Error detection for radiotherapy planning validation based on deep learning networks.

BACKGROUND: Quality assurance (QA) of patient-specific treatment plans for intensity-modulated radiation therapy (IMRT) and volumetric modulated arc therapy (VMAT) necessitates prior validation. However, the standard methodology exhibits deficiencies and lacks sensitivity in the analysis of positional dose distribution data, leading to difficulties in accurately identifying reasons for plan verification failure. This issue complicates and impedes the efficiency of QA tasks. PURPOSE: The primary aim of this research is to utilize deep learning algorithms for the extraction of 3D dose distribution maps and the creation of a predictive model for error classification across multiple machine models, treatment methodologies, and tumor locations. METHOD: We devised five categories of validation plans (normal, gantry error, collimator error, couch error, and dose error), conforming to tolerance limits of different accuracy levels and employing 3D dose distribution data from a sample of 94 tumor patients. A CNN model was then constructed to predict the diverse error types, with predictions compared against the gamma pass rate (GPR) standard employing distinct thresholds (3%, 3 mm; 3%, 2 mm; 2%, 2 mm) to evaluate the model's performance. Furthermore, we appraised the model's robustness by assessing its functionality across diverse accelerators. RESULTS: The accuracy, precision, recall, and F1 scores of CNN model performance were 0.907, 0.925, 0.907, and 0.908, respectively. Meanwhile, the performance on another device is 0.900, 0.918, 0.900, and 0.898. In addition, compared to the GPR method, the CNN model achieved better results in predicting different types of errors. CONCLUSION: When juxtaposed with the GPR methodology, the CNN model exhibits superior predictive capability for classification in the validation of the radiation therapy plan on different devices. By using this model, the plan validation failures can be detected more rapidly and efficiently, minimizing the time required for QA tasks and serving as a valuable adjunct to overcome the constraints of the GPR method.

Downregulation of SMAD4 protects HaCaT cells against UVB-induced damage and oxidative stress through the activation of EMT.

As a member of the SMAD family, SMAD4 plays a crucial role in several cellular biological processes. However, its function in UVB radiation-induced keratinocyte damage is not yet clarified. Our study aims to provide mechanistic insight for the development of future UVB protective therapies and therapeutics involving SMAD4. HaCaT cells were treated with UVB, and the dose dependence and time dependence of UVB were measured. The cell function of UVB-treated HaCaT cells and the activity of epithelial-mesenchymal transition (EMT) after overexpression or silencing of SMAD4 was observed by flow cytometry, quantitative reverse transcription PCR (qRT-PCR) and Western Blots (WB). We found that a significant decrease in SMAD4 was observed in HaCaT cells induced by UVB. Our data confirm SMAD4 as a direct downstream target of miR-664. The down-regulation of SMAD4 preserved the viability of the UVB-treated HaCaT cells by inhibiting autophagy or apoptosis. Furthermore, the silencing of SMAD4 activated the EMT process in UVB-treated HaCaT cells. Down-regulation of SMAD4 plays a protective role in UVB-treated HaCaT cells via the activation of EMT.

Immune Characteristic Genes and Neutrophil Immune Transformation Studies in Severe COVID-19.

As a disease causing a global pandemic, the progression of symptoms to severe disease in patients with COVID-19 often has adverse outcomes, but research on the immunopathology of COVID-19 severe disease remains limited. In this study, we used mRNA-seq data from the peripheral blood of COVID-19 patients to identify six COVID-19 severe immune characteristic genes (FPR1, FCGR2A, TLR4, S100A12, CXCL1, and L TF), and found neutrophils to be the critical immune cells in COVID-19 severe disease. Subsequently, using scRNA-seq data from bronchoalveolar lavage fluid from COVID-19 patients, neutrophil subtypes highly expressing the S100A family were found to be located at the end of cellular differentiation and tended to release neutrophil extracellular traps. Finally, it was also found that alveolar macrophages, macrophages, and monocytes with a high expression of COVID-19 severe disease immune characteristic genes may influence neutrophils through intercellular ligand-receptor pairs to promote neutrophil extracellular trap release. This study provides immune characteristic genes, critical immune pathways, and immune cells in COVID-19 severe disease, explores intracellular immune transitions of critical immune cells and pit-induced intercellular communication of immune transitions, and provides new biomarkers and potential drug targets for the treatment of patients with COVID-19 severe disease.

