transcription of butB and butA compared to the unmutated strain. Furthermore, micro-scale thermophoresis experiments confirmed that CcpA did not bind to the mutated cre. Furthermore, in a bacterial two-plasmid fluorescent hybridization system, it was similarly confirmed that the dissociation of CcpA from cre eliminated the repressive effect of CcpA on downstream genes. Finally, we investigated the differing catalytic capacities of the 2,3-butanediol dehydrogenase gene cluster in L. lactis N8-1 and L. lactis N8-2 for 2,3-butanediol. This led to increased expression of butB and butA, which were deregulated by CcpA repression. This is the first report on the elimination of the deterrent effect of CcpA in lactic acid bacteria, which changes the direction of enzymatic catalysis and alters the direction of carbon metabolism. This provides new perspectives and strategies for metabolizing 2,3-butanediol using bacteria in synthetic biology.

Scrutinizing a Lactococcus lactis mutant with enhanced capacity for extracellular electron transfer reveals a unique role for a novel type-II NADH dehydrogenase.

Lactococcus lactis, a lactic acid bacterium used in food fermentations and commonly found in the human gut, is known to possess a fermentative metabolism. L. lactis, however, has been demonstrated to transfer metabolically generated electrons to external electron acceptors, a process termed extracellular electron transfer (EET). Here, we investigated an L. lactis mutant with an unusually high capacity for EET that was obtained in an adaptive laboratory evolution (ALE) experiment. First, we investigated how global gene expression had changed, and found that amino acid metabolism and nucleotide metabolism had been affected significantly. One of the most significantly upregulated genes encoded the NADH dehydrogenase NoxB. We found that this upregulation was due to a mutation in the promoter region of NoxB, which abolished carbon catabolite repression. A unique role of NoxB in EET could be attributed and it was directly verified, for the first time, that NoxB could support respiration in L. lactis. NoxB, was shown to be a novel type-II NADH dehydrogenase that is widely distributed among gut microorganisms. This work expands our understanding of EET in Gram-positive electroactive microorganisms and the special significance of a novel type-II NADH dehydrogenase in EET.IMPORTANCEElectroactive microorganisms with extracellular electron transfer (EET) ability play important roles in biotechnology and ecosystems. To date, there have been many investigations aiming at elucidating the mechanisms behind EET, and determining the relevance of EET for microorganisms in different niches. However, how EET can be enhanced and harnessed for biotechnological applications has been less explored. Here, we compare the transcriptomes of an EET-enhanced L. lactis mutant with its parent and elucidate the underlying reason for its superior performance. We find that one of the most significantly upregulated genes is the gene encoding the NADH dehydrogenase NoxB, and that upregulation is due to a mutation in the catabolite-responsive element that abolishes carbon catabolite repression. We demonstrate that NoxB has a special role in EET, and furthermore show that it supports respiration to oxygen, which has never been done previously. In addition, a search reveals that this novel NoxB-type NADH dehydrogenase is widely distributed among gut microorganisms.

Detection and mitigation of coordinated cyber-physical attack in CPPS.

Cyber-Physical Power System (CPPS) refers to a system in which the elements of the internet and the physical power system communicate and work together. With the use of modern communication and information technology, grid monitoring and control have improved. However, the components of a cyber system are extremely vulnerable to cyberattacks via cyber connections due to inadequate cyber security measures. Therefore, an adaptive defence strategy is required for the analysis and mitigation of the coordinated attack. The conventional approach of using an offline controller requires tuning for changes in the operating conditions of the system, which is inappropriate for the modern CPPS. To counter the coordinated attack, a framework that integrates STATCOM based Adaptive Model Predictive Controller with RPME and time delay compensator is proposed. This paper addresses attack impact, detection, and mitigation methods in CPPS. In both time domain and frequency domain simulations the case studies are conducted for three distinct situations namely physical attack, cyberattack, and coordinated attack. Convolutional Neural Network (CNN), Support Vector Machine (SVM), Random Forest (RF), and K Nearest Neighbour (KNN) are four data-driven methods used for the detection of anomalies in PMU measurement data. Simulation studies show that CNN performs better in anomaly detection than other classifiers based on assessed performance metrics. For coordinated attack mitigation the proposed STATCOM based Adaptive Model Predictive Controller with RPME quickly recovers the system than the STATCOM based conventional lead-lag controller. The efficacy of the proposed strategy is validated on the WSCC 3 machine 9 bus system.

