Ten-Year Follow-Up of Lung Cancer Patients with Resected Stage IA Invasive Non-Small Cell Lung Cancer.

OBJECTIVE: The purpose of this study was to assess 10-year follow-up outcomes after surgical resection in patients with stage IA invasive non-small cell lung cancer (NSCLC) based on postoperative pathological diagnosis. METHODS: Patients with stage IA invasive NSCLC who underwent resection between December 2008 and December 2013 were reviewed. Patients were categorized into the pure-ground glass opacity (pGGO), mixed-ground glass opacity (mGGO), and solid groups based on consolidation to tumor ratio (CTR). Postoperative survival and risk of recurrence and developing secondary primary lung cancer were analyzed in each group. RESULTS: Among the 645 stage IA invasive NSCLC, the 10-year overall survival and recurrence-free survival rate was 79.38% and 77.44%, respectively. The 10-year overall survival for pGGO, mGGO, and solid group of patients was 95.08%, 86.21%, and 72.39%, respectively. The respective recurrence-free survival rate was 100%, 89.82%, and 65.83%. Multivariable Cox regression analysis associated tumor size and GGO components with recurrence and younger age, and tumors with GGO components were associated with longer overall survival. The cumulative incidence curve indicated no recurrence of GGO lung cancer ≥ 5 years postoperatively. Our cohort indicated that the number and stations of dissected lymph node did not influence long-term prognosis of IA invasive NSCLC. CONCLUSIONS: Recurrence of invasive stage IA NSCLC with GGO was more prevalent in patients with tumor size >1 cm and CTR > 0.5, occurring within 5 years after surgery. This will provide important evidence for follow-up strategies in these patients.

Clinicopathologic features, concurrent genomic alterations, and clinical outcomes of patients with KRAS G12D mutations in resected lung adenocarcinoma.

BACKGROUND: In light of the ongoing clinical development of KRAS G12D-specific inhibitors, we sought to investigate the clinicopathologic, co-occurring genomic features and outcomes of patients with KRAS G12D-mutant lung adenocarcinoma. METHODS: 3828 patients with completely resected primary lung adenocarcinomas were examined for KRAS mutations between 2008 and 2020. The association between KRAS G12D and clinicopathologic features, molecular profiles, and outcomes was investigated. RESULTS: 65 patients (1.7%) with KRAS G12D-mutant lung adenocarcinoma were identified. KRAS G12D mutation was more frequent in males, former/current smokers, radiologic solid tumors, and invasive mucinous adenocarcinoma. TP53 and STK11 were the two most frequent concomitant mutations in the KRAS G12D group. KRAS G12D mutation did not appear to be a prognostic factor in resected stage I-III lung adenocarcinomas, while KRAS non-G12D mutation was related to worse survival, especially in stage I tumors. KRAS G12D mutations were associated with positive but low (1-49%) PD-L1 expression compared to negative (<1%), while KRAS non-G12D mutation was associated with high PD-L1 expression (≥50%). TP53 co-mutation indicated higher PD-L1 expression, while STK11 co-mutation had a negligible impact on PD-L1 expression. Furthermore, data mining of MSK datasets from cBioPortal revealed that KRAS G12D and SKT11 co-mutation were associated with a diminished response to immunotherapy. CONCLUSIONS: KRAS G12D-mutant lung adenocarcinoma harbored unique clinicopathologic and genomic characteristics. Despite not being prognostic in resected lung adenocarcinoma, KRAS G12D might be a valuable biomarker in combination with certain co-mutations for identifying relevant subgroups of patients that could eventually influence treatment regimens.

Characterization of Heat-labile Uracil-DNA Glycosylase from Oncorhynchus mykiss and its Application for Carry-over Contamination Control in RT-qPCR.