Bleomycin induces senescence and repression of DNA repair via downregulation of Rad51.

BACKGROUND: Bleomycin, a potent antitumor agent, is limited in clinical use due to the potential for fatal pulmonary toxicity. The accelerated DNA damage and senescence in alveolar epithelial cells (AECs) is considered a key factor in the development of lung pathology. Understanding the mechanisms for bleomycin-induced lung injury is crucial for mitigating its adverse effects. METHODS: Human lung epithelial (A549) cells were exposed to bleomycin and subsequently assessed for cellular senescence, DNA damage, and double-strand break (DSB) repair. The impact of Rad51 overexpression on DSB repair and senescence in AECs was evaluated in vitro. Additionally, bleomycin was intratracheally administered in C57BL/6 mice to establish a pulmonary fibrosis model. RESULTS: Bleomycin exposure induced dose- and time-dependent accumulation of senescence hallmarks and DNA lesions in AECs. These effects are probably due to the inhibition of Rad51 expression, consequently suppressing homologous recombination (HR) repair. Mechanistic studies revealed that bleomycin-mediated transcriptional inhibition of Rad51 might primarily result from E2F1 depletion. Furthermore, the genetic supplement of Rad51 substantially mitigated bleomycin-mediated effects on DSB repair and senescence in AECs. Notably, decreased Rad51 expression was also observed in the bleomycin-induced mouse pulmonary fibrosis model. CONCLUSIONS: Our works suggest that the inhibition of Rad51 plays a pivotal role in bleomycin-induced AECs senescence and lung injury, offering potential strategies to alleviate the pulmonary toxicity of bleomycin.

Terminally differentiated cytotoxic CD4+ T cells were clonally expanded in the brain lesion of radiation-induced brain injury.

BACKGROUND: Accumulating evidence supports the involvement of adaptive immunity in the development of radiation-induced brain injury (RIBI). Our previous work has emphasized the cytotoxic function of CD8+ T cells in RIBI. In this study, we aimed to investigate the presence and potential roles of cytotoxic CD4+ T cells (CD4+ CTLs) in RIBI to gain a more comprehensive understanding of adaptive immunity in this context. MAIN TEXT: Utilizing single-cell RNA sequencing (scRNA-seq), we analyzed 3934 CD4+ T cells from the brain lesions of four RIBI patients and identified six subclusters within this population. A notable subset, the cytotoxic CD4+ T cells (CD4+ CTLs), was marked with high expression of cytotoxicity-related genes (NKG7, GZMH, GNLY, FGFBP2, and GZMB) and several chemokine and chemokine receptors (CCL5, CX3CR1, and CCL4L2). Through in-depth pseudotime analysis, which simulates the development of CD4+ T cells, we observed that the CD4+ CTLs exhibited signatures of terminal differentiation. Their functions were enriched in protein serine/threonine kinase activity, GTPase regulator activity, phosphoprotein phosphatase activity, and cysteine-type endopeptidase activity involved in the apoptotic signaling pathway. Correspondingly, mice subjected to gamma knife irradiation on the brain showed a time-dependent infiltration of CD4+ T cells, an increase of MHCII+ cells, and the existence of CD4+ CTLs in lesions, along with an elevation of apoptotic-related proteins. Finally, and most crucially, single-cell T-cell receptor sequencing (scTCR-seq) analysis at the patient level determined a large clonal expansion of CD4+ CTLs in lesion tissues of RIBI. Transcriptional factor-encoding genes TBX21, RORB, and EOMES showed positive correlations with the cytotoxic functions of CD4+ T cells, suggesting their potential to distinguish RIBI-related CD4+ CTLs from other subsets. CONCLUSION: The present study enriches the understanding of the transcriptional landscape of adaptive immune cells in RIBI patients. It provides the first description of a clonally expanded CD4+ CTL subset in RIBI lesions, which may illuminate new mechanisms in the development of RIBI and offer potential biomarkers or therapeutic targets for the disease.