Transcriptional regulation of cellobiose utilization by PRD-domain containing Sigma54-dependent transcriptional activator (CelR) and catabolite control protein A (CcpA) in Bacillus thuringiensis.

Cellobiose, a ß-1,4-linked glucose dimer, is a major cellodextrin resulting from the enzymatic hydrolysis of cellulose. It is a major source of carbon for soil bacteria. In bacteria, the phosphoenolpyruvate (PEP): carbohydrate phosphotransferase system (PTS), encoded by the cel operon, is responsible for the transport and utilization of cellobiose. In this study, we analyzed the transcription and regulation of the cel operon in Bacillus thuringiensis (Bt). The cel operon is composed of five genes forming one transcription unit. ß-Galactosidase assays revealed that cel operon transcription is induced by cellobiose, controlled by Sigma54, and positively regulated by CelR. The HTH-AAA+ domain of CelR recognized and specifically bound to three possible binding sites in the celA promoter region. CelR contains two PTS regulation domains (PRD1 and PRD2), which are separated by two PTS-like domains-the mannose transporter enzyme IIA component domain (EIIAMan) and the galactitol transporter enzyme IIB component domain (EIIBGat). Mutations of His-546 on the EIIAMan domain and Cys-682 on the EIIBGat domain resulted in decreased transcription of the cel operon, and mutations of His-839 on PRD2 increased transcription of the cel operon. Glucose repressed the transcription of the cel operon and catabolite control protein A (CcpA) positively regulated this process by binding the cel promoter. In the celABCDE and celR mutants, PTS activities were decreased, and cellobiose utilization was abolished, suggesting that the cel operon is essential for cellobiose utilization. Bt has been widely used as a biological pesticide. The metabolic properties of Bt are critical for fermentation. Nutrient utilization is also essential for the environmental adaptation of Bt. Glucose is the preferred energy source for many bacteria, and the presence of the phosphotransferase system allows bacteria to utilize other sugars in addition to glucose. Cellobiose utilization pathways have been of particular interest owing to their potential for developing alternative energy sources for bacteria. The data presented in this study improve our understanding of the transcription patterns of cel gene clusters. This will further help us to better understand how cellobiose is utilized for bacterial growth.

A New Mechanism of Carbon Metabolism and Acetic Acid Balance Regulated by CcpA.

Catabolite control protein A (CcpA) is a critical regulator in Gram-positive bacteria that orchestrates carbon metabolism by coordinating the utilization of different carbon sources. Although it has been widely proved that CcpA helps prioritize the utilization of glucose over other carbon sources, this global regulator's precise mechanism of action remains unclear. In this study, a mutant Bacillus licheniformis deleted for CcpA was constructed. Cell growth, carbon utilization, metabolites and the transcription of key enzymes of the mutant strain were compared with that of the wild-type one. It was found that CcpA is involved in the regulation of glucose concentration metabolism in Bacillus. At the same time, CcpA regulates glucose metabolism by inhibiting acetic acid synthesis and pentose phosphate pathway key gene zwF. The conversion rate of acetic acid is increased by about 3.5 times after ccpA is deleted. The present study provides a new mechanism of carbon metabolism and acetic acid balance regulated by CcpA. On the one hand, this work deepens the understanding of the regulatory function of CcpA and provides a new view on the regulation of glucose metabolism. On the other hand, it is helpful to the transformation of B. licheniformis chassis microorganisms.

luxS contributes to intramacrophage survival of Streptococcus agalactiae by positively affecting the expression of fruRKI operon.

The LuxS quorum sensing system is a widespread system employed by many bacteria for cell-to-cell communication. The luxS gene has been demonstrated to play a crucial role in intramacrophage survival of piscine Streptococcus agalactiae, but the underlying mechanism remains largely unknown. In this study, transcriptome analysis, followed by the luxS gene deletion and subsequent functional studies, confirmed that impaired bacterial survival inside macrophages due to the inactivation of luxS was associated with reduced transcription of the fruRKI operon, encoding the fructose-specific phosphotransferase system. Further, luxS was determined not to enhance the transcription of fruRKI operon by binding its promoter, but to upregulate the expression of this operon via affecting the binding ability of catabolite control protein A (CcpA) to the catabolite responsive element (cre) in the promoter of fruRKI. Collectively, our study identifies a novel and previously unappreciated role for luxS in bacterial intracellular survival, which may give a more thorough understanding of the immune evasion mechanism in S. agalactiae.

