Repurposing crizotinib to target RIPK1-dependent cell death.

Receptor-interacting protein kinase 1 (RIPK1) has emerged as a key regulator of cell death and inflammation, which are implicated in the pathogenesis of many inflammatory and degenerative diseases. RIPK1 is therefore a putative therapeutic target in many of these diseases. However, no pharmacological inhibitor of RIPK1-mediated cell death is currently in clinical use. Recognizing that a repurposed drug has an expedited clinical development pipeline, here we performed a high-throughput drug screen of Food and Drug Administration (FDA)-approved compounds and identified a novel use for crizotinib as an inhibitor of RIPK1-dependent cell death. Furthermore, crizotinib rescued TNF-α-induced death in mice with systemic inflammatory response syndrome. RIPK1 kinase activity was directly inhibited by crizotinib. These findings identify a new use for an established compound and are expected to accelerate drug development for RIPK1-spectrum disorders.

Quizartinib inhibits necroptosis by targeting receptor-interacting serine/threonine protein kinase 1.

Systemic inflammatory response syndrome (SIRS), at least in part driven by necroptosis, is characterized by life-threatening multiple organ failure. Blocking the progression of SIRS and consequent multiple organ dysfunction is challenging. Receptor-interacting serine/threonine protein kinase 1 (RIPK1) is an important cell death and inflammatory mediator, making it a potential treatment target in several diseases. Here, using a drug repurposing approach, we show that inhibiting RIPK1 is also an effective treatment for SIRS. We performed cell-based high-throughput drug screening of an US Food and Drug Administration (FDA)-approved drug library that contains 1953 drugs to identify effective inhibitors of necroptotic cell death by SYTOX green staining. Dose-response validation of the top candidate, quizartinib, was conducted in two cell lines of HT-22 and MEFs. The effect of quizartinib on necroptosis-related proteins was evaluated using western blotting, immunoprecipitation, and an in vitro RIPK1 kinase assay. The in vivo effects of quizartinib were assessed in a murine tumor necrosis factor α (TNFα)-induced SIRS model. High-throughput screening identified quizartinib as the top "hit" in the compound library that rescued cells from necroptosis in vitro. Quizartinib inhibited necroptosis by directly inhibiting RIPK1 kinase activity and blocking downstream complex IIb formation. Furthermore, quizartinib protected mice against TNFα-induced SIRS. Quizartinib, as an FDA-approved drug with proven safety and efficacy, was repurposed for targeted inhibition of RIPK1. This work provides essential preclinical data for transferring quizartinib to the treatment of RIPK1-dependent necroptosis-induced inflammatory diseases, including SIRS.

Creating a near-perfect circularly polarized terahertz beam through the nonreciprocity of a magnetoplasma.

Compared to other parts of the electromagnetic spectrum, the terahertz frequency range lacks efficient polarization manipulation techniques, which is impeding the proliferation of terahertz technology. In this work, we demonstrate a tunable and broadband linear-to-circular polarization converter based on an InSb plate containing a free-carrier magnetoplasma. In a wide spectral region (∼ 0.45 THz), the magnetoplasma selectively absorbs one circularly polarized mode due to electron cyclotron resonance and also reflects it at the edges of the absorption band. Both effects are nonreciprocal and contribute to form a near-zero transmission band with a high isolation of -36 dB, resulting in the output of a near-perfect circularly polarized terahertz wave for an incident linearly polarized beam. The near-zero transmission band is tunable with magnetic field to cover a wide frequency range from 0.3 to 4.8 THz.

Characterization of soundscape assessment in outdoor public spaces of urban high-rise residential communitiesa).

Soundscape perceptual models were developed in various contexts. However, as the outdoor public space in high-rise residential communities differs in terms of space planning and management, the soundscape perceptual characteristics are still unclear. In this study, an on-site survey was conducted to obtain the perceptual dimensions of soundscape in outdoor public spaces in urban high-rise residential communities based on evaluations of residents. Meantime, the soundscape of the space in different community layouts were compared. It was found that: (1) Four dimensions of outdoor soundscape in high-rise communities were extracted, namely Relaxation, Communication, Quietness, and Spatiality. The first three dimensions were positively correlated with overall soundscape satisfaction significantly. (2) Relaxation was mostly correlated with dominance of noise; Communication and Quietness were primarily related to sounds from human beings. (3) Lower traffic noise and higher levels of human sounds were perceived in enclosed communities, resulting in higher Relaxation and lower Quietness scores in outdoor public spaces in enclosed communities than non-enclosed ones. These findings evoke insights into the understanding of soundscape assessment in different contexts, and provide implications for sound environment design in urban high-rise communities.

