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Abstract The senescence of alveolar type II (AT2) cells impedes self-repair of the lung epithelium and contributes to lung injury in the setting of idiopathic pulmonary fibrosis (IPF). Yes-associated protein 1 (YAP1) is essential for cell growth and organ development; however, the role of YAP1 in AT2 cells during pulmonary fibrosis is still unclear. YAP1 expression was found to be downregulated in the AT2 cells of PF patients. Deletion of YAP1 in AT2 cells resulted in lung injury, exacerbated extracellular matrix (ECM) deposition, and worsened lung function. In contrast, overexpression of YAP1 in AT2 cells promoted alveolar regeneration, mitigated pulmonary fibrosis, and improved lung function. In addition, overexpression of YAP1 alleviated bleomycin (BLM) -induced senescence of alveolar epithelial cells both in vivo and in vitro. Moreover, YAP1 promoted the expression of peroxiredoxin 3 (Prdx3) by directly interacting with TEAD1. Forced expression of Prdx3 inhibited senescence and improved mitochondrial dysfunction in BLM-treated MLE-12 cells, whereas depletion of Prdx3 partially abrogated the protective effect of YAP1. Furthermore, overexpression of Prdx3 facilitated self-repair of the injured lung and reduced ECM deposition, while silencing Prdx3 attenuated the antifibrotic effect of YAP1. In conclusion, this study demonstrated that YAP1 alleviates lung injury and pulmonary fibrosis by regulating Prdx3 expression to improve mitochondrial dysfunction and block senescence in AT2 cells, revealing a potential novel therapeutic strategy for pulmonary fibrosis.

Abstract This multicentre, two-arm, phase 2 study aimed to explore the efficacy and safety of neoadjuvant camrelizumab plus chemotherapy or apatinib in patients with initially unresectable stage II–III non-small-cell lung cancer (NSCLC). Eligible patients regardless of PD-L1 expression received neoadjuvant camrelizumab 200 mg and platinum-doublet chemotherapy every 3 weeks (arm A) or those with PD-L1-positive tumors received neoadjuvant camrelizumab and apatinib 250 mg once daily (arm B), for 2–4 cycles, followed by surgery. The primary endpoint was major pathological response (MPR) rate. Thirty patients in arm A and 21 in arm B were enrolled. Surgery rates were 50.0% (15/30) in arm A and 42.9% (9/21) in arm B, with all patients achieving R0 resections. Of these patients, the MPR and pathological complete response rates were both 20.0% (95% CI 4.3–48.1) in arm A and were 55.6% (95% CI 21.2–86.3) and 11.1% (95% CI 0.3–48.2) in arm B, respectively. The corresponding objective response rates were 33.3% (95% CI 11.8–61.6) and 55.6% (95% CI 21.2–86.3). With a median follow-up of 22.4 months (95% CI 19.0–26.0), the median event-free survival was not reached (NR; 95% CI 13.6-NR) in arm A and 16.8 months (95% CI 8.6-NR) in arm B. Grade 3 or above treatment-related adverse events occurred in eight (26.7%) patients in arm A and three (14.3%) in arm B. Biomarker analysis showed baseline TYROBP expression was predictive of treatment response in arm B. Neoadjuvant camrelizumab plus chemotherapy or apatinib exhibits preliminary efficacy and manageable toxicity in patients with initially unresectable stage II–III NSCLC.

Abstract Background The erector spinae plane block (ESPB) is a new analgesic method used in thoracic surgery. However, few studies have characterized their effects on perioperative opioid consumption. We aimed to evaluate the effects of ESPB on perioperative opioid consumption in patients who underwent video-assisted thoracoscopic surgery (VATS). Methods This was a randomized, observer-blinded clinical trial at a single-centre academic hospital. Eighty patients were scheduled for thoracoscopic segmentectomy or lobectomy by VATS for lung cancer. Forty participants were randomly assigned to ESPB or control group. All patients received intravenous patient-controlled postoperative analgesia. Perioperative opioid consumption, visual analogue scale (VAS) scores, and adverse events were recorded. Results Intraoperative and postoperative opioid consumption and static/dynamic VAS scores were significantly lower in the early hours after VATS in the ESPB group (p

Abstract Understanding the regulation of normal erythroid development will help to develop new potential therapeutic strategies for disorders of the erythroid lineage. Cellular repressor of E1A‐stimulated genes 1 (CREG1) is a glycoprotein that has been implicated in the regulation of tissue homeostasis. However, its role in erythropoiesis remains largely undefined. In this study, it is found that CREG1 expression increases progressively during erythroid differentiation. In zebrafish, creg1 mRNA is preferentially expressed within the intermediate cell mass (ICM)/peripheral blood island (PBI) region where primitive erythropoiesis occurs. Loss of creg1 leads to anemia caused by defective erythroid differentiation and excessive apoptosis of erythroid progenitors. Mechanistically, creg1 deficiency results in reduced activation of TGF‐β/Smad2 signaling pathway. Treatment with an agonist of the Smad2 pathway (IDE2) could significantly restore the defective erythroid development in creg1−/− mutants. Further, Klf1, identified as a key target gene downstream of the TGF‐β/Smad2 signaling pathway, is involved in creg1 deficiency‐induced aberrant erythropoiesis. Thus, this study reveals a previously unrecognized role for Creg1 as a critical regulator of erythropoiesis, mediated at least in part by the TGF‐β/Smad2‐Klf1 axis. This finding may contribute to the understanding of normal erythropoiesis and the pathogenesis of erythroid disorders.

BackgroundLead exposure levels are closely linked to human health and can cause damage to multiple organ systems, including the blood system and liver. However, due to insufficient evidence, the effects of lead exposure on hematological and biochemical indices have not been fully established.ObjectiveThis study aims to explore the blood lead levels of permanent residents in Jiangxi Province and analyze the factors affecting blood lead levels and the impact of blood lead levels on hematological and biochemical indices.MethodsWe conducted a cross-sectional study including questionnaires, health examinations, and blood sample examinations on 720 randomly selected permanent residents (3–79 years) in Jiangxi Province in 2018. The blood lead levels were measured using inductively coupled plasma mass spectrometry. Routine hematological and biochemical tests were determined by qualified medical institutions using automated hematology analyzers and biochemistry analyzers.ResultsThe geometric mean of blood lead concentration in permanent residents of Jiangxi Province was 20.45 μg/L. Gender, age, annual household income, smoking, and hypertension were the influencing factors for blood lead levels. For each 1 μg/L increase in blood lead, the risks of elevated red blood cell count (from low to high), platelet volume distribution width, alkaline phosphatase (from low to high), and cholesterol increased by 2.4, 1.6, 3.6, and 2.3%, respectively, whereas the risks of elevation of direct bilirubin and total bilirubin both decreased by 1.7%.ConclusionThe blood lead level in permanent residents of Jiangxi Province is higher than the national average. Higher blood lead levels were found in men than in women; blood lead levels were positively correlated with age but negatively correlated with annual household income; smoking and hypertension are risk factors for elevated blood lead; and blood lead levels affect routine hematological and biochemical markers such as red blood cell count, platelet volume distribution width, direct bilirubin, total bilirubin, alkaline phosphatase, and cholesterol.