Sequence composition and location of CRE motifs affect the binding ability of CcpA protein.

Bacillus catabolite control protein (CcpA) mediates carbon catabolite repression (CCR) by binding with catabolite response elements (CREs) of genes or operons. Although numerous CREs had been predicted and identified, the influence of the changes in sequence and structure of CREs on recognition and binding for CcpA has yet to be unclear. This study aimed at revealing how CcpA could bind such diverse sites and focused on the analysis of multiple mutants of the CRE motif derived from the α-amylase promoter. Molecular docking and free energy calculation insights into the binding ability between the CRE sequences composition and CcpA protein. Disruption of conserved nucleotides in the CRE motifs, as well as altering the symmetric structure of the CRE sequences and the relative position of the displaced CRE motifs near the transcription start site contribute to some extent to weakening the strength of CcpA - dependent regulation. These main factors contribute to the understanding of the subtle changes in CRE motifs leading to differential regulatory effects of CcpA. Finally, an engineered promoter with a high level of transcription was obtained, and elevated extracellular enzyme activity was achieved in the expression system of Bacillus amyloliquefaciens, including alkaline protease, keratinase, aminopeptidase and acid-stable alpha amylase. The study also provides a reference for the application of other promoters with CRE motifts.

Two-carba cyclic phosphatidic acid treatment promotes phenotypic switch from M1 to M2 microglia and prevents behavioral abnormalities in a mouse model of neuropsychiatric systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease associated with the production of double-stranded DNA (dsDNA) antibodies and other antibodies that predominantly affects women with a wide range of lesions. Although neuropsychiatric lupus erythematosus (NPSLE), characterized by neuropsychiatric symptoms related to cerebrovascular diseases or depression, ranks high in severity, no specific treatment has been defined. Two-carba cyclic phosphatidic acid (2ccPA), a derivative of cyclic phosphatidic acid, was isolated from the true slime mold Physarum polycephalum in 1992. 2ccPA treatment suppresses neuroinflammation and promotes tissue repair in mouse multiple sclerosis and traumatic brain injury models. In this study, we performed behavioral tests on MRL/lpr mice as an NPSLE model. MRL/lpr mice showed increased depression-like behaviors compared with control mice, which were significantly suppressed by 2ccPA treatment. The expression of CD68, an M1 phenotypic marker of microglia, was significantly elevated in the prefrontal cortex and hippocampus of MRL/lpr mice, which was significantly suppressed by 2ccPA treatment. In contrast, the expression of Arginase1, an M2 phenotypic marker of microglia, was significantly increased by 2ccPA treatment. Compared to control mice, MRL/lpr mice showed higher plasma levels of anti-dsDNA antibodies, which are mainly involved in SLE pathogenesis. 2ccPA treatment decreased these levels in the MRL/lpr mice. These results suggest that 2ccPA treatment suppresses behavioral abnormalities by promoting a microglial phenotypic switch from M1 to M2 in MRL/lpr mice.

Role of C-Reactive Protein and Erythrocyte Sedimentation Rate in the Diagnosis and Monitoring of Treatment Response in Treatment Naïve Subjects with Chronic Pulmonary Aspergillosis.

PURPOSE: The role of C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) for diagnosing chronic pulmonary aspergillosis (CPA) remains unknown. Herein, we investigate the diagnostic performance of serum ESR and CRP in CPA. METHODS: We retrospectively analyzed the data of treatment-naïve subjects with CPA and diseased controls (post-tuberculosis lung disease on CT thorax). We treated CPA subjects with six months of oral itraconazole. Our primary objective was to evaluate the sensitivity and specificity of ESR and CRP in diagnosing CPA. The key secondary objective was to study the change in the inflammatory markers with treatment. RESULTS: We included 434 subjects and 20 diseased controls. The sensitivity and specificity of ESR (n = 434) and CRP (at cut-off value of 10 mg/L, n = 308) in diagnosing CPA were 42.9% and 65%, and 52.3% and 65%, respectively. Both ESR and CRP had erratic trend following treatment. ESR and CRP declined or remained stable in approximately 60% of subjects but increased in approximately 40% of the subjects despite treatment. CONCLUSION: Serum CRP and ESR have limited utility in diagnosing and following subjects with CPA.

