The chemokine CCL1 triggers an AMFR-SPRY1 pathway that promotes differentiation of lung fibroblasts into myofibroblasts and drives pulmonary fibrosis.

Recruitment of immune cells to the site of inflammation by the chemokine CCL1 is important in the pathology of inflammatory diseases. Here, we examined the role of CCL1 in pulmonary fibrosis (PF). Bronchoalveolar lavage fluid from PF mouse models contained high amounts of CCL1, as did lung biopsies from PF patients. Immunofluorescence analyses revealed that alveolar macrophages and CD4+ T cells were major producers of CCL1 and targeted deletion of Ccl1 in these cells blunted pathology. Deletion of the CCL1 receptor Ccr8 in fibroblasts limited migration, but not activation, in response to CCL1. Mass spectrometry analyses of CCL1 complexes identified AMFR as a CCL1 receptor, and deletion of Amfr impaired fibroblast activation. Mechanistically, CCL1 binding triggered ubiquitination of the ERK inhibitor Spry1 by AMFR, thus activating Ras-mediated profibrotic protein synthesis. Antibody blockade of CCL1 ameliorated PF pathology, supporting the therapeutic potential of targeting this pathway for treating fibroproliferative lung diseases.

Targeting Degradation of the Transcription Factor C/EBPß Reduces Lung Fibrosis by Restoring Activity of the Ubiquitin-Editing Enzyme A20 in Macrophages.

Although recent progress provides mechanistic insights into the pathogenesis of pulmonary fibrosis (PF), rare anti-PF therapeutics show definitive promise for treating this disease. Repeated lung epithelial injury results in injury-repairing response and inflammation, which drive the development of PF. Here, we report that chronic lung injury inactivated the ubiquitin-editing enzyme A20, causing progressive accumulation of the transcription factor C/EBPß in alveolar macrophages (AMs) from PF patients and mice, which upregulated a number of immunosuppressive and profibrotic factors promoting PF development. In response to chronic lung injury, elevated glycogen synthase kinase-3ß (GSK-3ß) interacted with and phosphorylated A20 to suppress C/EBPß degradation. Ectopic expression of A20 or pharmacological restoration of A20 activity by disturbing the A20-GSK-3ß interaction accelerated C/EBPß degradation and showed potent therapeutic efficacy against experimental PF. Our study indicates that a regulatory mechanism of the GSK-3ß-A20-C/EBPß axis in AMs may be a potential target for treating PF and fibroproliferative lung diseases.

Clock knockout in inhibitory neurons reduces predisposition to epilepsy and influences anxiety-like behaviors in mice.

Epilepsy is a brain disorder affecting up to 1 in 26 individuals. Despite its clinical importance, the molecular mechanisms of epileptogenesis are still far from clarified. Our previous study showed that disruption of Clock in excitatory neurons alters cortical circuits and leads to generation of focal epilepsy. In this study, a GAD-Cre;Clockflox/flox mouse line with conditional Clock gene knockout in inhibitory neurons was established. We observed that seizure latency was prolonged, the severity and mortality of pilocarpine-induced seizure were significantly reduced, and memory was improved in GAD-Cre;Clockflox/flox mice. We hypothesize that mice with CLOCK knockout in inhibitory neurons have increased threshold for seizure, opposite from mice with CLOCK knockout in excitatory neurons. Further investigation showed Clock knockout in inhibitory neurons upregulated the basal protein level of ARC, a synaptic plasticity-associated immediate-early gene product, likely through the BDNF-ERK pathway. Altered basal levels of ARC may play an important role in epileptogenesis after Clock deletion in inhibitory neurons. Although sEPSCs and intrinsic properties of layer 5 pyramidal neurons in the somatosensory cortex exhibit no changes, the spine density increased in apical dendrite of pyramidal neurons in CLOCK knockout group. Our results suggest an underlying mechanism by which the circadian protein CLOCK in inhibitory neurons participates in neuronal activity and regulates the predisposition to epilepsy.

microRNA profilings identify plasma biomarkers and targets associated with pediatric epilepsy patients.