Abstract Objectives The right lower sleeve lobectomy is a rarely performed major lung resection.This study aims to evaluate the safety and effectiveness of this procedure by comparing to right lower bilobectomy in non-small cell lung cancer patients. Methods We retrospectively reviewed a prospective database of non-small cell lung cancer patients who underwent right lower sleeve lobectomy (group S) or right lower bilobectomy (group B) from January 2014 to January 2020 in Shanghai Pulmonary Hospital. Propensity score matching method was applied to balance confounders between the two groups, resulting in 41 matched pairs.The analysis was performed to compare perioperative outcomes, long-term survival, and postoperative pulmonary volume between the two groups. Results No significant differences in the characteristics were observed between the two matched groups.Major postoperative complications developed in 31.7% of the patients in group B and 12.1% of the patients in group S (P = 0.032).Intervention rate for surgical residual cavity in group B is significantly higher than those patients in group S(21.9%vs7.3%,p = 0.037).The postoperative right lateral and overall lung volume in group S were both significantly larger than that in group B (P = 0.026,P = 0.001,respectively). Conclusions Compared to bi-lobectomy, a middle lobe sparing sleeve resection obtains a less prevalence of major complications, smaller postoperative residual air space and similar long-term survival for selected central right lower NSCLC patients.

Abstract The Illumina HiSeq platform has been a commonly used option for bacterial genome sequencing. Now the BGI DNA nanoball (DNB) nanoarrays platform may provide an alternative platform for sequencing of bacterial genomes. To explore the impact of sequencing platforms on bacterial genome assembly, quality assessment, sequence alignment, functional annotation, mutation detection, and metagenome mapping, we compared genome assemblies based on sequencing of cultured bacterial species using the HiSeq 2000 and BGISEQ-500 platforms. In addition, simulated reads were used to evaluate the impact of insert size on genome assembly. Genome assemblies based on BGISEQ-500 sequencing exhibited higher completeness and fewer N bases in high GC genomes, whereas HiSeq 2000 assemblies exhibited higher N50. The majority of assembly assessment parameters, sequences of 16S rRNA genes and genomes, numbers of single nucleotide variants (SNV), and mapping to metagenome data did not differ significantly between platforms. More insertions were detected in HiSeq 2000 genome assemblies, whereas more deletions were detected in BGISEQ-500 genome assemblies. Insert size had no significant impact on genome assembly. Taken together, our results suggest that DNBSEQ platforms would be a valid substitute for HiSeq 2000 for bacterial genome sequencing.

Abstract Background Postoperative pain remains a significant concern following uniportal thoracoscopic surgery. The analgesic efficacy of erector spinae plane block (ESPB) and serratus anterior plane block (SAPB) in terms of postoperative opioid consumption in uniportal thoracoscopic surgery still needs further studies. Methods A randomized controlled trial was conducted, enrolling 150 patients who underwent uniportal thoracoscopic lobectomy. The patients were randomly allocated to three groups in a 1:1:1 ratio: the ESPB group (administered 20 ml of 0.5% ropivacaine), the SAPB group (administered 20 ml of 0.5% ropivacaine), and the standard care (control) group. The primary endpoint was the consumption of sufentanil during the first 24 h following surgery. Secondary endpoints assessed the area under the curve (AUC) of pain numerical rating scale (NRS) scores, occurrence of moderate to severe pain, time to initial sufentanil request, and postoperative adverse events. Results No significant difference was observed in the consumption of sufentanil during the first 24 h following surgery between the ESPB and SAPB groups (adjusted difference, 1.53 [95% CI, -5.15 to 2.08]). However, in comparison to the control group, both intervention groups demonstrated a significant decrease in sufentanil consumption, with adjusted differences of -9.97 [95% CI, -13.10 to -6.84] for the ESPB group and -12.55 [95% CI, -15.63 to -9.47] for the SAPB group. There were no significant differences in AUC of NRS scores during rest and movement between the ESPB and SAPB groups, with adjusted differences of -7.10 [95% CI, 1.33 to -15.55] for the rest condition and 5.61 [95% CI, -13.23 to 2.01] for the movement condition. At 6 h postoperatively, there were fewer patients with moderate to severe pain in the ESPB group compared with those in the SAPB group (adjusted difference, -1.37% [95% CI, -2.29% to -0.45%]. The time to first sufentanil request significantly differed among the three groups (ESPB vs Control P

IntroductionFaecalibacterium is one of the most abundant bacteria in the gut microbiota of healthy adults, highly regarded as a next-generation probiotic. However, the functions of Faecalibacterium genomes from cultured strains and the distribution of different species in populations may differ among different sources.MethodsWe here performed an extensive analysis of pan-genomes, functions, and safety evaluation of 136 Faecalibacterium genomes collected from 10 countries.ResultsThe genomes are clustered into 11 clusters, with only five of them were characterized and validly nomenclated. Over 80% of the accessory genes and unique genes of Faecalibacterium are found with unknown function, which reflects the importance of expanding the collection of Faecalibacterium strains. All the genomes have the potential to produce acetic acid and butyric acid. Nine clusters of Faecalibacterium are found significantly enriched in the healthy individuals compared with patients with type II diabetes..DiscussionThis study provides a comprehensive view of genomic characteristic and functions and of culturable Faecalibacterium bacterium from human gut, and enables clinical advances in the future.