CcpA and CodY Regulate CRISPR-Cas System of Streptococcus mutans.

Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) genes are widely recognized as bacterial adaptive immune systems against invading viruses and bacteriophages. The oral pathogen Streptococcus mutans encodes two CRISPR-Cas loci (CRISPR1-Cas and CRISPR2-Cas), and their expression under environmental conditions is still under investigation. In this study, we investigated the transcriptional regulation of cas operons by CcpA and CodY, two global regulators that contribute to carbohydrate and (p)ppGpp metabolism. The possible promoter regions for cas operons and the binding sites for CcpA and CodY in the promoter regions of both CRISPR-Cas loci were predicted using computational algorithms. We found that CcpA could directly bind to the upstream region of both cas operons, and detected an allosteric interaction of CodY within the same region. The binding sequences of the two regulators were identified through footprinting analysis. Our results showed that the promoter activity of CRISPR1-Cas was enhanced under fructose-rich conditions, while deletion of the ccpA gene led to reduced activity of the CRISPR2-Cas promoter under the same conditions. Additionally, deletion of the CRISPR systems resulted in a significant decrease in fructose uptake ability compared to the parental strain. Interestingly, the accumulation of guanosine tetraphosphate (ppGpp) was reduced in the presence of mupirocin, which induces a stringent response, in the CRISPR1-Cas-deleted (ΔCR1cas) and both CRISPR-Cas-deleted (ΔCRDcas) mutant strains. Furthermore, the promoter activity of both CRISPRs was enhanced in response to oxidative or membrane stress, while the CRISPR1 promoter activity was reduced under low-pH conditions. Collectively, our findings demonstrate that the transcription of the CRISPR-Cas system is directly regulated by the binding of CcpA and CodY. These regulatory actions play a crucial role in modulating glycolytic processes and exerting effective CRISPR-mediated immunity in response to nutrient availability and environmental cues. IMPORTANCE An effective immune system has evolved not only in eukaryotic organisms but also in microorganisms, enabling them to rapidly detect and neutralize foreign invaders in the environment. Specifically, the CRISPR-Cas system in bacterial cells is established through a complex and sophisticated regulatory mechanism involving specific factors. In this study, we demonstrate that the expression of two CRISPR systems in S. mutans can be controlled by two global regulators, CcpA and CodY, which play critical roles in carbohydrate metabolism and amino acid biosynthesis. Importantly, our results show that the expression of the CRISPR-Cas system in S. mutans influences (p)ppGpp production during the stringent response, which is a gene expression regulatory response that aids in environmental stress adaptation. This transcriptional regulation by these regulators enables a CRISPR-mediated immune response in a host environment with limited availability of carbon sources or amino acids, while ensuring efficient carbon flux and energy expenditure to support multiple metabolic processes.

Characterization of Treatment Response Outcomes in Chronic Pulmonary Aspergillosis: CPAnet Definitions Versus the Existing Criteria.

BACKGROUND: The chronic pulmonary aspergillosis network (CPAnet) has recently proposed definitions for treatment outcomes in CPA. However, these definitions need to be validated. Herein, we evaluate the agreement between the existing and the CPAnet definitions for response assessment. METHODS: We enrolled consecutive treatment-naïve CPA subjects (between January 2021 and June 2021) who received six months of itraconazole therapy and followed them for an additional six months after treatment discontinuation. We retrospectively applied the CPAnet criteria and compared the agreement between the existing and the CPAnet criteria for response assessment (primary objective). We also assessed if adding weight loss (> 5% from baseline) as a component improved the performance of the CPAnet criteria. RESULTS: We included 43 (mean age, 47.4 years) CPA subjects. The existing and the CPAnet criteria categorized 29 (67.4%) and 30 (69.8%) subjects as treatment success, respectively, at treatment completion. There was substantial (kappa = 0.73; p < 0.0001) agreement between the two definitions. However, both criteria did not identify eight subjects requiring treatment re-initiation within three months. There was an increment in the sensitivity of both criteria (by 36%) for identifying treatment failure after incorporating ≥ 5% weight loss as an element of worsening. CONCLUSION: The CPAnet definitions correctly categorized treatment outcomes in most cases of CPA. The addition of weight change would further enhance the performance of the CPAnet treatment outcome definitions.