Distribution of soil organic carbon and carbon sequestration potential of different geomorphic units in Shiyang river basin, China.

Accurate assessment of soil C storage patterns and control factors on a regional and global scale is essential for predicting and mitigating soil C feedback to global environmental change. We used soil samples collected in the Shiyang River Basin in 2018, combined with remote sensing data, climate and meteorological data, watershed hydrological data, and soil physical and chemical properties to discuss the change characteristics and influencing factors of soil organic carbon (SOC) under different soil depths in the Shiyang River Basin in an arid area and analyze the storage model and carbon sequestration potential of soil organic carbon in different geomorphic units. The research results show that, (1) in spatial distribution, the SOC content in the Shiyang River Basin shows an obvious regional difference, and the average content of SOC in the oasis area in the middle reaches significantly higher than that in the mountain area in the upstream and desert area in the downstream. In vertical distribution, the content of SOC in the whole watershed decreases with the soil depth increase. (2) Soil carbon sequestration potential decreased with the increase in soil depth, but there were regional differences. The Oasis area in the middle reaches of the Shiyang River Basin is a high-potential area. In contrast, most of the upper mountain areas and the lower reaches of the desert area are low-potential areas. Environmental factors such as vegetation cover, meteorological factors, and physical and chemical properties of soil are important factors that promote the spatial variability of SOC content. The decisive effect of environmental factors on the SOC content is most significant in the surface layer 0-20 cm.

Reverse design of pixel-type terahertz band-pass filters.

Reverse design is a frontier direction in the optical research field. In this work, reverse design is applied to the design of terahertz devices. We have employed direct binary search (DBS) and binary particle swarm optimization (BPSO) algorithms to design pixel-type terahertz band-pass filters, respectively. Through a comparative analysis of the designed devices, we found that BPSO algorithm converged faster than DBS algorithm, and the device performance is better on out-of-band suppression. We have fabricated a sample utilizing femtosecond laser micromachining and characterized it by terahertz time-domain spectroscopy. The experimental results were consistent with the finite difference time domain (FDTD) simulation. Our method can simultaneously optimize multiple characteristics of the band-pass filters, including the peak transmittance, out-of-band transmittance, bandwidth, and polarization stability, which can not be achieved by traditional optical design methods.

OPTN variants in ALS cases: a case report of a novel mutation and literature review.

INTRODUCTION: Optineurin (OPTN)-associated mutations have been implicated in the development of type 12 amyotrophic lateral sclerosis (ALS12). We reported a case of ALS with a new OPTN variant (p.D527fs) and reviewed relevant literature to better understand the phenotypes and pathophysiological mechanisms of ALS12. METHODS: We report a case of a 55-year-old female patient with a new heterozygous variant of the OPTN gene. A literature search of ALS cases associated with the OPTN gene mutations was performed in PubMed with the search criteria as [("amyotrophic lateral sclerosis") OR ("motor neuron disease")] AND ("OPTN"). RESULTS: The case of ALS with a new OPTN variant (p.D527fs) in our report manifested with bulbar involvement in onset and a rapidly progressive course. A literature review of 37 ALS patients with OPTN mutations included 20 males and 16 females with another patient whose gender was not described. The mean onset age of 37 ALS12 patients was 48 with the youngest 23 and the oldest 83 years old. Differences in onset age between male and female patients were not significant. Mean time from initiation to death was 61.8 ± 12.0 months. Patients present with either limb onset (73.5% cases) or bulbar onset (23.5% cases). CONCLUSION: Through the literature review, we summarized the clinical characteristics of ALS12. The phenotypes of the reported patients elucidate the genetic profiles and clinical phenotypes of ALS12. Clinicians should pay close attention to the role of receptor-interacting kinase 1 (RIPK1)-dependent necroptosis in the pathophysiologic development of ALS12, since necroptosis inhibitors are expected as potential therapeutic agents for treating ALS12.