Introduction The subtransverse process interligamentary (STIL) plane block is an emerging interfascial plane block that has garnered attention for its potential to provide effective postoperative analgesia for breast and thoracic surgeries. However, a direct comparative assessment between the STIL plane block and the paravertebral block is currently lacking. Consequently, our study aims to assess the analgesic efficacy of the STIL block in comparison to paravertebral block for patients undergoing video-assisted thoracoscopic surgery (VATS).Methods and analysis This study is a randomised, parallel-controlled, double-blind, non-inferiority trial, with the goal of enrolling 114 participants scheduled for uniportal VATS at Shanghai Pulmonary Hospital. Participants will be randomly assigned in a 1:1 ratio through block randomisation to receive either the STIL plane block (n=57) or the paravertebral block (n=57). The primary outcome of the study is the area under the curve of Numerical Rating Scale(NRS) scores recorded over a 48-hour period following the surgical procedure. Secondary outcomes encompass the evaluation of Quality of Recovery-40, cumulative sufentanil consumption, serum inflammatory factors, rescue medication usage, the incidence of adverse events and the patient satisfaction scores.Ethics and dissemination This study has received approval from the Medical Ethics Committee of Shanghai Pulmonary Hospital (approval no. L22-329). Written informed consent will be obtained from all participants. The findings will be submitted for publication in peer-reviewed journals.Trial registration number ChiCTR2200066909.

The genus Bifidobacterium widely exists in human gut and has been increasingly used as the adjuvant probiotics for the prevention and treatment of diseases. However, the functional differences of Bifidobacterium genomes from different regions of the world remain unclear. We here describe an extensive study on the genomic characteristics and function annotations of 1512 genomes (clustered to 849 non-redundant genomes) of Bifidobacterium cultured from human gut. The distribution of some carbohydrate-active enzymes varied among different Bifidobacterium species and continents. More than 36% of the genomes of B. pseudocatenulatum harbored biosynthetic gene clusters of lanthipeptide-class-iv. 99.76% of the cultivated genomes of Bifidobacterium harbored genes of bile salt hydrolase. Most genomes of B. adolescentis, and all genomes of B. dentium harbored genes involved in gamma-aminobutyric acid synthesis. B. longum subsp. infantis were characterized harboring most genes related to human milk oligosaccharide utilization. Significant differences between the distribution of antibiotic resistance genes among different species and continents revealed the importance to use antibiotics precisely in the clinical treatment. Phages infecting Bifidobacterium and horizontal gene transfers occurring in genomes of Bifidobacterium were dependent on species and region sources, and might help Bifidobacterium adapt to the environment. In addition, the distribution of Bifidobacterium in human gut was found varied from different regions of the world. This study represents a comprehensive view of characteristics and functions of genomes of cultivated Bifidobacterium from human gut, and enables clinical advances in the future.

Abstract Afatinib, an irreversible ErbB-family blocker, could improve the survival of advanced epidermal growth factor receptor (EGFR)-mutant non-small cell lung cancer patients (NSCLCm+). This phase II trial (NCT04201756) aimed to assess the feasibility of neoadjuvant Afatinib treatment for stage III NSCLCm+. Forty-seven patients received neoadjuvant Afatinib treatment (40 mg daily). The primary endpoint was objective response rate (ORR). Secondary endpoints included pathological complete response (pCR) rate, pathological downstaging rate, margin-free resection (R0) rate, event-free survival, disease-free survival, progression-free survival, overall survival, treatment-related adverse events (TRAEs). The ORR was 70.2% (95% CI: 56.5% to 84.0%), meeting the pre-specified endpoint. The major pathological response (MPR), pCR, pathological downstaging, and R0 rates were 9.1%, 3.0%, 57.6%, and 87.9%, respectively. The median survivals were not reached. The most common TRAEs were diarrhea (78.7%) and rash (78.7%). Only three patients experienced grade 3/4 TRAEs. Biomarker analysis and tumor microenvironment dynamics by bulk RNA sequencing were included as predefined exploratory endpoints. CISH expression was a promising marker for Afatinib response (AUC = 0.918). In responders, compared to baseline samples, increasing T-cell- and B-cell-related features were observed in post-treatment tumor and lymph-node samples, respectively. Neoadjuvant Afatinib is feasible for stage III NSCLC+ patients and leads to dynamic changes in the tumor microenvironment.

Abstract The oral cavity harbors highly diverse communities of microorganisms. However, the number of isolated species and high-quality genomes is limited. Here we present a Cultivated Oral Bacteria Genome Reference (COGR), comprising 1089 high-quality genomes based on large-scale aerobic and anaerobic cultivation of human oral bacteria isolated from dental plaques, tongue, and saliva. COGR covers five phyla and contains 195 species-level clusters of which 95 include 315 genomes representing species with no taxonomic annotation. The oral microbiota differs markedly between individuals, with 111 clusters being person-specific. Genes encoding CAZymes are abundant in the genomes of COGR. Members of the Streptococcus genus make up the largest proportion of COGR and many of these harbor entire pathways for quorum sensing important for biofilm formation. Several clusters containing unknown bacteria are enriched in individuals with rheumatoid arthritis, emphasizing the importance of culture-based isolation for characterizing and exploiting oral bacteria.

Abstract Background Prolonged mechanical ventilation (PMV), mostly defined as mechanical ventilation > 72 h after lung transplantation with or without tracheostomy, is associated with increased mortality. Nevertheless, the predictive factors of PMV after lung transplant remain unclear. The present study aimed to develop a novel scoring system to identify PMV after lung transplantation. Methods A total of 141 patients who underwent lung transplantation were investigated in this study. The patients were divided into PMV and non-prolonged ventilation (NPMV) groups. Univariate and multivariate logistic regression analyses were performed to assess factors associated with PMV. A risk nomogram was then established based on the multivariate analysis, and model performance was further examined regarding its calibration, discrimination, and clinical usefulness. Results Eight factors were finally identified to be significantly associated with PMV by the multivariate analysis and therefore were included as risk factors in the nomogram as follows: the body mass index (BMI, P = 0.036); primary diagnosis as idiopathic pulmonary fibrosis (IPF, P = 0.038); pulmonary hypertension (PAH, P = 0.034); primary graft dysfunction grading (PGD, P = 0.011) at T0; cold ischemia time (CIT P = 0.012); and three ventilation parameters (peak inspiratory pressure [PIP, P