Role of serum procalcitonin in the diagnosis and monitoring of treatment response in treatment-naïve subjects with chronic pulmonary aspergillosis.

Background: Interferon-gamma (IFN-γ) down-regulates plasma procalcitonin (PCT), marker of inflammation. Chronic pulmonary aspergillosis (CPA) is associated with low IFN-γ levels. Thus, plasma PCT may be elevated in CPA and could have a role in diagnosing and monitoring treatment response in CPA. Herein, we investigate the diagnostic performance of plasma PCT in CPA. Methods: We extracted the demographic, clinical, radiological, treatment outcomes, and plasma PCT levels of CPA subjects and controls (previously treated pulmonary tuberculosis with radiological abnormalities on CT chest [diseased controls] and treatment naïve active pulmonary tuberculosis [PTB]). We treated CPA subjects with six months of oral itraconazole. We took 0.25 ng/mL as a cut-off value for PCT. The study's primary objective was to ascertain the diagnostic performance of PCT in diagnosing CPA. The key secondary outcome was to study the change in the plasma PCT levels after itraconazole therapy. Results: We included 190 CPA cases and 40 controls (diseased controls [n = 20] and active PTB [n = 20]). PCT was elevated (≥0.25 ng/mL) in only 7 (3.7%) subjects with CPA. The sensitivity and specificity of PCT (≥0.25 ng/mL) were 3.7% (1.5-7.4%) and 100 (91.2-100%), respectively. The area under the curve for plasma PCT was 0.48 (95% confidence interval, 0.39-0.58). The plasma PCT values were available in 93 subjects at six months. There was a significant decline in the median plasma levels of PCT after treatment; however, the PCT levels either increased or remained the same in 45% of the subjects. Conclusion: Plasma procalcitonin has poor performance in diagnosing and following subjects with CPA.

comCDE (Competence) Operon Is Regulated by CcpA in Streptococcus pneumoniae D39.

Natural transformation plays an important role in the formation of drug-resistant bacteria. Exploring the regulatory mechanism of natural transformation can aid the discovery of new antibacterial targets and reduce the emergence of drug-resistant bacteria. Competence is a prerequisite of natural transformation in Streptococcus pneumoniae, in which comCDE operon is the core regulator of competence. To date, only ComE has been shown to directly regulate comCDE transcription. In this study, a transcriptional regulator, the catabolite control protein A (CcpA), was identified that directly regulated comCDE transcription. We confirmed that CcpA binds to the cis-acting catabolite response elements (cre) in the comCDE promoter region to regulate comCDE transcription and transformation. Moreover, CcpA can coregulate comCDE transcription with phosphorylated and dephosphorylated ComE. Regulation of comCDE transcription and transformation by CcpA was also affected by carbon source signals. Together, these insights demonstrate the versatility of CcpA and provide a theoretical basis for reducing the emergence of drug-resistant bacteria. IMPORTANCE Streptococcus pneumoniae is a major cause of bacterial infections in humans, such as pneumonia, bacteremia, meningitis, otitis media, and sinusitis. Like most streptococci, S. pneumoniae is naturally competent and employs this ability to augment its adaptive evolution. The current study illustrates CcpA, a carbon catabolite regulator, can participate in the competence process by regulating comCDE transcription, and this process is regulated by different carbon source signals. These hidden abilities are likely critical for adaptation and colonization in the environment.

StkP- and PhpP-Mediated Posttranslational Modifications Modulate the S. pneumoniae Metabolism, Polysaccharide Capsule, and Virulence.