BACKGROUND: Heat-labile uracil-DNA glycosylase (HL-UDG) is commonly employed to eliminate carry-over contamination in DNA amplifications. However, the prevailing HL-UDG is markedly inactivated at 50°C, rendering it unsuitable for specific one-step RT-qPCR protocols utilizing reverse transcriptase at an optimal temperature of 42°C. OBJECTIVE: This study aimed to explore novel HL-UDG with lower inactivation temperature and for recombinant expression. METHODS: The gene encoding an HL-UDG was cloned from the cold-water fish rainbow trout (Oncorhynchus mykiss) and expressed in Escherichia coli with high yield. The thermostability of this enzyme and other enzymatic characteristics were thoroughly examined. The novel HL-UDG was then applied for controlling carry-over contamination in one-step RT-qPCR. RESULTS: This recombinantly expressed truncated HL-UDG of rainbow trout (OmUDG) exhibited high amino acids similarity (84.1% identity) to recombinant Atlantic cod UDG (rcUDG) and was easily denatured at 40°C. The optimal pH of OmUDG was 8.0, and the optimal concentrations of both Na+ and K+ were 10 mM. Since its inactivation temperature was lower than that of rcUDG, the OmUDG could be used to eliminate carry-over contamination in one-step RT-qPCR with moderate reverse transcription temperature. CONCLUSION: We successfully identified and recombinantly expressed a novel HL-UDG with an inactivation temperature of 40°C. It is suitable for eliminating carry-over contamination in one-step RT-qPCR.

The paracrine isthmin1 transcriptionally regulated by C/EBPß exacerbates pulmonary vascular leakage in murine sepsis.

It is known that pulmonary vascular leakage, a key pathological feature of sepsis-induced lung injury, is largely regulated by perivascular cells. However, the underlying mechanisms have not been fully uncovered. In the present study, we aimed to evaluate the role of isthmin1, a secretory protein originating from alveolar epithelium, in the pulmonary vascular leakage during sepsis and to investigate the regulatory mechanisms of isthmin1 gene transcription. We observed an elevated isthmin1 gene expression in the pulmonary tissue of septic mice induced by cecal ligation and puncture (CLP), as well as in primary murine alveolar type II epithelial cells (ATII) exposed to lipopolysaccharide (LPS). Furthermore, we confirmed that isthmin1 derived from ATII contributes to pulmonary vascular leakage during sepsis. Specifically, adenovirus-mediated isthmin1 disruption in ATII led to a significant attenuation of the increased pulmonary microvascular endothelial cell (PMVEC) hyperpermeability in a PMVEC/ATII coculture system when exposed to LPS. In addition, adeno-associated virus 9 (AAV9)-mediated knockdown of isthmin1 in the alveolar epithelium of septic mice significantly attenuated pulmonary vascular leakage. Finally, mechanistic studies unveiled that nuclear transcription factor CCAAT/enhancer binding protein (C/EBP)ß participates in isthmin1 gene activation by binding directly to the cis-regulatory element of isthmin1 locus and may contribute to isthmin1 upregulation during sepsis. Collectively, the present study highlighted the impact of the paracrine protein isthmin1, derived from ATII, on the exacerbation of pulmonary vascular permeability in sepsis and revealed a new regulatory mechanism for isthmin1 gene transcription.NEW & NOTEWORTHY This article addresses the role of the alveolar epithelial-secreted protein isthmin1 on the exacerbation of pulmonary vascular permeability in sepsis and identified nuclear factor CCAAT/enhancer binding protein (C/EBP)ß as a new regulator of isthmin1 gene transcription. Targeting the C/EBPß-isthmin1 regulatory axis on the alveolar side would be of great value in the treatment of pulmonary vascular leakage and lung injury induced by sepsis.

Cerebral Lactate Participates in Hypoxia-induced Anapyrexia Through its Receptor G Protein-coupled Receptor 81.