Anatomical changes and dosimetric analysis of the neck region based on FBCT for nasopharyngeal carcinoma patients during radiotherapy.

BACKGROUND: The study aimed to investigate anatomical changes in the neck region and their impact on dose distribution in patients with nasopharyngeal carcinoma (NPC) undergoing intensity modulated radiation therapy (IMRT), as well as to determine the optimal time for replanning during treatment. METHODS: Twenty NPC patients received IMRT with weekly pretreatment in-room kV fan beam computed tomography (FBCT) scans. Metastasized lymph nodes in the neck region and organs at risk (OARs) were recontoured based on the FBCT scans. The original treatment plan (PLAN0) was copied to each FBCT scan to create new plans of PLAN 1-6, correspondingly. The dose-volume histograms (DVH) of the new plans and the original plan were compared. One-way repeated measure ANOVA was employed to define threshold(s) at any timepoint. The presence of a threshold(s) would indicate significant anatomical change(s) such that replanning should be suggested. RESULTS: Progressive volume reductions in the neck region, gross target volume for metastatic lymph nodes (GTVnd), submandibular glands, and parotids were observed over time. Compared to PLAN0, Dmean of GTVnd-L significantly increased in PLAN5, while the D95% of PGTVnd-L showed a significant decrease from PLAN3 to PLAN6. Similarly, the Dmean of GTVnd-R significantly increased from PLAN4 to PLAN6, whereas the D95% of PGTVnd-R exhibited a significant decrease from PLAN3 to PLAN6. Furthermore, a gradual increase in the dose delivered to the bilateral parotid glands, bilateral submandibular glands, brainstem, and spinal cord from PLAN0 to PLAN6. CONCLUSION: Significant anatomic and dosimetric changes were observed in the target volumes and OARs. Based on the identified thresholds, replanning at approximately 20 fractions is crucial to ensure adequate target volumes dose and avoid overdosing to the OARs. This approach is clinically feasible and strongly recommended, particularly for centers without access to an adaptive planning system.

Stroke as initial manifestation of non-small cell lung cancer with Trousseau syndrome.

OBJECTIVE: Stroke is a rare but fatal complication of advanced cancer with Trousseau syndrome, especially as initial symptoms. Here, we report the clinical characteristics, treatment, and prognosis of patients with non-small cell lung cancer (NSCLC) who initially presenting with acute multiple cerebral infarction. METHODS: The clinical characteristics, imaging, treatment, and oncological outcomes of 10 patients diagnosed with Trousseau syndrome and NSCLC between 2015 and 2021 at Guangdong Sanjiu Brain Hospital were retrospectively collected and analyzed. The clinical course of two typical cases were presented. RESULTS: All 10 patients with pathologically confirmed lung adenocarcinoma initially presented with neurological symptoms, including hemiplegic paralysis (7 patients, 70%), dizziness (5 patients, 50%), and unclear speech (3 patients, 30%). The median age was 63.5 years. Eight and two cases were stage III and IV, respectively, at the initial diagnosis. Five patients underwent driver gene testing, revealing three patients with EGFR-sensitive mutations, one patient with ALK fusion, and one patient with wild-type EGFR. All 10 patients received antiplatelet therapy, and six patients subsequently received anti-cancer treatment. The median overall survival of the patients was 8.5 months (95% confidence interval) and 1-year survival rate was 57.1%. Patients who received antitumor treatment, especially those harboring driver gene mutations and received tyrosine kinase inhibitors, had better neurological symptom recovery and superior oncological prognosis (median overall survival, not reached versus 7.4 months, p = 0.038). CONCLUSION: Trousseau syndrome, presenting as multiple cerebral infarctions, is a rare complication of lung adenocarcinoma. Both antiplatelet and antitumor treatment are recommended to achieve better neurological recovery and oncological prognosis in these patients.