BACKGROUND: Although previous studies show that microRNAs (miRNAs) can potentially be used as diagnostic markers for epilepsy, there are very few analyses of pediatric epilepsy patients. METHODS: miRNA profiles using miRNA-seq was performed on plasma samples from 14 pediatric epileptic patients and 14 healthy children. miRNA miR-27a-3p that were significantly changed between two groups were further evaluated. The potential target genes of miR-27a-3p were screened through unbiased mRNA-seq and further validated using Western blot and immunohistochemistry in HEK-293T cells and in the brains of mice with epilepsy induced by lithium chloride-pilocarpine. RESULTS: We found 82 upregulated and 76 downregulated miRNAs in the plasma from pediatric patients compared with controls (p < 0.01), of which miR-27a-3p exhibited a very low p value (p < 0.0001) and validated in additional plasma samples. Two genes, GOLM1 and LIMK1, whose mRNA levels were decreased (p < 0.001) with the increase of miR-27a-3p were further validated in both HEK-293T cells and in epileptic mice. CONCLUSIONS: MiR-27a-3p exhibits potential as a diagnostic and therapeutic marker for epilepsy. We postulate that additional studies on the downstream targets of miR-27a-3p will unravel its roles in epileptogenesis or disease progression. IMPACT: A total of 158 differentially expressed miRNAs were detected in plasma between epileptic and control children. Plasma miR-27a-3p was one of the miRNAs with a low p value. GOLM1 and LIMK1 were validated as downstream target genes of miR-27a-3p. miR-27a-3p has potential as a diagnostic and therapeutic marker for epilepsy.

Evidence for an intra-tumoral microbiome in pituitary neuroendocrine tumors with different clinical phenotypes.

PURPOSE: Bacteria have been observed in the tumor environment for decades and have been demonstrated to play important roles in the pathogenesis and development of several different tumors. So far there is a clear lack of specific studies relating to the presence of bacteria in pituitary neuroendocrine tumors (PitNETs). METHODS: In this study, we performed five region-based amplification and bacterial 16 S rRNA sequencing to identify the microbiome of PitNET tissues across four clinical phenotypes. Multiple filter procedures were performed to inhibit the risk of contamination with bacteria and bacterial DNA. Histological analysis was also conducted to validate the localization of bacteria in the intra-tumoral region. RESULTS: We identified common and diverse bacterial types across the four clinical phenotypes of PitNET. We also predicted the potential functions of these bacteria in tumor phenotypes and found that these functions were reported in certain previous mechanistic studies. Our data indicate that the pathogenesis and development of tumors may correlate with the behavior of intra-tumoral bacteria. Histological results, including lipopolysaccharide (LPS) staining and fluorescence in situ hybridization (FISH) for bacterial 16 S rRNA clearly demonstrated the localization of bacteria in the intra-tumoral region. Staining for Iba-1 suggested that the proportion of microglia was more abundant in FISH-positive regions than in FISH-negative regions. Furthermore, in FISH-positive regions, the microglia exhibited a longitudinally branched morphology that was different to the compact morphology observed in FISH-negative regions. CONCLUSION: In summary, we provide an evidence for the existence of intra-tumoral bacteria in PitNET.

Pan-Cancer Study of the Prognosistic Value of Selenium Phosphate Synthase 1.

Objective: This study sought to determine the mean prognostic usefulness of seleniumphosphate synthase (SEPHS1) by investigating its expression in 33 human malignancies and its relationship to tumor immunity.Methods: The expression of selenophosphate synthase 1 (SEPHS1) in 33 human malignant tumors was examined using the Genotype-Tissue Expression (GTEx), Cancer Genome Atlas (TCGA), and TIMER databases. Furthermore, the TCGA cohort was used to investigate relationships between SEPHS1 and immunological checkpoint genes (ICGs), tumor mutation burden (TMB), microsatellite instability (MSI), and DNA mismatch repair genes (MMRs). To establish independent risk factors and calculate survival probabilities for liver hepatocellular carcinoma (LIHC) and brain lower-grade glioma (LGG), Cox regression models and Kaplan-Meier curves were utilized. Eventually, the Genomics of Cancer Drug Sensitivity (GDSC) database was used to evaluate the drug sensitivity in LGG and LIHC patients with high SEPHS1 expression.Results: Overall, in numerous tumor tissues, SEPHS1 was highly expressed, and it significantly linked with the prognosis of LGG, ACC, and LIHC (P < .05). Furthermore, in numerous cancers, SEPHS1 expression was linked to tumor-infiltrating immune cells (TIICs), TMB, MSI, and MMRs. According to univariate and multivariate Cox analyses, SEPHS1 expression was significant for patients with LGG and LIHC.Conclusion: High SEPHS1 expression has a better prognosis for LGG, while low SEPHS1 expression has a better prognosis for LIHC. Chemotherapy was advised for LGG patients, particularly for those with high SEPHS1 expression because it can predict how responsive patients will be to 5-Fluorouracil and Temozolomide. This interaction between SEPHS1 and chemoradiotherapy has a positive clinical impact and may be used as evidence for chemotherapy for LGG and LIHC patients.