Hypoxia-induced anapyrexia is thought to be a regulated decrease in body core temperature (Tcore), but the underlying mechanism remains unclear. Recent evidence suggests that lactate, a glycolysis product, could modulate neuronal excitability through the G protein-coupled receptor 81 (GPR81). The present study aims to elucidate the role of central lactate and GPR81 in a rat model of hypoxia-induced anapyrexia. The findings revealed that hypoxia (11.1% O2, 2 h) led to an increase in lactate in cerebrospinal fluid (CSF) and a decrease in Tcore. Injection of dichloroacetate (DCA, 5 mg/kg, 1 µL), a lactate production inhibitor, to the third ventricle (3 V), alleviated the increase in CSF lactate and the decrease in Tcore under hypoxia. Immunofluorescence staining showed GPR81 was expressed in the preoptic area of hypothalamus (PO/AH), the physiological thermoregulation integration center. Under normoxia, injection of GPR81 agonist 3-chloro-5-hydroxybenzoic acid (CHBA, 0.05 mg/kg, 1 µL) to the 3 V, reduced Tcore significantly. In addition, hypoxia led to a dramatic increase in tail skin temperature and a decrease in interscapular brown adipose tissue skin temperature. The number of c-Fos+ cells in the PO/AH increased after exposure to 11.1% O2 for 2 h, but administration of DCA to the 3 V blunted this response. Injection of CHBA to the 3 V also increased the number of c-Fos+ cells in the PO/AH under normoxia. In light of these, our research has uncovered the pivotal role of central lactate-GPR81 signaling in anapyrexia, thereby providing novel insights into the mechanism of hypoxia-induced anapyrexia.

Value of adjuvant chemotherapy for patients with pT2N0M0 non-small cell lung cancer receiving radical resection.

BACKGROUND: Associations between adjuvant chemotherapy (ACT) and the improvement in survival for patients with pT2N0M0 non-small cell lung cancer (NSCLC) who received R0 resection remain controversial. This study aimed to evaluate the value of ACT for patients with pT2N0M0 NSCLCs, and to identify the subgroups who could benefit from ACT. METHODS: Multivariable Cox proportional hazards regression models were used to estimate independent prognostic factors. High-risk factor (HRF) included visceral pleural invasion (VPI), lymphovascular invasion (LVI) and poor differentiation/undifferentiated tumors. RESULTS: Of the 899 patients, 277 (30.8%) patients received ACT. More younger patients (p < 0.001) and male patients (p = 0.007) received ACT. With the increase of pathological tumor size (p < 0.001) and the number of HRFs (p < 0.001), there was a significant rise in the proportion of patients receiving ACT. For all patients, ACT could not improve recurrence-free survival (RFS) (p = 0.672) and overall survival (OS) (p = 0.306). For patients with pathological stage IIA or radiological pure-solid tumors, ACT could significantly improve the OS (p = 0.011 and p = 0.037, respectively), and multivariate analysis revealed that ACT was an independent prognostic factor for patients with pathological stage IIA (p = 0.005). ACT could improve the OS significantly in patients with pathological stage IB pure-solid lung adenocarcinoma (LUAD) (p = 0.043). CONCLUSION: ACT was valuable for patients with pathological stage IIA (pT2bN0M0) and patients with radiological pure-solid LUAD of pathological stage IB. A combination of radiological features and pathological subtypes could be helpful when selecting patients with pT2N0M0 NSCLCs for ACT.

NOTCH1 Mutations Predict Superior Outcomes of Immune Checkpoint Blockade in Non-Small Cell Lung Cancer.

Background: NOTCH1 is frequently mutated in non-small cell lung cancer (NSCLC), and also is a poor therapeutic target. It is of clinical importance to investigate the effects of NOTCH1 mutations on anti-tumor immunity and response to immune checkpoint blockade (ICB). Methods: An observational study with targeted sequencing in 963 NSCLC patients at our center were performed (FUSCC cohort). Data of the Cancer Genome Atlas Pan-Lung Cancer study (TCGA cohort) were analyzed, and gene set enrichment analysis (GSEA) was performed. The Samstein et al cohort included 350 patients with advanced NSCLC undergoing genomic profiling with the MSK-IMPACT assay, and receiving at least one dose of ICB therapy. Results: NOTCH1 mutations were more common in smokers and patients with squamous-cell carcinoma (SCC) (all P value <0.05). For patients who did not receive ICB therapy (TCGA cohort), the overall survival (OS) of NOTCH1-mutant and -WT patients were comparable (log-rank P = 0.72), while for patients who received ICB therapy in the Samstein et al cohort, NOTCH1-mutant patients had significantly superior OS than WT patients (log-rank P = 0.041). On multivariate Cox analysis, the predictive value of NOTCH1 mutations reached marginal statistical significance (HR, 0.42; 95% CI, 0.17-1.04; P = 0.059). The median of TMB for NOTCH1-mutant tumors was significantly higher than that for NOTCH1-WT tumors, and GSEA revealed that NOTCH1 mutations manifested various defects in the repair of DNA damage. NOTCH1-mutant tumors displayed an inflamed tumor microenvironment (TME), manifesting as increased PD-L1 expression and tumor-infiltrating CD8+ T cells. Conclusion: NOTCH1 mutations define a molecular subtype of NSCLC, which are more common in smokers and patients with SCC, are characterized with higher TMB, inflamed TME, and display improved survival of ICB therapy for NSCLC patients.