Pneumococcal Ser/Thr kinase (StkP) and its cognate phosphatase (PhpP) play a crucial role in bacterial cytokinesis. However, their individual and reciprocal metabolic and virulence regulation-related functions have yet to be adequately investigated in encapsulated pneumococci. Here, we demonstrate that the encapsulated pneumococcal strain D39-derived D39ΔPhpP and D39ΔStkP mutants displayed differential cell division defects and growth patterns when grown in chemically defined media supplemented with glucose or nonglucose sugars as the sole carbon source. Microscopic and biochemical analyses supported by RNA-seq-based global transcriptomic analyses of these mutants revealed significantly down- and upregulated polysaccharide capsule formation and cps2 genes in D39ΔPhpP and D39ΔStkP mutants, respectively. While StkP and PhpP individually regulated several unique genes, they also participated in sharing the regulation of the same set of differentially regulated genes. Cps2 genes were reciprocally regulated in part by the StkP/PhpP-mediated reversible phosphorylation but independent of the MapZ-regulated cell division process. StkP-mediated dose-dependent phosphorylation of CcpA proportionately inhibited CcpA-binding to Pcps2A, supporting increased cps2 gene expression and capsule formation in D39ΔStkP. While the attenuation of the D39ΔPhpP mutant in two mouse infection models corroborated with several downregulated capsules-, virulence-, and phosphotransferase systems (PTS)-related genes, the D39ΔStkP mutant with increased amounts of polysaccharide capsules displayed significantly decreased virulence in mice compared to the D39 wild-type, but more virulence compared to D39ΔPhpP. NanoString technology-based inflammation-related gene expression and Meso Scale Discovery-based multiplex chemokine analysis of human lung cells cocultured with these mutants confirmed their distinct virulence phenotypes. StkP and PhpP may, therefore, serve as critical therapeutic targets.

2-carba-cyclic phosphatidic acid modulates astrocyte-to-microglia communication and influences microglial polarization towards an anti-inflammatory phenotype.

2-carba-cyclic phosphatidic acid (2ccPA) suppresses microglial and astrocyte inflammation for neuronal survival following traumatic brain injury. However, it remains unknown how 2ccPA regulates microglial activation. In this study, to elucidate the 2ccPA behavior in glial communication, we collected the astrocyte conditioned media (ACM) from primary astrocyte cultures that were treated by lipopolysaccharide (LPS) and 2ccPA and analyzed the alteration of microglial inflammation caused by the ACM treatment. The addition of the ACM derived from LPS- and 2ccPA-double treated astrocytes to microglia decreased the CD86+ pro-inflammatory M1 microglia, which were upregulated with the ACM collected from astrocytes treated by LPS without 2ccPA, while the direct addition of LPS and 2ccPA to microglia failed to decrease the CD86+ microglia to the basal level. We confirmed that the ACM from LPS- and 2ccPA-treated astrocytes increased the ratio of CD206+ anti-inflammatory M2 microglia to total microglia, whereas direct treatment of microglia with LPS and 2ccPA had no effect on the CD206+ microglia ratio, demonstrating the importance of astrocyte intervention in microglial polarization. In addition, we examined whether astrocytes modulate the 2ccPA-regulated proinflammatory cytokine production derived from microglia. The addition of the ACM from LPS- and 2ccPA-treated astrocytes to microglia remarkably canceled the LPS-induced upregulation of IL-1ß, IL-6, and TNF-α secreted from microglia, while the direct addition of LPS and 2ccPA to microglia showed no affect. Therefore, our results indicate that astrocytes mediate the 2ccPA function to shift microglia towards the M2 phenotype by interfering with the polarization of M1 microglia and to suppress cytokine production.

Astrocytic Neuroimmunological Roles Interacting with Microglial Cells in Neurodegenerative Diseases.

Both astrocytic and microglial functions have been extensively investigated in healthy subjects and neurodegenerative diseases. For astrocytes, not only various sub-types were identified but phagocytic activity was also clarified recently and is making dramatic progress. In this review paper, we mostly focus on the functional role of astrocytes in the extracellular matrix and on interactions between reactive astrocytes and reactive microglia in normal states and in neurodegenerative diseases, because the authors feel it is necessary to elucidate the mechanisms among activated glial cells in the pathology of neurological diseases in order to pave the way for drug discovery. Finally, we will review cyclic phosphatidic acid (cPA), a naturally occurring phospholipid mediator that induces a variety of biological activities in the brain both in vivo and in vitro. We propose that cPA may serve as a novel therapeutic molecule for the treatment of brain injury and neuroinflammation.

Effects of 2-carba-cyclic phosphatidic acid derivatives on IL-1ß-stimulated human chondrocytes.