Jasmonates modulate sphingolipid metabolism and accelerate cell death in the ceramide kinase mutant acd5.

Sphingolipids are structural components of the lipid bilayer that acts as signaling molecules in many cellular processes, including cell death. Ceramides, key intermediates in sphingolipid metabolism, are phosphorylated by the ceramide kinase ACCELERATED CELL DEATH5 (ACD5). The loss of ACD5 function leads to ceramide accumulation and spontaneous cell death. Here, we report that the jasmonate (JA) pathway is activated in the Arabidopsis (Arabidopsis thaliana) acd5 mutant and that methyl JA treatment accelerates ceramide accumulation and cell death in acd5. Moreover, the double mutants of acd5 with jasmonate resistant1-1 and coronatine insensitive1-2 exhibited delayed cell death, suggesting that the JA pathway is involved in acd5-mediated cell death. Quantitative sphingolipid profiling of plants treated with methyl JA indicated that JAs influence sphingolipid metabolism by increasing the levels of ceramides and hydroxyceramides, but this pathway is dramatically attenuated by mutations affecting JA pathway proteins. Furthermore, we showed that JAs regulate the expression of genes encoding enzymes in ceramide metabolism. Together, our findings show that JAs accelerate cell death in acd5 mutants, possibly by modulating sphingolipid metabolism and increasing ceramide levels.

Arabidopsis alkaline ceramidase ACER functions in defense against insect herbivory.

Plant sphingolipids are important membrane components and bioactive molecules in development and defense responses. However, the function of sphingolipids in plant defense, especially against herbivores, is not fully understood. Here, we report that Spodoptera exigua feeding affects sphingolipid metabolism in Arabidopsis, resulting in increased levels of sphingoid long-chain bases, ceramides, and hydroxyceramides. Insect-induced ceramide and hydroxyceramide accumulation is dependent on the jasmonate signaling pathway. Loss of the Arabidopsis alkaline ceramidase ACER increases ceramides and decreases long-chain base levels in plants; in this work, we found that loss of ACER enhances plant resistance to S. exigua and improves response to mechanical wounding. Moreover, acer-1 mutants exhibited more severe root-growth inhibition and higher anthocyanin accumulation than wild-type plants in response to methyl jasmonate treatment, indicating that loss of ACER increases sensitivity to jasmonate and that ACER functions in jasmonate-mediated root growth and secondary metabolism. Transcript levels of ACER were also negatively regulated by jasmonates, and this process involves the transcription factor MYC2. Thus, our findings reveal that ACER is involved in mediating jasmonate-related plant growth and defense and that jasmonates function in regulating the expression of ACER.

Licochalcone A inhibits cell growth through the downregulation of the Hippo pathway via PES1 in cholangiocarcinoma cells.

Overexpression or activation of Yes-associated protein (YAP) is common in cancer cells. Thus, targeting YAP may be a strategy for cancer therapy. Licochalcone A (LicA) is a primary active compound of licorice root and is known to have medicinal effects, such as antioxidant, antibacterial, antiviral, and anticancer effects. However, the anticancer pharmacological mechanism of LicA has not been investigated in cholangiocarcinoma. In this study, we investigated the antiproliferative effect of LicA and the underlying molecular mechanism in HCCC-9810 and RBE human cholangiocarcinoma cells. Our experiments indicated that LicA suppressed the growth of cholangiocarcinoma cells through inactivation of the Hippo pathway. Pescadillo ribosomal biogenesis factor 1 (PES1) was notably upregulated and related to carcinogenesis. We also found that LicA suppressed the expression and nuclear localization of PES1, which was associated with the inhibition of YAP expression and transcriptional activity.