6-Gingerol alleviates placental injury in preeclampsia by inhibiting oxidative stress via BNIP3/LC3 signaling-mediated trophoblast mitophagy.

Background and aims: Preeclampsia (PE) is the leading cause of maternal and fetal morbidity and mortality worldwide. Apoptosis of trophoblast cells induced by oxidative stress is a principal reason of placental injury in PE. 6-Gingerol, an antioxidant from ginger, plays an important role in many disease models, but its effect on obstetric diseases has not been elucidated. In this study, we investigated the protective effect of 6-gingerol against placental injury. Methods: In vitro hypoxia/reoxygenation (H/R) model of HTR8/Svneo cells and preeclamptic mice model were established to simulate PE. The effects of 6-Gingerol on PE were evaluated by morphological detection, biochemical analysis, and Western blot. Results: We found that H/R treatment induced cell apoptosis, increased the production of reactive oxygen species, malondialdehyde and lactate dehydrogenase, and decreased superoxide dismutase in trophoblast. In addition, the polarization of mitochondrial membrane potential and the cellular calcium flux were also destroyed under H/R condition, which also activated BCL2-interacting protein 3 (BNIP3) and provoked excessive mitophagy. Importantly, 6-Gingerol reversed these corrosive effects. Furthermore, the placenta damage in PE-like mouse caused by the cell apoptosis, oxidative stress and mitophagy was mitigated by 6-Gingerol. Conclusion: These findings suggest that 6-Gingerol exerts a protective effect against placental injury in PE by reducing oxidative stress and inhibiting excessive mitophagy caused by mitochondrial dysfunction.

Causal Associations between Gut Microbiota and Different Types of Dyslipidemia: A Two-Sample Mendelian Randomization Study.

The determination of a causal association between gut microbiota and a range of dyslipidemia remains uncertain. To clarify these associations, we employed a two-sample Mendelian randomization (MR) analysis utilizing the inverse-variance weighted (IVW) method. This comprehensive analysis investigated the genetic variants that exhibited a significant association (p < 5 × 10-8) with 129 distinct gut microbiota genera and their potential link to different types of dyslipidemia. The results indicated a potential causal association between 22 gut microbiota genera and dyslipidemia in humans. Furthermore, these findings suggested that the impact of gut microbiota on dyslipidemia regulation is dependent on the specific phylum, family, and genus. Bacillota phylum demonstrated the greatest diversity, with 15 distinct genera distributed among eight families. Notably, gut microbiota-derived from the Lachnospiraceae and Lactobacillaceae families exhibit statistically significant associations with lipid levels that contribute to overall health (p < 0.05). The sensitivity analysis indicated that our findings possess robustness (p > 0.05). The findings of our investigation provide compelling evidence that substantiates a causal association between the gut microbiota and dyslipidemia in the human body. It is noteworthy to highlight the significant influence of the Bacillota phylum as a crucial regulator of lipid levels, and the families Lachnospiraceae and Lactobacillaceae should be recognized as probiotics that significantly contribute to this metabolic process.

PM2.5 leads to adverse pregnancy outcomes by inducing trophoblast oxidative stress and mitochondrial apoptosis via KLF9/CYP1A1 transcriptional axis.

Epidemiological studies have demonstrated that fine particulate matter (PM2.5) is associated with adverse obstetric and postnatal metabolic health outcomes, but the mechanism remains unclear. This study aimed to investigate the toxicological pathways by which PM2.5 damaged placental trophoblasts in vivo and in vitro. We confirmed that PM2.5 induced adverse gestational outcomes such as increased fetal mortality rates, decreased fetal numbers and weight, damaged placental structure, and increased apoptosis of trophoblasts. Additionally, PM2.5 induced dysfunction of the trophoblast cell line HTR8/SVneo, including in its proliferation, apoptosis, invasion, migration and angiogenesis. Moreover, we comprehensively analyzed the transcriptional landscape of HTR8/SVneo cells exposed to PM2.5 through RNA-Seq and observed that PM2.5 triggered overexpression of pathways involved in oxidative stress and mitochondrial apoptosis to damage HTR8/SVneo cell biological functions through CYP1A1. Mechanistically, PM2.5 stimulated KLF9, a transcription factor identified as binding to CYP1A1 promoter region, which further modulated the CYP1A1-driven downstream phenotypes. Together, this study demonstrated that the KLF9/CYP1A1 axis played a crucial role in the toxic progression of PM2.5 induced adverse pregnancy outcomes, suggesting adverse effects of environmental pollution on pregnant females and putative targeted therapeutic strategies.