Osteoarthritis (OA) is a common joint disease characterized by the breakdown of subchondral bone and cartilage damage, most often affecting middle-aged and elderly people. Although the etiology of OA is still unknown, some reports suggest that inflammatory factors such as interleukin (IL)- 1ß mediate the progression of OA. To investigate the effect of IL-1ß and the possibility of treatment for OA, we applied 2-carba-cyclic phosphatidic acid (2ccPA) and its derivatives on human chondrocytes. 2ccPA is a synthesized phospholipid derived from a bioactive phospholipid mediator: cyclic phosphatidic acid (cPA). It has been previously reported that 2ccPA exhibits anti-inflammatory and chondroprotective effects in an OA animal model. 2ccPA and its ring-opened body (ROB) derivative significantly suppressed IL-1ß-induced upregulation of IL-6, matrix metalloproteinase-13, and cyclooxygenase-2, as well as the degradation of type II collagen and aggrecan. However, the other two derivatives, namely the deacylated and ring-opened deacylated bodies, showed little effect on an IL-1ß-exposed human chondrosarcoma cell-line. These data suggest that the intactness of 2ccPA and ROB is essential for anti-inflammatory effects on OA. Collectively, this study provides evidence that 2ccPA and ROB would be novel therapeutic agents for OA.

Implementation of data protection laws in the European Union and in California is associated with a move of clinical trials to countries with fewer data protections.

The European Union implemented data privacy laws in mid-2018 and the state of California enacted a similar law several weeks later. These regulations affect medical data collection and analysis. It is unclear if they achieve this goal in the realm of clinical trials. Here we investigate the effect of these laws on clinical trials through analysis of clinical trials recorded on the US's ClinicalTrials.gov, the World Health Organization's International Clinical Trials Registry Platform and scientific papers describing clinical trials. Our findings show that the number of phase 1 and 2 trials in countries not adhering to these data privacy laws rose significantly after implementation of these laws. The largest rise occurred in countries which are less free, as indicated by the negative correlation (-0.48, p = 0.008) between the civil liberties freedom score of countries and the increase in the number of trials. This trend was not observed in countries adhering to data privacy laws nor in the paper publication record. The rise was larger (and statistically significant) among industry funded trials and interventional trials. Thus, the implementation of data privacy laws is associated a change in the location of clinical trials, which are currently executed more often in countries where people have fewer protections for their data.

The Role of Regulator Catabolite Control Protein A (CcpA) in Streptococcus agalactiae Physiology and Stress Response.

Streptococcus agalactiae is a leading cause of infections in neonates. This opportunistic pathogen colonizes the vagina, where it has to cope with acidic pH and hydrogen peroxide produced by lactobacilli. Thus, in the host, this bacterium possesses numerous adaptation mechanisms in which the pleiotropic regulators play a major role. The transcriptional regulator CcpA (catabolite control protein A) has previously been shown to be the major regulator involved in carbon catabolite repression in Gram-positive bacteria but is also involved in other functions. By transcriptomic analysis, we characterized the CcpA-dependent gene regulation in S. agalactiae. Approximately 13.5% of the genome of S. agalactiae depends on CcpA for regulation and comprises genes involved in sugar uptake and fermentation, confirming the role of CcpA in carbon metabolism. We confirmed by electrophoretic mobility shift assays (EMSAs) that the DNA binding site called cis-acting catabolite responsive element (cre) determined for other streptococci was effective in S. agalactiae. We also showed that CcpA is of capital importance for survival under acidic and oxidative stresses and is implicated in macrophage survival by regulating several genes putatively or already described as involved in stress response. Among them, we focused our study on SAK_1689, which codes a putative UspA protein. We demonstrated that SAK_1689, highly downregulated by CcpA, is overexpressed under oxidative stress conditions, this overexpression being harmful for the bacterium in a ΔccpA mutant. IMPORTANCE Streptococcus agalactiae is a major cause of disease burden leading to morbidity and mortality in neonates worldwide. Deciphering its adaptation mechanisms is essential to understand how this bacterium manages to colonize its host. Here, we determined the regulon of the pleiotropic regulator CcpA in S. agalactiae. Our findings reveal that CcpA is not only involved in carbon catabolite repression, but is also important for acidic and oxidative stress resistance and survival in macrophages.