GLCCI1 reduces collagen deposition and airway hyper-responsiveness in a mouse asthma model through binding with WD repeat domain 45B.

Asthma is a serious public health problem worldwide, without effective therapeutic methods. Our previous study indicated that glucocorticoid-induced transcript 1 gene (GLCCI1) knockout reduces the sensitivity to glucocorticoid in asthmatic mouse. Here, we explored the role and action mechanism of GLCCI1 in asthma development. In ovalbumin-sensitized mice, airway resistance and tissue damage increased, the production of inflammatory cytokines were up-regulated, GLCCI1 expression was reduced and autophagy was activated. Increasing of GLCCI1 inhibited human and mouse airway epithelial cell (AEC) autophagy, while decreasing of GLCCI1 promoted autophagy. Furthermore, we found that GLCCI1 bound with WD repeat domain 45B (WDR45B) and inhibited its expression. Increasing of WDR45B partly reversed the inhibition of GLCCI1 to autophagy-related proteins expression and autophagosome formation in vitro. Increasing of WDR45B in vivo reversed the improvement of GLCCI1 on airway remodelling in asthma and the inhibition to autophagy level in lung tissues. Overall, our data showed that GLCCI1 improved airway remodelling in ovalbumin-sensitized mice through inhibiting autophagy via combination with WDR45B and inhibiting its expression. Our results proved a new idea for asthma treatment.

Systematic Evaluation of Mycobacterium tuberculosis Proteins for Antigenic Properties Identifies Rv1485 and Rv1705c as Potential Protective Subunit Vaccine Candidates.

The lack of efficacious vaccines against Mycobacterium tuberculosis (MTB) infection is a limiting factor in the prevention and control of tuberculosis (TB), the leading cause of death from an infectious agent. Improvement or replacement of the BCG vaccine with one that reliably protects all age groups is urgent. Concerns exist that antigens currently being evaluated are too homogeneous. To identify new protective antigens, we screened 1,781 proteins from a high-throughput proteome-wide protein purification study for antigenic activity. Forty-nine antigens (34 previously unreported) induced antigen-specific gamma interferon (IFN-γ) release from peripheral blood mononuclear cells (PBMCs) derived from 4,452 TB and suspected TB patients and 167 healthy donors. Three (Rv1485, Rv1705c, and Rv1802) of the 20 antigens evaluated in a BALB/c mouse challenge model showed protective efficacy, reducing lung CFU counts by 66.2%, 75.8%, and 60%, respectively. Evaluation of IgG2a/IgG1 ratios and cytokine release indicated that Rv1485 and Rv1705c induce a protective Th1 immune response. Epitope analysis of PE/PPE protein Rv1705c, the strongest candidate, identified a dominant epitope in its extreme N-terminal domain accounting for 90% of its immune response. Systematic preclinical assessment of antigens Rv1485 and Rv1705c is warranted.

Fatty acylCoA synthetase FadD13 regulates proinflammatory cytokine secretion dependent on the NF-κB signalling pathway by binding to eEF1A1.

Mycobacterium tuberculosis (Mtb) manipulates multiple host defence pathways to survive and persist in host cells. Understanding Mtb-host cell interaction is crucial to develop an efficient means to control the disease. Here, we applied the Mtb proteome chip, through separately interacting with H37Ra and H37Rv stimulated macrophage lysates, screened 283 Mtb differential proteins. Through primary screening, we focused on fatty acylCoA synthetase FadD13. Mtb FadD13 is a potential drug target, but its role in infection remains unclear. Deletion of FadD13 in Mtb reduced the production of proinflammatory cytokines IL-1ß, IL-18, and IL-6. Bimolecular fluorescence complementation and colocalization showed that the binding partner of FadD13 in macrophage was eEF1A1 (a translation elongation factor). Knockdown eEF1A1 expression in macrophage abrogated the promotion of proinflammatory cytokines induced by FadD13. In addition, ΔfadD13 mutant decreased the expression of the NF-κB signalling pathway related proteins p50 and p65, so did the eEF1A1 knockdown macrophage infected with H37Rv. Meanwhile, we found that deletion of FadD13 reduced Mtb survival in macrophages during Mtb infection, and purified FadD13 proteins induced broken of macrophage membrane. Taken together, FadD13 is crucial for Mtb proliferation in macrophages, and it plays a key role in the production of proinflammatory cytokines during Mtb infection.