Screening of the Pandemic Response Box identifies anti-microsporidia compounds.

Microsporidia are fungal obligate intracellular pathogens, which infect most animals and cause microsporidiosis. Despite the serious threat that microsporidia pose to humans and agricultural animals, few drugs are available for the treatment and control of microsporidia. To identify novel inhibitors, we took advantage of the model organism Caenorhabditis elegans infected with its natural microsporidian Nematocida parisii. We used this system to screen the Pandemic Response Box, a collection of 400 diverse compounds with known antimicrobial activity. After testing these compounds in a 96-well format at high (100 µM) and low (40 µM) concentrations, we identified four inhibitors that restored the ability of C. elegans to produce progeny in the presence of N. parisii. All four compounds reduced the pathogen load of both N. parisii and Pancytospora epiphaga, a C. elegans-infecting microsporidia related to human-infecting species. One of these compounds, a known inhibitor of a viral protease, MMV1006203, inhibited invasion and prevented the firing of spores. A bis-indole derivative, MMV1593539, decreased spore viability. An albendazole analog, MMV1782387, inhibited proliferation of N. parisii. We tested albendazole as well as 5 other analogs and observed that MMV1782387 was amongst the strongest inhibitors of N. parisii and displayed the least host toxicity. Our study further demonstrates the effectiveness of the C. elegans-N. parisii system for discovering microsporidia inhibitors and the compounds we identified provide potential scaffolds for anti-microsporidia drug development.

Droplet manipulation on an adjustable closed-open digital microfluidic system utilizing asymmetric EWOD.

The closed-open digital microfluidic (DMF) system offers a versatile and powerful platform for various applications by combining the advantages of both closed and open structures. The current closed-open DMF system faces challenges in scaling up due to electrode structural differences between closed and open regions. Here we developed an adjustable closed-open DMF platform by utilizing the modified slippery liquid-infused porous surfaces (SLIPS) with asymmetric electrowetting on dielectric (AEWOD) as a hydrophobic dielectric layer. The consistent electrode structures of the bottom printed circuit board (PCB) electrode array on both the closed and open regions, and the utilization of a transparent acrylic with floating potential as the top plate allow a low-cost and easily scalable closed-open DMF system to be achieved. The impacts of applied voltage, parallel plate spacing, electrode switching interval, and electrode driving strategies on various droplet manipulations were investigated. The results show that the optimal plate spacings range from 340-510 µm within the closed region. Meanwhile, we also studied the influence of the thickness, geometry, and position of the top plate on the droplet movement at the closed-open boundary. Through force analysis and experimentation, it is found that a thin top plate and a bevel of ∼4° can effectively facilitate the movement of droplets at the boundary. Finally, we successfully achieved protein staining experiments on this platform and developed a customized smartphone application for the accurate detection of protein concentration. This innovative closed-open DMF system provides new possibilities for future applications in real-time biological sample processing and detection.

Cold stress causes liver damage by inducing ferroptosis through the p38 MAPK/Drp1 pathway.