Abstract Background This trial aimed to analyse the safety, effectiveness and transcriptomic characteristics of neoadjuvant toripalimab plus chemotherapy in II–III non-small-cell lung cancer (NSCLC). Methods Patient eligibility mainly involved treatment-naive, clinical stage II–III and wild-type EGFR/ALK NSCLC. The patients received 2–4 cycles of toripalimab (240 mg q3w) plus carboplatin-based chemotherapy. After the second treatment cycle, all patients were re-evaluated by a multidisciplinary team. Candidates eligible for surgery underwent surgery; otherwise, patients received the remaining treatment cycles. The primary endpoints were safety and major pathological response (MPR). Secondary endpoints were R0 resection rate, progression-free survival (PFS) and overall survival (OS). RNA sequencing of baseline and post-treatment samples was conducted to explore the transcriptomic characteristics of the therapeutic response. Results In total, 50 eligible patients were enrolled, including 12 (24.0%) with resectable disease (RD) and 38 (76.0%) with potentially resectable disease (PRD). Treatment-related adverse events (TRAEs) were recorded in 48 cases (96.0%). Severe TRAEs occurred in 3 (6.0%) cases, including myelosuppression, drug-induced liver injury and death related to haemoptysis. The objective response rate (ORR) was 76.0%, with 8 (16.0%) patients having a complete response (CR), 30 (60.0%) partial response (PR), 10 (20.0%) stable disease (SD) and 2 (4.0%) progressive disease (PD). Surgery could be achieved in 12 (100%) patients with RD and 25 (65.8%) with PRD; 1 (2.0%) with PRD refused surgery. Therefore, R0 resection was performed for all 36 (100%) patients who underwent surgery; 20 (55.6%) achieved MPR, including 10 (27.8%) with a complete pathological response (pCR). The CHI3L1 (chitinase-3-like protein 1) immunohistochemistry (IHC) expression of baseline tumour samples could predict the therapeutic response (AUC=0.732), OS (P=0.017) and PFS (P=0.001). Increased PD-1 expression, T cell abundance and immune-related pathway enrichment were observed in post-treatment samples compared to baseline in the response group (CR+PR) but not in the non-response group (SD+PD). Conclusions Neoadjuvant toripalimab plus chemotherapy was safe and effective, with a high MPR and manageable TRAEs for II–III NSCLC, even converting initially PRD to RD. Disparate transcriptomic characteristics of therapeutic efficiency were observed, and CHI3L1 expression predicted therapeutic response and survival. Trial registration ChiCTR1900024014, June 22, 2019.

Idiopathic pulmonary fibrosis (IPF) is a progressive disease with unknown etiology and limited therapeutic options. Activation of fibroblasts is a prominent feature of pulmonary fibrosis. Here we report that lncRNA DACH1 (dachshund homolog 1) is downregulated in the lungs of IPF patients and in an experimental mouse model of lung fibrosis. LncDACH1 knockout mice develop spontaneous pulmonary fibrosis, whereas overexpression of LncDACH1 attenuated TGF-β1-induced aberrant activation, collagen deposition and differentiation of mouse lung fibroblasts. Similarly, forced expression of LncDACH1 not only prevented bleomycin (BLM)-induced lung fibrosis, but also reversed established lung fibrosis in a BLM model. Mechanistically, LncDACH1 binding to the serine/arginine-rich splicing factor 1 (SRSF1) protein decreases its activity and inhibits the accumulation of Ctnnb1. Enhanced expression of SRSF1 blocked the anti-fibrotic effect of LncDACH1 in lung fibroblasts. Furthermore, loss of LncDACH1 promoted proliferation, differentiation, and extracellular matrix (ECM) deposition in mouse lung fibroblasts, whereas such effects were abolished by silencing of Ctnnb1. In addition, a conserved fragment of LncDACH1 alleviated hyperproliferation, ECM deposition and differentiation of MRC-5 cells driven by TGF-β1. Collectively, LncDACH1 inhibits lung fibrosis by interacting with SRSF1 to suppress CTNNB1 accumulation, suggesting that LncDACH1 might be a potential therapeutic target for pulmonary fibrosis.

Rehabilitation medicine (RM) requires more personalisation than any other treatment in the medical domain. Additive manufacturing (AM) is a promising candidate for providing customised tools to improve performance but reduce cost and complexity of personalisation. Research and applications of AM in the RM field are receiving increasing attention. This paper aims to identify the research status and development trend via visual analysis of literature. At first, core publications from representative domain journals and conference proceedings were extracted as source data for the inputs of statistical analysis. Secondly, keyword visualisation analysis provides research hotspots and technological developments to summarise the research frontiers. Finally, visual knowledge graphs were constructed to show the emerging trends, including digital solutions, lifecycle treatment, and multiple-supporter rehabilitation services, in this special interdisciplinary field. Specially, digital methods with AM and associated services will be one of the principle roadmaps for personalised RM in the future.

BackgroundEpidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) could provide survival benefits for locally advanced EGFR-mutant (EGFRm) non-small cell lung cancer (NSCLC). However, the role of radical surgery for EGFR-TKI treated stage IIIB EGFRm NSCLC remains controversial. This study attempted to assess the feasibility of neoadjuvant EGFR-TKI followed by radical surgery for stage IIIB EGFRm NSCLC.Patients and MethodsBetween 2013 and 2020, EGFRm lung adenocarcinoma (LUAD) patients in clinical stage IIIB undergoing neoadjuvant EGFR-TKI followed by surgery (T-S-Arm) and EGFR-TKI alone (T-Arm) were reviewed retrospectively in Shanghai Pulmonary Hospital (SPH). The chi-square test, Student’s t-test or Fisher’s exact test was performed for analysis of baseline characteristics. Progression-free survival (PFS) was estimated using the Kaplan-Meier analysis. Multivariate Cox regression analysis was used to identify independent predictors of progression.ResultsA total of 43 patients were divided into T-S-Arm (n = 21) and T-Arm (n = 22). Patients were well-balanced between the two arms. The majority of patients were female (n = 25, 58.1%), non-smokers (n = 35, 81.4%), first-generation of EGFR-TKI treatment (n = 39, 90.7%), and exon 19 deletions (19-DEL) (n = 26, 60.5%). The median diagnostic age was 63.0 years [interquartile range (IQR), 54.0-67.5 years). At the cut-off date with June 30th 2022, median follow-up time was 28 months (IQR, 20-39 months). Neoadjuvant EGFR-TKI treatment followed by radical surgery could significantly improve the median PFS compared with patients underwent EGFR-TKI alone (23.0 months vs 14.5 months, P = 0.002). Multivariate Cox regression analysis demonstrated that radical surgery (T-S-Arm vs. T-Arm, HR: 0.406; 95% CI: 0.207-0.793, P = 0.027) was the only independent predictor for disease progression. The stratified analysis demonstrated patients with N2 disease could benefit from radical surgery (HR, 0.258; 95% CI, 0.107-0.618), especially for patients harboring L858R mutation (HR, 0.188; 95% CI, 0.059-0.604).ConclusionsFor stage IIIB EGFRm NSCLC patients, the prognosis might be improved by neoadjuvant EGFR-TKI followed by radical surgery versus EGFR-TKI alone, especially for those with N2 disease and harboring L858R mutation.