Retrospective Analysis of False-Positive and Disputed Rifampin Resistance Xpert MTB/RIF Assay Results in Clinical Samples from a Referral Hospital in Hunan, China.

Concerns about the specificity of the Xpert MTB/RIF (Xpert) assay have arisen, as false-positive errors in the determination of Mycobacterium tuberculosis complex (MTBC) infection and rifampin (RIF) resistance in clinical practice have been reported. Here, we investigated 33 cases where patients were determined to be RIF susceptible using the Bactec MGIT 960 (MGIT) culture system but RIF resistant using the Xpert assay. Isolates from two of these patients were found not to have any mutations in the rifampin resistance determining region (RRDR) region of rpoB and had good treatment outcomes with first-line antituberculosis (anti-TB) drugs. The remaining 31 patients included 5 new cases and 26 previously treated patients. A large number of well-documented disputed mutations, including Leu511Pro, Asp516Tyr, His526Asn, His526Leu, His526Cys, and Leu533Pro, were detected, and mutations, including a 508 to 509 deletion and His526Gly, were described here as disputed mutations for the first time. Twenty-one (81%) of the 26 previously treated patients had poor treatment outcomes, and isolates from 19 (90%) of these 21 patients were resistant to isoniazid (INH) as determined using the MGIT culture system. Twenty-seven of the 31 isolates with disputed rpoB mutations were phenotypically resistant to INH, 21 (78%) being predicted by GenoType MTBDRplus to have a high level of INH resistance. Most (77.4%) of the isolates with disputed mutations were of the Beijing lineage. These findings have implications for the interpretation of false-positive and disputed rifampin resistance Xpert MTB/RIF results in clinical samples and provide guidance on how clinicians should manage patients carrying isolates with disputed rpoB mutations.

Mycobacterium tuberculosis universal stress protein Rv2623 interacts with the putative ATP binding cassette (ABC) transporter Rv1747 to regulate mycobacterial growth.

We have previously shown that the Mycobacterium tuberculosis universal stress protein Rv2623 regulates mycobacterial growth and may be required for the establishment of tuberculous persistence. Here, yeast two-hybrid and affinity chromatography experiments have demonstrated that Rv2623 interacts with one of the two forkhead-associated domains (FHA I) of Rv1747, a putative ATP-binding cassette transporter annotated to export lipooligosaccharides. FHA domains are signaling protein modules that mediate protein-protein interactions to modulate a wide variety of biological processes via binding to conserved phosphorylated threonine (pT)-containing oligopeptides of the interactors. Biochemical, immunochemical and mass spectrometric studies have shown that Rv2623 harbors pT and specifically identified threonine 237 as a phosphorylated residue. Relative to wild-type Rv2623 (Rv2623WT), a mutant protein in which T237 has been replaced with a non-phosphorylatable alanine (Rv2623T237A) exhibits decreased interaction with the Rv1747 FHA I domain and diminished growth-regulatory capacity. Interestingly, compared to WT bacilli, an M. tuberculosis Rv2623 null mutant (ΔRv2623) displays enhanced expression of phosphatidyl-myo-inositol mannosides (PIMs), while the ΔRv1747 mutant expresses decreased levels of PIMs. Animal studies have previously shown that ΔRv2623 is hypervirulent, while ΔRv1747 is growth-attenuated. Collectively, these data have provided evidence that Rv2623 interacts with Rv1747 to regulate mycobacterial growth; and this interaction is mediated via the recognition of the conserved Rv2623 pT237-containing FHA-binding motif by the Rv1747 FHA I domain. The divergent aberrant PIM profiles and the opposing in vivo growth phenotypes of ΔRv2623 and ΔRv1747, together with the annotated lipooligosaccharide exporter function of Rv1747, suggest that Rv2623 interacts with Rv1747 to modulate mycobacterial growth by negatively regulating the activity of Rv1747; and that Rv1747 might function as a transporter of PIMs. Because these glycolipids are major mycobacterial cell envelope components that can impact on the immune response, our findings raise the possibility that Rv2623 may regulate bacterial growth, virulence, and entry into persistence, at least in part, by modulating the levels of bacillary PIM expression, perhaps through negatively regulating the Rv1747-dependent export of the immunomodulatory PIMs to alter host-pathogen interaction, thereby influencing the fate of M. tuberculosis in vivo.