Metabolic reprogramming by traditional Chinese medicine and its role in effective cancer therapy.

Metabolic reprogramming, characterized by alterations of cellular metabolic patterns, is fundamentally important in supporting the malignant behaviors of cancer cells. It is considered as a promising therapeutic target against cancer. Traditional Chinese medicine (TCM) and its bioactive components have been used in cancer therapy for an extended period, and they are well-known for their multi-target pharmacological functions and fewer side effects. However, the detailed and advanced mechanisms underlying the anticancer activities of TCM remain obscure. In this review, we summarized the critical processes of cancer cell metabolic reprogramming, including glycolysis, mitochondrial oxidative phosphorylation, glutaminolysis, and fatty acid biosynthesis. Moreover, we systemically reviewed the regulatory effects of TCM and its bioactive ingredients on metabolic enzymes and/or signal pathways that may impede cancer progress. A total of 46 kinds of TCMs was reported to exert antitumor effects and/or act as chemosensitizers via regulating metabolic processes of cancer cells, and multiple targets and signaling pathways were revealed to contribute to the metabolic-modulating functions of TCM. In conclusion, TCM has its advantages in ameliorating cancer cell metabolic reprogramming by its poly-pharmacological actions. This review may shed some new light on the explicit recognition of the mechanisms of anticancer actions of TCM, leading to the development of natural antitumor drugs based on reshaping cancer cell metabolism.

A novel Rap-Phr system in Bacillus velezensis NAU-B3 regulates surfactin production and sporulation via interaction with ComA.

Several quorum sensing systems occurring in Bacillus subtilis, e.g. Rap-Phr systems, were reported to interact with major regulatory proteins, such as ComA, DegU, and Spo0A, in order to regulate competence, sporulation, and synthesis of secondary metabolites. In this study, we characterized a novel Rap-Phr system, RapA4-PhrA4, in Bacillus velezensis NAU-B3. We found that the rapA4 and phrA4 genes were co-transcribed in NAU-B3. When rapA4 was expressed in the heterologous host Bacillus subtilis OKB105, surfactin production and sporulation were severely inhibited. However, when the phrA4 was co-expressed, the RapA4 activity was inhibited. The transcription of the surfactin synthetase srfA gene and sporulation-related genes were also regulated by the RapA4-PhrA4 system. In vitro results obtained from electrophoretic mobility shift assay (EMSA) proved that RapA4 inhibits ComA binding to the promoter of the srfA operon, and the PhrA4 pentapeptide acts as anti-activator of RapA4. We also found that the F24 residue plays a key role in RapA4 function. This study indicated that the novel RapA4-PhrA4 system regulates the surfactin synthesis and sporulation via interaction with ComA, thereby supporting the bacterium to compete and to survive in a hostile environment. KEY POINTS: •Bacillus velezensis NAU-B3 has a novel Rap-Phr quorum sensing system, which does not occur in model strains Bacillus subtilis 168 and B. velezensis FZB42. •RapA4-PhrA4 regulates surfactin production and sporulation. •RapA4-PhrA4 interacts with the ComA protein from ComP/ComA two-component system.

[Variant analysis on steroid 5-reductase type 2 deficiency caused by a novel SRD5A2 mutation].

This article reported the clinical characteristics and SRD5A2 gene mutation pattern of a child with steroid 5-α reductase type 2 deficiency. The 2-month-old boy showed hypospadias and short penis shortly after birth. DNA was extracted from the peripheral blood of the child and his parents. The endocrine disease-related genes were captured and sequenced by high-throughput sequencing technology, and the family DNA samples were verified by Sanger sequencing. The results showed that c.680G>A(p.R227Q) and c.608G>A(p.G203D) compound heterozygous mutations existed in the SRD5A2 gene of the child. The c.680G>A mutation inherited from his father, which was a known pathogenic mutation. The c.608G>A mutation originated from his mother, which was a novel mutation discovered in this study. These results provide molecular evidence for the etiological diagnosis of the child and genetic counseling for the family, as well as extend the mutation spectrum of SRD5A2 gene.