Abstract Background Enhancer of zeste homolog 2 (EZH2) is a novel oncogene that can specifically trimethylate the histone H3 lysine 27 (H3K27me3) to transcriptionally inhibit the expression of downstream tumor-suppressing genes. As a small molecular inhibitor of EZH2, 3-Deazaneplanocin (DZNep) has been widely studied due to the role of tumor suppression. With the roles of epigenetic regulation of bone cells emerged in past decades, the property and molecular mechanism of DZNep on enhancing osteogenesis had been reported and attracted a great deal of attention recently. This study aims to elucidate the role of DZNep on EZH2-H3K27me3 axis and downstream factors during both osteoclasts and osteoblasts formation and the therapeutic possibility of DZNep on bone defect healing. Methods Bone marrow-derived macrophages (BMMs) cells were cultured, and their responsiveness to DZNep was evaluated by cell counting kit-8, TRAP staining assay, bone resorption assay, podosome actin belt. Bone marrow-derived mesenchymal stem cells (BMSC) were cultured and their responsiveness to DZNep was evaluated by cell counting kit-8, ALP and AR staining assay. The expression of nuclear factor-κB (NF-κB), mitogen-activated protein kinase (MAPK), Wnt signaling pathway was determined by qPCR and western blotting. Mouse bone defect models were created, rescued by DZNep injection, and the effectiveness was evaluated by X-ray and micro-CT and histological staining. Results Consistent with the previous study that DZNep enhances osteogenesis via Wnt family member 1(Wnt1), Wnt6, and Wnt10a, our results showed that DZNep also promotes osteoblasts differentiation and mineralization through the EZH2-H3K27me3-Wnt4 axis. Furthermore, we identified that DZNep promoted the receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-induced osteoclast formation via facilitating the phosphorylation of IKKα/β, IκB, and subsequently NF-κB nuclear translocation, which credit to the EZH2-H3K27me3-Foxc1 axis. More importantly, the enhanced osteogenesis and osteoclastogenesis result in accelerated mice bone defect healing in vivo. Conclusion DZNep targeting EZH2-H3K27me3 axis facilitated the healing of mice bone defect via simultaneously enhancing osteoclastic bone resorption and promoting osteoblastic bone formation.

BackgroundCeritinib is used for the treatment of patients with anaplastic lymphoma kinase (ALK)-rearranged non-small cell lung cancer (NSCLC), who are at the risk of developing bone metastasis. During bone metastasis, tumor cells release factors that induce osteoclast formation, resulting in osteolysis. However, the effect of ceritinib on osteoclast formation remains unclear.MethodsOsteoclastogenesis was induced to assess the effect of ceritinib on osteoclast formation and osteoclast-specific gene expression. Western blotting was used to examine the molecular mechanisms underlying the effect of ceritinib on osteoclast differentiation. An in vivo ovariectomized mouse model was established to validate the effect of ceritinib in suppressing osteoclast formation and preventing bone loss.ResultsThe differentiation of osteoclasts and the expression of osteoclast-specific genes were inhibited upon ceritinib stimulation. Ceritinib suppressed Akt and p65 phosphorylation during the receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclastogenesis. The administration of ceritinib to ovariectomized mice ameliorated trabecular bone loss by inhibiting osteoclast formation.ConclusionsCeritinib is beneficial in preventing bone loss by suppressing osteoclastic Akt and nuclear factor κB (NF-κB) signaling.

Abstract Long non-coding RNAs (lncRNAs) have emerged as critical factors for regulating multiple biological processes during organ fibrosis. However, the mechanism of lncRNAs in idiopathic pulmonary fibrosis (IPF) remains incompletely understood. In the present study, two sets of lncRNAs were defined: IPF pathogenic lncRNAs and IPF progression lncRNAs. IPF pathogenic and progression lncRNAs-mRNAs co-expression networks were constructed to identify essential lncRNAs. Network analysis revealed a key lncRNA CTD-2528L19.6, which was up-regulated in early-stage IPF compared to normal lung tissue, and subsequently down-regulated during advanced-stage IPF. CTD-2528L19.6 was indicated to regulate fibroblast activation in IPF progression by mediating the expression of fibrosis related genes LRRC8C, DDIT4, THBS1, S100A8 and TLR7 et al. Further studies showed that silencing of CTD-2528L19.6 increases the expression of Fn1 and Collagen I both at mRNA and protein levels, promoted the transition of fibroblasts into myofibroblasts and accelerated the migration and proliferation of MRC-5 cells. In contrast, CTD-2528L19.6 overexpression alleviated fibroblast activation in MRC-5 cells induced by TGF-β1. LncRNA CTD-2528L19.6 inhibited fibroblast activation through regulating the expression of LRRC8C in vitro assays. Our results suggest that CTD-2528L19.6 may prevent the progression of IPF from early-stage and alleviate fibroblast activation during the advanced-stage of IPF. Thus, exploring the regulatory effect of lncRNA CTD-2528L19.6 may provide new sights for the prevention and treatment of IPF.

Abstract Purine rich element binding protein A (Purα), encoded by the Purα gene, is an important transcriptional regulator that binds to DNA and RNA and is involved in processes such as DNA replication and RNA translation. Purα also plays an important role in the nervous system. To identify the function of Pura, we performed RNA sequence (RNA-seq) analysis of Purɑ-KO mouse hippocampal neuron cell line (HT22) to analyze the effect of Purα deletion on neuronal expression profiles. And combined with ChIP-seq analysis to explore the mechanism of Purα on gene regulation. In the end, totaly 656 differentially expressed genes between HT22 and Purα-KO HT22 cells have been found, which include 7 Alzheimer’s disease (AD)-related genes and 5 Aβ clearance related genes. 47 genes were regulated by Purα directly, the evidence based on CHIP-seq, which include Insr, Mapt, Vldlr, Jag1, etc. Our study provides the important informations of Purα in neuro-development. The possible regulative effects of Purα on AD-related genes consist inthe direct and indirect pathways of Purα in the pathogenesis of AD.