Gene Signatures for Latent Radiation-Induced Lung Injury Post X-ray Exposure in Mouse.

Objective: To investigate the X-ray-specific sensitive genes and potential signaling pathways involved in the latent period of radiation-induced lung injury (RILI) in mouse models. Method: Mice were randomized into groups for whole thoracic irradiation with a single fraction of 20 Gy X-ray or 12.5 Gy carbon heavy ion. Lungs were harvested 3 weeks after the irradiation, whole RNA was extracted and detected with the genome-wide transcriptional microarrays. Differentially expressed genes (DEGs) were calculated for each group and the X-ray-specific sensitive genes were determined, followed by the gene enrichment analysis of those DEGs exploring the potentially relevant signaling pathways and biological processes in latent RILI. Results: Three weeks after irradiation, gene expression levels varied between groups. 76 up-regulated DEGs were determined with mice in the X-ray group and gene ontology enrichment analysis for biological process (GO-BP) obtained several processes which were associated with radiation reaction, mitotic, immune cell chemotaxis or metastasis, immune factors, p53 apoptosis, and tissue remodeling. KEGG signaling pathway enrichment analysis showed that those 76 up-regulated DEGs were enriched in p53, IL-17, FoXO, melanoma, and non-small-cell lung cancer signaling pathways. By comparing the DEGs in X-ray and heavy ion groups, X-ray-specific sensitive genes were determined, the top 10 genes were Adamts9, Aacs, Col6a2, Fdps, Mdk, Mcam, Stbd1, Lbh, Ak3, and Emid1. The expression level of the top 10 genes was found to be significantly higher in the X-ray group than in the control and heavy ion groups. Conclusion: Our research determined the X-ray-specific sensitive gene set in mice lungs after exposure to radiation. The gene set could be used as a genetic marker to suggest the latency of RILI. The enrichment analysis results suggested that the relevant signaling pathways were potentially involved in the development of RILI. Further validation of those genes and signaling pathways is needed to confirm these findings.

Effect of peroxiredoxin 1 on the regulation of trophoblast function by affecting autophagy and oxidative stress in preeclampsia.

PURPOSE: PE is a pregnancy-specific syndrome and one of the main causes of maternal, fetal, and neonatal mortality. PRDX1 is an antioxidant that regulates cell proliferation, differentiation, and apoptosis. The aim of this study is to investigate the effect of PRDX1 on the regulation of trophoblast function by affecting autophagy and oxidative stress in preeclampsia. METHODS: Western blotting, RT-qPCR, and immunofluorescence were used to examine the expression of PRDX1 in placentas. PRDX1-siRNA was transfected to knockdown PRDX1 in HTR-8/SVneo cells. The biological function of HTR-8/SVneo cells was detected by wound healing, invasion, tube formation, CCK-8, EdU, flow cytometry, and TUNEL assays. Western blotting was used to detect the protein expression of cleaved-Caspase3, Bax, LC3II, Beclin1, PTEN, and p-AKT. DCFH-DA staining was used to detect ROS levels by flow cytometry. RESULTS: PRDX1 was significantly decreased in placental trophoblasts in PE patients. Following the exposure of HTR-8/SVneo cells to H2O2, PRDX1 expression was significantly decreased, LC3II and Beclin1 expression was notably increased, and ROS level was also markedly increased. PRDX1 knockdown impaired migration, invasion, and tube-formation abilities and promoted apoptosis, which was accompanied by an increased expression of cleaved-Caspase3 and Bax. PRDX1 knockdown induced a significant decrease in LC3II and Beclin1 expression, along with an elevated p-AKT expression and a decreased PTEN expression. PRDX1 knockdown increased intracellular ROS levels, and NAC attenuated PRDX1 knockdown-induced apoptosis. CONCLUSION: PRDX1 regulated trophoblast function through the PTEN/AKT signaling pathway to affect cell autophagy and ROS level, which provided a potential target for the treatment of PE.