Quantitative proteomic analysis of host responses triggered by Mycobacterium tuberculosis infection in human macrophage cells.

Macrophages are primary host of Mycobacterium tuberculosis (M.tb) and the central effector of in vivo innate immune responses against bacteria. Though the interaction between macrophages and mycobacteria has been widely investigated, the molecular mechanisms of M.tb pathogenesis in macrophages are still not clear. In this work, we investigated the altered protein expression profiles of macrophages after virulent H37Rv strain and avirulent H37Ra strain infection by tandem mass tag-based quantitative proteomics. Among 6762 identified proteins of macrophages, the expression levels of 235 proteins were significantly altered, which is supposed to be related to the infection of different strains. By bioinformatics analysis at systems level, we found that these proteins are mainly involved in the biological process of apoptosis, blood coagulation, oxidative phosphorylation, and others. The enormous variation in protein profiles between macrophages infected with H37Ra and H37Rv suggests the existence of four different immunity mechanisms that decide the fates of macrophages and M.tb. These data may provide a better understanding of M.tb pathogenesis within the host, which contributes to the prevention and clinical treatment of tuberculosis.

Later emergence of acquired drug resistance and its effect on treatment outcome in patients treated with Standard Short-Course Chemotherapy for tuberculosis.

BACKGROUNDS: The failure of current Standard Short-Course Chemotherapy (SCC) in new and previously treated cases with tuberculosis (TB) was mainly due to drug resistance development. But little is known on the characteristics of acquired drug resistant TB during SCC and its correlation with SCC failure. The objective of the study is to explore the traits of acquired drug resistant TB emergence and evaluate their impacts on treatment outcomes. METHODS: A prospective observational study was performed on newly admitted smear positive pulmonary TB (PTB) cases without drug resistance pretreatment treated with SCC under China's National TB Control Program (NTP) condition from 2008 to 2010. Enrolled cases were followed up through sputum smear, culture and drug susceptibility testing (DST) at the end of 1, 2, and 5 months after treatment initiation. The effect factors of early or late emergence of acquired drug resistant TB , such as acquired drug resistance patterns, the number of acquired resistant drugs and previous treatment history were investigated by multivariate logistic regression; and the impact of acquired drug resistant TB emergence on treatment failure were further evaluated. RESULTS: Among 1671 enrolled new and previously treated cases with SCC, 62 (3.7%) acquired different patterns of drug resistant TB at early period within 2 months or later around 3-5 months of treatment. Previously treated cases were more likely to develop acquired multi-drug resistant TB (MDR-TB) (OR, 3.8; 95%CI, 1.4-10.4; P = 0.015). Additionally, acquired MDR-TB cases were more likely to emerge at later period around 3-5 months after treatment starting than that of non-MDR-TB mainly appeared within 2 months (OR, 8.3; 95%CI, 1.7-39.9; P = 0.008). Treatment failure was associated with late acquired drug resistant TB emergence (OR, 25.7; 95%CI, 4.3-153.4; P < 0.001) with the reference of early acquired drug resistant TB emergence. CONCLUSIONS: This study demonstrates that later development of acquired drug resistant TB during SCC is liable to suffer treatment failure and acquired MDR-TB pattern may be one of the possible causes.

Restorative effect of audio and visual elements in urban waterfront spaces.