Breast cancer bone metastasis poses significant challenge for therapeutic strategies. Inside the metastatic environment, osteoclasts and tumor cells interact synergistically to promote cancer progression. In this study, the proprotein convertase furin is targeted due to its critical roles in both tumor cell invasion and osteoclast function. Importantly, the furin inhibitor is specifically delivered by bone targeting superparamagnetic iron oxide (SPIO) nanoparticles. Our in vitro and in vivo data demonstrate that this system can effectively inhibit both osteoclastic bone resorption and breast cancer invastion, leading to alleviated osteolysis. Therefore, the bone targeting & furin inhibition nanoparticle system is a promising therapeutic and diagnostic strategy for breast cancer bone metastasis.

The land use and land cover pattern of the Qinghai-Tibet Plateau (QTP) is an important basis for the structure and function of the QTP ecological barrier. It is of great significance to simulate the land use pattern and landscape ecological risk of the QTP under future scenarios for the construction of the QTP barrier area and to promote the sustainable use of land resources. The QTP was selected as the study area. Based on the spatial pattern of land use in 2010 and 2020, the PLUS model was used to predict the land use patterns of the QTP in 2030 under the two scenarios of natural development and ecological conservation. The landscape ecological risk index was constructed to evaluate the past, present, and future landscape ecological risk of the QTP. The natural break point method was used to divide the landscape ecological risk index into five levels: lower ecological risk, low ecological risk, medium ecological risk, high ecological risk, and higher ecological risk. The results showed that: (1) Under the natural development scenario, the area of cropland, forestland, grassland, and unused land decreased continuously, while the areas of water and built-up land increased gradually. Under the ecological conservation scenario, the areas of forestland and grassland increased by 130 km2 and 2293 km2, respectively, compared with the natural development scenario. (2) Under the natural development scenario, the overall ecological risk of the QTP increased from 2010 to 2030, which showed that the proportions of lower ecological risk area decreased, while the proportion of medium and high ecological risk area increased. Under the ecological conservation scenario, compared with the natural development scenario, the area of lower, low, and high ecological risk increased by 4044 km2, 2484 km2, and 6401 km2, respectively, while the areas of medium and higher ecological risk decreased by 6333 km2 and 6597 km2, respectively.

Osteolytic bone disorders are characterized by an overall reduction in bone mineral density which enhances bone ductility and vulnerability to fractures. This disorder is primarily associated with superabundant osteoclast formation and bone resorption activity. Nicorandil (NIC) is a vasodilatory anti-anginal drug with ATP-dependent potassium (KATP) channel openings. However, NIC is adopted to manage adverse cardiovascular and coronary events. Recent research has demonstrated that NIC also possesses anti-inflammatory peculiarity through the regulation of p38 MAPK and NF-κB signaling pathways. Both MAPK and NF-κB signaling pathways play pivotal roles in RANKL-induced osteoclast formation and bone resorption function. Herein, we hypothesized that NIC may exert potential biological effects against osteoclasts, and revealed that NIC dose-dependently suppressed bone marrow macrophage (BMM) precursors to differentiate into TRAP + multinucleated osteoclasts in vitro. Furthermore, osteoclast resorption assays demonstrated anti-resorptive effects exhibited by NIC. NIC had no impact on osteoblast differentiation or mineralization function. Based on Biochemical analyses, NIC relieved RANKL-induced ERK, NF-κB and p38 MAPK signaling without noticeable effects on JNK MAPK activation. However, the attenuation of NF-κB and p38 MAPK activation was sufficient to hamper the downstream induction of c-Fos and NFATc1 expression. Meanwhile, NIC administration markedly protected mice from ovariectomy (OVX)-induced bone loss through in vivo inhibition of osteoclast formation and bone resorption activity. Collectively, this work demonstrated the potential of NIC in the management of osteolytic bone disorders mediated by osteoclasts.

Abstract Background Helminths immunomodulate the host immune system by secreting proteins to create an inhibitory environment as a strategy for survival in the host. As a bystander effect, this balances the host immune system to reduce hypersensitivity to allergens or autoantigens. Based on this, helminth therapy has been used to treat some allergic or autoimmune diseases. As a tissue-dwelling helminth, Trichinella spiralis infection has been identified to have strong immunomodulatory effects; the effective components in the worm have not yet been identified. Methods The soluble extracts of T. spiralis adult worms and muscle larvae were used to treat airway inflammation before and after an ovalbumin (OVA)-sensitization/challenge in an OVA-induced asthma mouse model. The therapeutic effects were observed by measuring the level of inflammation in the lungs. Results The soluble products derived from T. spiralis parasites, especially from adult worms, were able to ameliorate OVA-induced airway inflammatory responses which were associated with reduced eosinophil infiltration, OVA-specific IgE, Th2 cytokine IL-4, and increased IL-10 and TGF-β. The stimulation of the Treg response may contribute to the alleviated allergic inflammation. Conclusions Trichinella spiralis worm extracts stimulate regulatory cytokines that are associated with reduced allergic airway inflammation. The identification of effective components in the adult worm extracts will be a crucial approach for developing a novel therapeutic for allergic and autoimmune diseases.

Osteolysis resulting from osteoclast overactivation is one of the severe complications of breast cancer metastasis to the bone. Previous studies reported that the anti-cancer agent DZNep induces cancer cell apoptosis by activating Akt signaling. However, the effect of DZNep on breast cancer bone metastasis is unknown. We previously found that DZNep enhances osteoclast differentiation by activating Akt. Therefore, we explored the use of the anti-cancer agent AZD3463 (an Akt inhibitor) along with DZNep, as AZD3463 can act as an anti-cancer agent and can also potentially ameliorate bone erosion. We evaluated osteoclast and breast cancer cell phenotypes and Akt signaling in vitro by treating cells with DZNep and AZD3463. Furthermore, we developed a breast cancer bone metastasis animal model in mouse tibiae to further determine their combined effects in vivo. Treatment of osteoclast precursor cells with DZNep alone increased osteoclast differentiation, bone resorption, and expression of osteoclast-specific genes. These effects were ameliorated by AZD3463. The combination of DZNep and AZD3463 inhibited breast cancer cell proliferation, colony formation, migration, and invasion. Finally, intraperitoneal injection of DZNep and AZD3463 ameliorated tumor progression and protected against bone loss. In summary, DZNep combined with AZD3463 prevented skeletal complications and inhibited breast cancer progression by suppressing Akt signaling.