The shape effect and its consequences for polar surfaces and for heterogeneous catalysis.

In this paper we develop the shape effect, which is relevant for crystalline materials whose size is larger than that of the thermodynamic limit. According to this effect the electronic properties of one surface of a crystal depend upon all of its surfaces, i.e. on the overall shape. At first, qualitative mathematical arguments are presented for the existence of this effect based on the conditions for the stability of polar surfaces. Our treatment explains why such surfaces are observed even though earlier theory indicated that they should not exist. Then, models are developed from which it is found computationally that changing the shape of a polar crystal can substantially alter the magnitude of its surface charges. Apart from surface charges, it follows that the crystal shape will also significantly affect bulk properties, most notably polarization and piezoelectric responses. Additional model calculations show a strong shape effect on the activation energy for heterogeneous catalysis primarily through local surface charges rather than a non-local/long range electrostatic potential.

TIGAR deficiency induces caspase-1-dependent trophoblasts pyroptosis through NLRP3-ASC inflammasome.

Introduction: Gestational diabetes mellitus (GDM), a common complication of pregnancy, is risky for both mother and fetus. Previous studies about TP53-induced glycolysis and apoptosis regulator (TIGAR) focused on the occurrence and development of cancer, cardiovascular disease, and neurological disease, however, it is still unclear whether TIGAR plays a regulatory role in gestational diabetes mellitus (GDM). Methods: Utilizing HG exposure, we explored the role of TIGAR in oxidative stress limitation, excessive inflammatory toxicity defense, and pyroptosis prevention. Results: TIGAR was up-regulated in vivo and in vitro under HG condition, and loss of TIGAR increased ROS in trophoblast cells which drove a phenotypic switch and hindered the capacity of migration, invasion, and tube formation. This switch depended on the increased activation of NLRP3-ASC-caspase-1 signaling, which caused a distinctive characteristic of pyroptosis, and these findings could finally be reverted by antioxidant treatment (NAC) and receptor block (MCC950). Collectively, trophoblast pyroptosis is an upstream event of TIGAR deficiency-induced inflammation, which is promoted by ROS accumulation through NLRP3-ASC inflammasome. Conclusion: Taken together, our results uncovered that, as the upstream event of TIGAR deficiency-induced inflammation, pyroptosis is stimulated by ROS accumulation through NLRP3-ASC inflammasome.

The PTEN-associated immune prognostic signature reveals the landscape of the tumor microenvironment in glioblastoma.

Glioblastoma (GBM) is a common brain tumor with a complex and diverse tumor microenvironment (TME). As PTEN mutation is the most common mutation in GBM, we aimed to investigate how PTEN mutation regulates the immune response in GBM TME and thus affects the prognosis of GBM patients. In this study, we conducted a comprehensive analysis of multiple levels of data, including whole-exome sequencing (WES), transcriptome RNA sequencing, patient survival and immune signatures, to study the relationship between PTEN mutation and TME in GBM. We developed an immune-related prognostic signature (IPS) based on the PTEN-associated immune-related genes (IRGs), and the IPS exhibited a powerful prognosis prediction capacity in different GBM cohorts. A scoring nomogram based on the IPS was also established for clinical application. In addition, the correlations of the IPS with tumor immune cell infiltration and immune checkpoints were systematically analyzed. This study illustrates the influence of PTEN mutation on the immune microenvironment of GBM. Our IPS, which is sensitive to PTEN mutation status, can enhance the prognosis prediction ability for GBM patients and provides potential targets for immunotherapy.

Mechanistic insights into aerobic oxidative cleavage of glycol catalyzed by an Anderson-type polyoxometalate [IMo6O24]5.