Introduction: Urban waterfront spaces are often composed of built infrastructures and nature elements. Though citizens could take advantage of these public spaces to relax from daily work, its restorative potential has not been paid enough attention. In this study, the restorative effect and mechanism of different audio and visual elements in urban waterfront spaces was systematically studied. Methods: At the first stage, restorative potential of waterfront spaces was investigated and different elements with restorative effects were identified through an on-site survey, in which visual and auditory forms of environmental-nature, animal-nature, on-water human activities and on-shore human activities were identified. At the second stage, a series of laboratory experiments were conducted to explore the restorative function of the audio and visual elements. Results and discussion: It is found that the degree of artificiality of waterfront space was a crucial factor influencing the restoration level of the space, and higher artificiality level of waterfront space resulted in lower level of perceived restoration. However it was available by adding visual and audio elements to the scene to facilitate the restorative effect in waterfront spaces with high-level artificiality. The effects of adding visual and auditory elements on psychophysiological restoration were explored, and elements that should be recommended and restrained were discussed. Prospects: These findings would provide applicable suggestions for future design and rebuilding of urban waterfront spaces.

The association between type 2 diabetes and pulmonary cavitation revealed among IGRA-positive tuberculosis patients.

The co-occurrence of tuberculosis (TB) and diabetes mellitus (DM) presents a significant obstacle to TB eradication. Pulmonary cavitation can occur in severe cases of TB, particularly in patients with DM. From 1 May 2014 through 30 June 2019, we conducted a cross-sectional study of 1,658 smear- or culture-confirmed pulmonary TB (PTB) patients at the Second Department of Pulmonary Medicine and Tuberculosis, Shenzhen, China. A total of 861 participants who satisfied the criteria (chest CT scan for cavitation, interferon-gamma release assay (IGRA), diagnosis of diabetes mellitus), with the median age of 36.7 years, 63.6% of male, 79.7% IGRA positive, 13.8% with diabetes, and 40.8% with pulmonary cavitation, were included in the study. The association between diabetes and pulmonary cavitation was confirmed in these TB patients (adjusted OR, 2.54; 95% CI, 1.66-3.94; p < 0.001). No associations were observed between diabetes and IGRA, as well as between lung cavitary and IGRA. Based on the criteria of IGRA+/-, pulmonary cavitation+/-, and DM+/-, the further analysis with univariate and multivariate logistic regression were conducted in six subgroups. The significant association between diabetes and pulmonary cavitation was further confirmed in the IGRA+ subgroup (adjusted OR, 3.07; 95% CI, 1.86-5.16; p < 0.001) but not observed in IGRA- individuals. This observation suggests that different immunological mechanisms of pulmonary cavitary/DM may be employed in IGRA+ TB patients from IGRA- TB patients.

Effects of agricultural activities on hydrochemistry in the Shiyang River Basin, China.

Agricultural water accounts for more than 80% of the available water in arid areas. Agricultural activities have a great impact on surface water and groundwater. If the impact of agricultural activities on hydrochemistry is not prevented, the risk of water quality change in arid areas may be greatly intensified. Based on the hydrochemical data of the whole Shiyang River Basin from April 2014 to October 2019, this paper analyzes the impact of agricultural activities on hydrochemistry in the basin. The results show that (i) in the middle and lower reaches of farmland with high intensity of agricultural activities, the ion concentration of groundwater in summer and autumn is significantly higher than that in winter and spring due to the influence of irrigation; (ii) the runoff ion concentration in the backflow of the river reaches recharged by irrigation water is significantly higher than that of other reaches; (iii) due to strong evaporation, different types of reservoirs will lead to an overall increase in ion concentration, which is more obvious in plain reservoirs and river tail lakes. In addition, the reservoirs have a certain removal effect on nitrates.

Identification of Piperlongumine as Potent Inhibitor of Necroptosis.