The progression of intervertebral disc degeneration (IDD) is multifactorial with the senescence of nucleus pulposus (NP) cells and closely related to inflammation in NP cells. Dehydrocostus lactone (DHE) is a natural sesquiterpene lactone isolated from medicinal plants that has anti-inflammatory properties. Thus, DHE may have a therapeutic effect on the progression of IDD. In this study, NP cells were used to determine the appropriate concentration of DHE in vitro. The role of DHE in tumor necrosis factor-α (TNF-α)–induced activation of inflammatory signaling pathways and cellular senescence, together with anabolism and catabolism of extracellular matrix (ECM) in NP cells, was examined in vitro. The therapeutic effect of DHE in vivo was determined using a spinal instability model of IDD in mice. The TNF-α–induced ECM degradation and the senescence of NP cells were partially attenuated by DHE. Mechanistically, DHE inhibited the activation of NF-κB and MAPK inflammatory signaling pathways and ameliorated the senescence of NP cells caused by the activation of STING-TBK1/NF-κB signaling induced by TNF-α. Furthermore, a spinal instability model in mice demonstrated that DHE treatment could ameliorate progression of IDD. Together, our findings indicate that DHE can alleviate IDD changes and has a potential therapeutic function for the treatment of IDD.

Parallel microdispensing of high-viscous liquid is a fundamental task in many industrial processes. Herein, a smart printing head is developed, including the probe array, the electric control module, the contact force measurement module, and the extra force balance module. The parallel dispensing of high-viscous liquid in nL level is achieved. The interacting effect between probes on the loading process is analyzed too. According to the result, the interacting effect between probes has a strong influence on the loading process. Therefore, the strategy of serial electrical loading and parallel transfer printing is utilized. Finally, the dependency of transfer printing volume on probe size, etc., is experimentally investigated. The volume of the loaded droplet can be controlled by the lifting velocity of the probe array, and the volume of the transferred droplet can be adjusted by the size of the probe instead of the contact force. The advantage of the proposed method is to realize the highly repeatable parallel dispensing of high-viscous liquid with a relatively simple device.

Dravet syndrome (DS) is a refractory epilepsy typically caused by heterozygous mutations of the Scn1a gene, which encodes the voltage-gated sodium channel Nav1.1. Glucagon-like peptide-1 (GLP-1) analogues, effective therapeutic agents for the treatment of diabetes, have recently become attractive treatment modalities for patients with nervous system disease; however, the impact of GLP-1 analogues on DS remains unknown. This study aimed to determine the neuroprotective role of liraglutide in mouse and cell models of Scn1a KO-induced epilepsy. Epileptic susceptibility, behavioral changes, and behavioral seizures were assessed using electroencephalography (EEG), IntelliCage (TSE Systems, Bad Homburg, Germany), and the open field task. Morphological changes in brain tissues were observed using hematoxylin and eosin (HE) and Nissl staining. Expression of apoptosis-related proteins and the mammalian target of rapamycin (mTOR) signaling pathway were determined using immunofluorescence and western blotting in Scn1a KO-induced epileptic mice in vitro. Scn1a KO model cell proliferation was evaluated using the Cell Counting Kit-8 assay, and the effect of liraglutide on cellular apoptosis levels was examined using Annexin V-FITC/PI flow cytometry. Apoptotic signal proteins and mTOR were assessed using reverse transcription - quantitative polymerase chain reaction (RT-qPCR) and western blotting. Our results showed that liraglutide significantly increased mRNA ((0.31 ± 0.04) *10-3 vs. (1.07 ± 0.08) * 10-3, P = 0.0004) and protein (0.10 ± 0.02 vs. 0.27 ± 0.02, P = 0.0006) expression of Scn1a in Scn1a KO-induced epileptic mice. In addition, liraglutide significantly alleviated electroencephalographic seizures, the severity of responses to epileptic seizures (96.53 ± 0.45 % vs. 85.98 ± 1.24 %, P = 0.0003), cognitive dysfunction, and epileptic-related necrotic neurons (9.76 ± 0.91 % vs. 19.65 ± 2.64 %, P = 0.0005) in Scn1a KO-induced epileptic mice. Moreover, liraglutide protected against Scn1a KO-induced apoptosis, which was manifested in the phosphorylation of mTOR (KO+NS: 1.99 ± 0.31 vs. KO+Lira: 0.97 ± 0.18, P = 0.0004), as well as the downregulation of cleaved caspase-3 (KO+NS: 0.49 ± 0.04 vs. KO+Lira: 0.30 ± 0.01, P = 0.0003) and restoration of the imbalance between BAX (KO+NS: 0.90 ± 0.02 vs. KO+Lira: 0.75 ± 0.04, P = 0.0005) and BCL-2 (KO+NS: 0.46 ± 0.02 vs. KO+Lira: 0.61 ± 0.02, P = 0.0006). Collectively, these results show that liraglutide reduces seizure susceptibility and cognitive dysfunction in the mouse model of Dravet syndrome, and exerts anti-apoptotic and neuroprotective effects in Scn1a KO mice and cells.

Background and objective As computed tomography (CT) screening for lung cancer becomes more common in China, so too does detection of pulmonary ground-glass nodules (GGNs). Although anumber of national or international guidelines about pulmonary GGNs have been published,most of these guidelines are produced by respiratory, oncology or radiology physicians, who might not fully understand the progress of modern minimal invasive thoracic surgery, and these current guidelines may overlook or underestimate the value of thoracic surgery in the management of pulmonary GGNs. In addition, the management for pre-invasive adenocarcinoma is still controversial. Based onthe available literature and experience from Shanghai Pulmonary Hospital, we composed this consensus about diagnosis and treatment of pulmonary GGNs. For lesions which are considered as adenocarcinoma in situ, chest thin layer CT scan follow-up is recommended and resection can only be adopt in some specific cases and excision should not exceed single segment resection. For lesions which are considered as minimal invasive adenocarcinoma, limited pulmonary resection or lobectomy is recommended. For lesions which are considered as early stage invasive adenocarcinoma, pulmonary resection is recommend and optimal surgical methods depend on whether ground glass component exist, location, volume and number of the lesions and physical status of patients. Principle of management of multiple pulmonary nodules is that primary lesions should be handled with priority, with secondary lesions taking into account.