A computational investigation of the aerobic oxidative C-C bond cleavage reaction of glycol catalyzed by an Anderson-type heteropolyanion HPA [IMo6O24]5- in the presence of acetonitrile as solvent has been performed at the WB97XD/6-31G(d,p)/lanl2dz level. Two reaction pathways have been identified. The catalytic cycle of each pathway consists of three steps: oxidation cleavage of a glycol molecule by the HPA, oxidation of the HPA by one dioxygen molecule, and, finally, oxidation of a second glycol and regeneration of the catalyst. These reaction pathways have been thoroughly investigated in terms of energetic, natural bond orbital (NBO), natural charges, and geometrical parameters. It is found that (i) even though the top oxygen atoms of the Anderson heteropolyanion are not the most negatively charged ones, they are more likely to react with the diol hydroxyl groups, (ii) a direct relationship between the presence of the iodine ion I(VII) and the studied oxidation reaction could not be identified, and (iii) in terms of energy, the transfer of the two hydrogen atoms is the most energetic step.

Partial Ablation of Astrocytes Exacerbates Cerebral Infiltration of Monocytes and Neuronal Loss After Brain Stab Injury in Mice.

In traumatic brain injury (TBI), mechanical injury results in instantaneous tissue damages accompanied by subsequent pro-inflammatory cascades composed of microgliosis and astrogliosis. However, the interactive roles between microglia and astrocytes during the pathogenesis of TBI remain unclear and sometimes debatable. In this study, we used a forebrain stab injury mouse model to investigate the pathological role of reactive astrocytes in cellular and molecular changes of inflammatory response following TBI. In the ipsilateral hemisphere of stab-injured brain, monocyte infiltration and neuronal loss, as well as increased elevated astrogliosis, microglia activation and inflammatory cytokines were observed. To verify the role of reactive astrocytes in TBI, local and partial ablation of astrocytes was achieved by stereotactic injection of diphtheria toxin in the forebrain of Aldh1l1-CreERT2::Ai9::iDTR transgenic mice which expressed diphtheria toxin receptor (DTR) in astrocytes after tamoxifen induction. This strategy achieved about 20% of astrocytes reduction at the stab site as validated by immunofluorescence co-staining of GFAP with tdTomato-positive astrocytes. Interestingly, reduction of astrocytes showed increased microglia activation and monocyte infiltration, accompanied with increased severity in stab injury-induced neuronal loss when compared with DTR-/- mice, together with elevation of inflammatory chemokines such as CCL2, CCL5 and CXCL10 in astrogliosis-reduced mice. Collectively, our data verified the interactive role of astrocytes as an immune modulator in suppressing inflammatory responses in the injured brain. Schematic diagram shows monocyte infiltration and neuronal loss, as well as increased elevated astrogliosis, microglia activation and chemokines were observed in the injured site after stab injury. Local and partial ablation of astrocytes led to increased microglia activation and monocyte infiltration, accompanied with increased severity in neuronal loss together with elevation of inflammatory chemokines as compared with control mice subjected stab injury.

Transcriptome Analysis Reveals Hub Genes Regulating Autophagy in Patients With Severe COVID-19.

Background: The COVID-19 pandemic has currently developed into a worldwide threat to humankind. Importantly, patients with severe COVID-19 are believed to have a higher mortality risk than those with mild conditions. However, despite the urgent need to develop novel therapeutic strategies, the biological features and pathogenic mechanisms of severe COVID-19 are poorly understood. Methods: Here, peripheral blood mononuclear cells (PBMCs) from four patients with severe COVID-19, four patients with mild COVID-19, and four healthy controls were examined by RNA sequencing (RNA-Seq). We conducted gene expression analysis and Venn diagrams to detect specific differentially expressed genes (DEGs) in patients with severe disease compared with those with mild conditions. Gene Ontology (GO) enrichment analysis was performed to identify the significant biological processes, and protein-protein interaction networks were constructed to extract hub genes. These hub genes were then subjected to regulatory signatures and protein-chemical interaction analysis for certain regulatory checkpoints and identification of potent chemical agents. Finally, to demonstrate the cell type-specific expression of these genes, we performed single-cell RNA-Seq analyses using an online platform. Results: A total of 144 DEGs were specifically expressed in severe COVID-19, and GO enrichment analysis revealed a significant association of these specific DEGs with autophagy. Hub genes such as MVB12A, CHMP6, STAM, and VPS37B were then found to be most significantly involved in the biological processes of autophagy at the transcriptome level. In addition, six transcription factors, including SRF, YY1, CREB1, PPARG, NFIC, and GATA2, as well as miRNAs, namely, hsa-mir-1-3p, and potent chemical agents such as copper sulfate and cobalt chloride, may cooperate in regulating the autophagy hub genes. Furthermore, classical monocytes may play a central role in severe COVID-19. Conclusion: We suggest that autophagy plays a crucial role in severe COVID-19. This study might facilitate a more profound knowledge of the biological characteristics and progression of COVID-19 and the development of novel therapeutic approaches to achieve a breakthrough in the current COVID-19 pandemic.