A validated high-performance liquid chromatography-tandem mass spectrometry method for quantification of gefitinib and its main metabolites in xenograft mouse tumor: Application to a pharmacokinetics study.

Monitoring gefitinib and its metabolites may help to explore the underlying mechanisms of gefitinib resistance. The concentration of gefitinib and its metabolites in tumor tissues could influence its anticancer activities more than that in the plasma. In the present study, a rapid and specific HPLC-MS/MS method was developed and validated to simultaneously determine gefitinib, M387783, M523595, M537194 and M608236 in tumor tissues of H1975 human lung cancer xenografts of nude mice. The established HPLC-MS/MS method was validated for specificity, linearity, accuracy and precision, matrix effect and recovery, carryover and dilution integrity, and analyte stability. The standard curves were linear (r2 ≥ 0.99) over the range of 0.5-100 ng/mL for M608236 and 1-200 ng/mL for gefitinib, M523595 and M537194 as well as M387783. The accuracy ranged from -8.35 to 6.03% relative error; and the precision was <15% relative standard deviation. Recoveries (87.74-99.96%) and matrix effects (86.60-106.40%) were satisfactory in the biological matrix examined. Stability studies showed that the analytes were stable during the assay procedure and storage. Finally, the validated method was successfully applied to study the pharmacokinetics profiles for gefitinib and its metabolites in nonsmall cell lung cancer (NSCLC) xenograft mouse tumors. Meanwhile, MTT assay showed that gefitinib had a more powerful inhibitory effect than its four major metabolites in H1975 NSCLC cells. This validated HPLC-MS/MS method may be applied to help understand the mechanisms of gefitinib resistance in EGFR-mutant nonsmall cell lung cancer.

Control on helical filaments by twisted beams in a nonlinear CS2 medium.

Curved filament with large bent angle and controllable propagation behavior has always been great expectation and challenge due to its novelty and complexity. The unique properties of curved filaments make it possible to achieve many applications in micro-fabrication, spectroscopy and meteorology. Here we realize experimentally and theoretically control on helical filaments induced by twisted beams in CS2. The results show that helical filaments exhibit a robust pattern and high rotation rate. Specific intensity pattern of the twisted beam confines the filaments in fixed relative position and the azimuthal energy flux drives the rotating of the filamentation pattern. In addition, we demonstrated that the global orbital angular momentum (OAM) of twisted beams is still conservative to be zero, but local OAMs exhibit distinct variation during nonlinear propagation. Our idea has its significance which realizes the construction of helical filaments with flexibility and controllability and then facilitates to push the development of related researches.

Femtosecond polarization-structured optical field meets an anisotropic nonlinear medium.

Filamentation, as a universal femtosecond phenomenon that could occur in various nonlinear systems, has aroused extensive interest, owing to its underlying physics, complexity and applicability. It is always anticipated to realize the controllable and designable filamentation. For this aim, the crucial problem is how to actively break the symmetry of light-matter nonlinear interaction. A kind of extensively used approaches is based on the controllable spatial structure of optical fields involving phase, amplitude and polarization. Here we present an idea to control the optical field collapse by introducing optical anisotropy of matter as an additional degree of freedom, associated with polarization structure. Our theoretical prediction and experimental results reveal that the synergy of optical anisotropy and polarization structure is indeed a very effective means for controlling the optical field collapse, which has the robust feature against random noise.

Measurement of the topological charge and index of vortex vector optical fields with a space-variant half-wave plate.

Vortex vector optical fields (VVOFs) refer to a kind of vector optical field with an azimuth-variant polarization and a helical phase, simultaneously. Such a VVOF is defined by the topological index of the polarization singularity and the topological charge of the phase vortex. We present a simple method to measure the topological charge and index of VVOFs by using a space-variant half-wave plate (SV-HWP). The geometric phase grating of the SV-HWP diffracts a VVOF into ±1 orders with orthogonally left- and right-handed circular polarizations. By inserting a polarizer behind the SV-HWP, the two circular polarization states project into the linear polarization and then interfere with each other to form the interference pattern, which enables the direct measurement of the topological charge and index of VVOFs.