Purpose: Excessive necroptosis contributes to the pathogenesis of several inflammatory and neurodegenerative diseases. Here, using a high-throughput screening approach, we investigated the anti-necroptosis effects of piperlongumine, an alkaloid isolated from the long pepper plant, in vitro and in a mouse model of systemic inflammatory response syndrome (SIRS). Methods: A natural compound library was screened for anti-necroptosis effects in cellular. The underlying mechanism of action of the top candidate piperlongumine was explored by quantifying the necroptosis marker phosphorylated receptor-interacting protein kinase 1 (p-RIPK1) by Western blotting. The anti-inflammatory effect of piperlongumine was assessed in a tumor necrosis factor α (TNFα)-induced SIRS model in mice. Results: Among the compounds investigated, piperlongumine significantly rescued cell viability. The half maximal effective concentration (EC50) of piperlongumine for inhibiting necroptosis was 0.47 µM in HT-29 cells, 6.41 µM in FADD-deficient Jurkat cells, and 2.33 µM in CCRF-CEM cells, while the half maximal inhibitory concentration (IC50) was 95.4 µM in HT-29 cells, 93.02 µM in FADD-deficient Jurkat cells, and 161.1 µM in CCRF-CEM cells. Piperlongumine also significantly inhibited TNFα-induced intracellular RIPK1 Ser166 phosphorylation in cell lines and significantly prevented decreases in body temperature and improved survival in SIRS mice. Conclusion: As a potent necroptosis inhibitor, piperlongumine prevents phosphorylation of RIPK1 at its activation residue Ser166. Piperlongumine thus potently inhibits necroptosis at concentrations safe enough for human cells in vitro and inhibits TNFα-induced SIRS in mice. Piperlongumine has potential clinical translational value for the treatment of the spectrum of diseases associated with necroptosis, including SIRS.

Modular pathway engineering of diterpenoid synthases and the mevalonic acid pathway for miltiradiene production.

Microbial production can be advantageous over the extraction of phytoterpenoids from natural plant sources, but it remains challenging to rationally and rapidly access efficient pathway variants. Previous engineering attempts mainly focused on the mevalonic acid (MVA) or methyl-d-erythritol phosphate (MEP) pathways responsible for the generation of precursors for terpenoids biosynthesis, and potential interactions between diterpenoids synthases were unexplored. Miltiradiene, the product of the stepwise conversion of (E,E,E)-geranylgeranyl diphosphate (GGPP) catalyzed by diterpene synthases SmCPS and SmKSL, has recently been identified as the precursor to tanshionones, a group of abietane-type norditerpenoids rich in the Chinese medicinal herb Salvia miltiorrhiza . Here, we present the modular pathway engineering (MOPE) strategy and its application for rapid assembling synthetic miltiradiene pathways in the yeast Saccharomyces cerevisiae . We predicted and analyzed the molecular interactions between SmCPS and SmKSL, and engineered their active sites into close proximity for enhanced metabolic flux channeling to miltiradiene biosynthesis by constructing protein fusions. We show that the fusion of SmCPS and SmKSL, as well as the fusion of BTS1 (GGPP synthase) and ERG20 (farnesyl diphosphate synthase), led to significantly improved miltiradiene production and reduced byproduct accumulation. The MOPE strategy facilitated a comprehensive evaluation of pathway variants involving multiple genes, and, as a result, our best pathway with the diploid strain YJ2X reached miltiradiene titer of 365 mg/L in a 15-L bioreactor culture. These results suggest that terpenoids synthases and the precursor supplying enzymes should be engineered systematically to enable an efficient microbial production of phytoterpenoids.

Increased density of macrophage migration inhibitory factor (MIF) in tuberculosis granuloma.

Granulomas, the pathologic hallmarks of tuberculosis, are composed of tightly numerous immune cells that respond to a variety of persistent stimuli during pathogen-host interaction. The granuloma is essential for host containment of mycobacterial infection, however, the mechanism of host and pathogen determinants to recruit immune cells at the site of inflammation and the formation of granulomas remains elusive until now. Macrophage migration inhibitory factor (MIF), a cytokine produced by many cell types, modulates cellular and humoral immune responses and promote lymphocytes migration to the site of infection. In this study, we evaluate the expression of MIF in tuberculous granulomas by three different models of diseases: mouse, human tissues and zebrafish. The overall results demonstrated that the expression of MIF positive signals markedly increased in the tissues which have been infected with mycobacterium, whereas a few presence of MIF in the PBS-treated animals (means the control group). In the mycobacterial-infected animals, the MIF positives distributed extensively within the granuloma especially in the multinucleated giant cells. Thus, three independent lines of evidence support the hypothesis that MIF may be an important player in aggregate immune cells to the granuloma microenvironments in these animal models of tuberculosis.