Efficient model photosensitizers based on metallocenyl complexes with thiophene-N = N-pyrimidine as π-conjugated bridge and cyanoacrylate as an anchoring group: a density functional theory study.

Eight push-pull systems involving containing four transition metals (iron, ruthenium, cobalt, and nickel), metallocenes as donor groups, cyanoacrylate as electron attractor group, and thiophene-N = N- pyrimidine derivatives as π-conjugated bridges were designed and studied using DFT and TD-DFT methods involving B3LYP and CAM-B3LYP functionals combined with the cc-pVDZ/LANL2DZ basis sets. The main purpose of this work is to determine the effect of metallocene in improving the photosensitization property of such chromophores. This was done by calculating their light-harvesting efficiency LHE as well as other properties employed for DSSC application. The considered dyes were first studied in the gas phase, then in the presence of TiO2 nanoparticles representing the semi-conductor, and finally in the presence of a specific implicit solvent. The presence of iron as metal involved in the metallocene group supplemented by extending the π-conjugated bridge by a cyanovinyl spacer was demonstrated so as to give the most optimal response taking into account the lower cost and toxicity as well as the friendliness to the environment of iron as metal.

TRPA1 promotes melanosome phagocytosis in keratinocytes via PAR-2/CYLD axis.

BACKGROUND: Keratinocytes are recipients of melanosomes. Although the chemical basis of melanogenesis is well documented, the molecular mechanism of melanosome transfer must be elucidated. TRPA1 is a member of the transient receptor potential A subfamily. Previous studies have shown that inhibition of TRPA1 activity reduces melanin synthesis in human epidermal melanocytes; however, the mechanism remains unknown. OBJECTIVE: This study aimed to investigate the roles and mechanism(s) of action of TRPA1 in keratinocytes. METHODS: The correlation between TRPA1 expression levels and the ability of keratinocytes to phagocytize melanosomes was examined using melanin silver staining. TRPA1 depleted human epidermal keratinocytes and keratinocyte cell lines HaCaT were established using adenovirus-expressing shRNAs against TRPA1. The effects of TRPA1 on keratinocytes and HaCaT cells were determined using cell-based analyses, including light stimulation, calcium imaging, melanin phagocytosis, immunoblotting, and co-immunoprecipitation assays. The degree of epidermal pigmentation was determined in a guinea pig model. RESULTS: TRPA1 mediated the phagocytic activity of keratinocytes. TRPA1 knockdown markedly suppressed melanosome transport to keratinocytes. Mechanistically, TRPA1 was required for PAR-2-induced melanosome phagocytosis in keratinocytes. Furthermore, TRPA1 activation indirectly stabilized microtubules by promoting the competitive binding of CYLD and acetylated α-tubulin. In addition, bortezomib (PS-341), a proteasome inhibitor, increased TRPA1 and CYLD expression and promoted phagocytic activity both in vitro and in vivo. CONCLUSIONS: Our findings firstly suggest that TRPA1 promotes melanosome transport in keratinocytes and reveal that TRPA1 is a regulator of PAR-2 activation and microtubule stability via the PAR-2/CYLD axis.