Marsdenia tenacissima extract induces apoptosis and suppresses autophagy through ERK activation in lung cancer cells.

BACKGROUND: Marsdenia tenacissima is an herb medicine which has been utilized to treat malignant diseases for decades. The M. tenacissima extract (MTE) shows significant anti-proliferation activity against non-small cell lung cancer (NSCLC) cells, but the underlying mechanisms remain unclear. In this study, we explored the potential anti-proliferation mechanisms of MTE in NSCLC cells in relation to apoptosis as well as autophagy, which are two critical forms to control cancer cell survival and death. METHODS: The proliferation of H1975 and A549 cells was evaluated by MTT assay. Cell apoptosis was assessed by Annexin V and PI staining, Caspase 3 expression and activity. Autophagy flux proteins were detected by Western blot with or without autophagy inducer and inhibitor. Endogenous LC3-II puncta and LysoTracker staining were monitored by confocal microscopy. The formation of autophagic vacuoles was measured by acridine orange staining. ERK is a crucial molecule to interplay with cell autophagy and apoptosis. The role of ERK on cell apoptosis and autophagy influenced by MTE was determined in the presence of MEK/ERK inhibitor U0126. RESULTS: The significant growth inhibition and apoptosis induction were observed in MTE treated NSCLC cells. MTE induced cell apoptosis coexisted with elevated Caspase 3 activity. MTE also impaired autophagic flux by upregulated LC3-II and p62 expression. Autophagy inducer EBSS could not abolish the impaired autophagic flux by MTE, while it was augmented in the presence of autophagy inhibitor Baf A1. The autophagosome-lysosome fusion was blocked by MTE via affecting lysosome function as evidenced by decreased expression of LAMP1 and Cathepsin B. The molecule ERK became hyperactivated after MTE treatment, but the MEK/ERK inhibitor U0126 abrogated autophagy inhibition and apoptosis induction caused by MTE, suggested that ERK signaling pathways partially contributed to cell death caused by MTE. CONCLUSION: Our results demonstrate that MTE caused apoptosis induction as well as autophagy inhibition in NSCLC cells. The activated ERK is partially associated with NSCLC apoptotic and autophagic cell death in response to MTE treatment. The present findings reveal new mechanisms for the anti-tumor activity of MTE against NSCLC.

Clinical trial on the effects of thalidomide on hemoglobin synthesis in patients with moderate thalassemia intermedia.

To investigate the efficacy and safety of thalidomide in patients with thalassemia intermedia (TI). Patients with a confirmed diagnosis of TI who met the trial criteria and signed consent forms were prescribed oral thalidomide 50 mg qn for 3 months from February 2017. Complete blood counts, Hb analysis, and liver and kidney functions were monitored monthly during treatment and any differences were compared before and after treatment. Patients with Hb increments > 2.0 g/dL were termed main responders (MaR), and those with Hb increments between 1.0 and 2.0 g/dL as minor responders (MiR), otherwise they were termed non-responders. Relevance analysis was performed to explore parameters predicting Hb increments after treatment. Adverse effects during treatment were carefully recorded. The overall response rate (ORR = MaR + MiR) and MaR rates were 78.6 and 50% after 1 month of treatment, respectively, and 85.7 and 71.4% after 3 months treatment. At the end of the treatment period, Hb and HbF increased by 2.5 ± 1.8 g/dL and 2.5 ± 1.6 g/dL, while bilirubin, lactate dehydrogenase, and the nucleated red blood cell count (NRBC) were significantly decreased, while the reticulocyte count significantly increased. Correlation analysis showed that the Hb increments correlated significantly with the ratio of HbF before treatment (r = 0.683, P = 0.007) rather than age, Hb, reticulocyte count, and NRBC before treatment. Adverse events during treatment were mild, and drug reduction or withdrawal from the trial was not required. Thalidomide had rapid and significant effects in patients with TI, and also, it is safe and convenient. But larger scale clinical trials will be required to confirm our conclusions. TRIAL REGISTRATION: NCT02995707, https://www.clinicaltrials.gov/ct2/show/NCT03184844?term=thalidomide+thalassemia&rank=1 .