Abstract Background Sepsis is a life-threatening complication of an infection and remains one of the leading causes of mortality in surgical patients. Bacteremia induces excessive inflammatory responses that result in multiple organ damage. Chronic helminth infection and helminth-derived materials have been found to immunomodulate host immune system to reduce inflammation against some allergic or inflammatory diseases. Schistosoma japonicum cystatin (Sj-Cys) is a cysteine protease inhibitor that induces regulatory T-cells and a potential immunomodulatory. The effect of Sj-Cys on reducing sepsis inflammation and mortality was investigated. Methods Sepsis was induced in BALB/c mice using cecal ligation and puncture (CLP), followed by intraperitoneal injection of different doses (10, 25 or 50 μg) of recombinant Sj-Cys (rSj-Cys). The therapeutic effect of rSj-Cys on sepsis was evaluated by observing the survival rates of mice for 96 h after CLP and the pathological injury of liver, kidney and lung by measuring the levels of alanine transaminase (ALT), aspartate transaminase (AST), blood urea nitrogen (BUN) and creatinine (Cr) in sera and the tissue sections pathology, and the expression of MyD88 in liver, kidney and lung tissues. The immunological mechanism was investigated by examining pro-inflammatory cytokines (TNF-α, IL-6, IL-1β) and IL-10 and TGF-β1 in mice sera and in culture of macrophages stimulated by lipopolysaccharides (LPS). Results rSj-Cys treatment provided significant therapeutic effects on CLP-induced sepsis in mice demonstrated with increased survival rates, alleviated overall disease severity and tissue injury of liver, kidney and lung. The rSj-Cys conferred therapeutic efficacy was associated with upregualted IL-10 and TGF-β1 cytokines and reduced pro-inflammatory cytokines TNF-α, IL-6, IL-1β. MyD88 expression in liver, kidney and lung tissues of rSj-Cys-treated mice was reduced. In vitro assay with macrophages also showed that rSj-Cys inhibited the release of pro-inflammatory cytokines and mediator nitric oxide (NO) after being stimulated by lipopolysaccharide (LPS). Conclusions The results suggest the anti-inflammatory potential of rSj-Cys as a promising therapeutic agent on sepsis. The immunological mechanism underlying its therapeutic effect may involve the downregulation of pro-inflammatory cytokines and upregulation of IL-10 and TGF-β1 cytokines possibly via downregulation of the TLR adaptor-transducer MyD88 pathway. The findings suggest rSj-Cys is a potential therapeutic agent for sepsis and other inflammatory diseases.

OBJETIVO: A cirurgia torácica vídeo-assistida (CTVA) tem sido uma intervenção de escolha para o tratamento de pneumotórax espontâneo (PS) com bolha pulmonar. Nosso objetivo foi apresentar uma abordagem de CTVA uniportal unilateral para bulectomia bilateral e avaliar sua eficácia terapêutica. MÉTODOS: Entre maio de 2011 e janeiro de 2012, cinco pacientes foram submetidos a bulectomia bilateral por essa abordagem. Todos apresentavam PS bilateral. A TCAR pré-operatória mostrou que todos os pacientes tinham bolhas bilaterais no pulmão apical. As indicações cirúrgicas, os procedimentos de operação e os desfechos foram revisados. RESULTADOS: Todos os pacientes foram submetidos com sucesso a essa abordagem para bulectomia bilateral, sem complicações intraoperatórias. A mediana de tempo para a retirada do dreno torácico foi de 4,2 dias, e a mediana do tempo de hospitalização no pós-operatório foi de 5,2 dias. A mediana de seguimento pós-operatório foi de 11,2 meses. Um paciente teve recidiva de PE do lado esquerdo três semanas após a cirurgia e foi submetido a abrasão pleural. CONCLUSÕES: A bulectomia bilateral utilizando CTVA uniportal combinada com acesso contralateral ao mediastino anterior é tecnicamente confiável e promove desfechos favoráveis para pacientes com PS que desenvolvem bolhas bilaterais no pulmão apical. Entretanto, para a realização desse procedimento cirúrgico, são necessários cirurgiões com experiência em CTVA, instrumentos toracoscópicos longos, entre outras exigências.OBJECTIVE: Video-assisted thoracoscopic surgery (VATS) has been a surgical intervention of choice for the treatment of spontaneous pneumothorax (SP) with lung bulla. Our objective was to introduce a uniportal VATS approach for simultaneous bilateral bullectomy and to evaluate its therapeutic efficacy. METHODS: Between May of 2011 and January of 2012, five patients underwent bilateral bullectomy conducted using this approach. All of the patients presented with bilateral SP. Preoperative HRCT revealed that all of the patients had bilateral apical bullae. We reviewed the surgical indications, surgical procedures, and outcomes. RESULTS: All of the patients were successfully submitted to this approach for bilateral bullectomy, and there were no intraoperative complications. The median time to chest tube removal was 4.2 days, and the median length of the postoperative hospital stay was 5.2 days. The median postoperative follow-up period was 11.2 months. One patient experienced recurrence of left SP three weeks after the surgery and underwent pleural abrasion. CONCLUSIONS: Bilateral bullectomy through uniportal VATS combined with contralateral access to the anterior mediastinum is technically reliable and provides favorable surgical outcomes for patients with bilateral SP who develop bilateral apical bullae. However, among other requirements, this surgical procedure demands that surgeons be experienced in VATS and that the appropriate thoracoscopic instruments are available.

It is still a difficult clinical issue to decide whether a patient is a suitable candidate for a cochlear implant and to plan postoperative rehabilitation, especially for some special cases, such as auditory neuropathy. A partial solution to these problems is to preoperatively evaluate the functional integrity of the auditory neural pathways. For evaluating the strength of phase-locking of auditory neurons, which was not reflected in previous methods using electrically evoked auditory brainstem response (EABR), a new method for recording phase-locking related auditory responses to electrical stimulation, called the electrically evoked frequency-following response (EFFR), was developed and evaluated using guinea pigs. The main objective was to assess feasibility of the method by testing whether the recorded signals reflected auditory neural responses or artifacts. The results showed the following: 1) the recorded signals were evoked by neuron responses rather than by artifact; 2) responses evoked by periodic signals were significantly higher than those evoked by the white noise; 3) the latency of the responses fell in the expected range; 4) the responses decreased significantly after death of the guinea pigs; and 5) the responses decreased significantly when the animal was replaced by an electrical resistance. All of these results suggest the method was valid. Recording obtained using complex tones with a missing fundamental component and using pure tones with various frequencies were consistent with those obtained using acoustic stimulation in previous studies.