Acute extreme cold exposure impairs human health and even causes hypothermia which threatens human life. Liver, as a hub in metabolism and thermogenesis, is vital for cold acclimatization. Although accumulating evidence has suggested that cold exposure can cause liver damage, the underlying mechanisms remain poorly understood. This study investigated the role and underlying mechanisms of ferroptosis in cold stress-induced liver damage. To evaluate the role of ferroptosis in cold stress-induced liver damage, rats were pretreated with ferroptosis inhibitor liproxstatin-1 (Lip-1) before exposed to -10 °C for 8 h. Core body temperature was recorded. The levels of ferroptosis-related indicators were examined with the corresponding assay kits or by western blotting. Hepatic pathological changes were analyzed by hematoxylin-eosin staining and ultrastructural observation. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were measured to assess liver function. Rats were also pretreated with p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 or Dynamin-related protein 1 (Drp1) inhibitor Mdivi-1 to determine the underlying mechanisms. We found that Lip-1 inhibited ferroptosis, attenuated hepatic pathological damages and blocked the increased ALT and AST levels in cold-exposed rats. Moreover, Mdivi-1 inhibited mitochondrial fission and suppressed ferroptosis. Furthermore, SB203580 and Mdivi-1 administration alleviated cold stress-induced liver injury. Our results suggested that cold stress caused liver damage partially by inducing ferroptosis through the p38 MAPK/Drp1 pathway. These findings might provide an effective preventive and therapeutic target for cold stress-induced liver injury.

Germination of Microsporidian Spores: The Known and Unknown.

Microsporidia are a large group of mysterious obligate intracellular eukaryotic parasites. The microsporidian spore can survive in the absence of nutrients for years under harsh conditions and germinate within seconds under the stimulation of environmental changes like pH and ions. During germination, microsporidia experience an increase in intrasporal osmotic pressure, which leads to an influx of water into the spore, followed by swelling of the polaroplasts and posterior vacuole, which eventually fires the polar filament (PF). Infectious sporoplasm was transported through the extruded polar tube (PT) and delivered into the host cell. Despite much that has been learned about the germination of microsporidia, there are still several major questions that remain unanswered, including: (i) There is still a lack of knowledge about the signaling pathways involved in spore germination. (ii) The germination of spores is not well understood in terms of its specific energetics. (iii) Limited understanding of how spores germinate and how the nucleus and membranes are rearranged during germination. (iv) Only a few proteins in the invasion organelles have been identified; many more are likely undiscovered. This review summarizes the major resolved and unresolved issues concerning the process of microsporidian spore germination.

[Acute hypoxia blunts cold sensitivity through the inhibition of the lateral parabrachial nucleus in rats].

To explore the changes of cold sensitivity after exposure to acute hypoxia and its mechanisms, Sprague-Dawley rats were divided into normoxia control group (21% O2, 25 °C), 10% O2 hypoxia group (10% O2, 25 °C), 7% O2 hypoxia group (7% O2, 25 °C), normoxia cold group (21% O2, 10 °C) and hypoxia cold group (7% O2, 10 °C). Cold foot withdrawal latency and preference temperature of each group were measured, skin temperatures were estimated using an infrared thermographic imaging camera, body core temperature was recorded by wireless telemetry system, immunohistochemical staining was used to detect the expression of c-Fos in the lateral parabrachial nucleus (LPB). The results showed that acute hypoxia significantly prolonged the latency of cold foot withdrawal and significantly enhanced the intensity of cold stimulation for foot withdrawal, and the rats under hypoxia preferred cold temperature. Cold exposure (10 °C) for 1 h significantly enhanced the expression of c-Fos in LPB of rats in normoxia, while hypoxia inhibited cold-induced c-Fos expression. Acute hypoxia significantly increased the skin temperature of feet and tails, decreased the skin temperature of interscapular region, and decreased the body core temperature of rats. These results indicate that acute hypoxia can significantly blunt cold sensitivity through the inhibition of LPB, suggesting actively keeping warm measures should be taken at the early stage after ascent to high altitude to prevent the upper respiratory infection and acute mountain sickness.

Selective Mediastinal Lymph Node Dissection Strategy for Clinical T1N0 Invasive Lung Cancer: A Prospective, Multicenter, Clinical Trial.

INTRODUCTION: We aimed to prospectively evaluate our previously proposed selective mediastinal lymph node (LN) dissection strategy for peripheral clinical T1N0 invasive NSCLC. METHODS: This is a multicenter, prospective clinical trial in China. We set six criteria for predicting negative LN stations and finally guiding selective LN dissection. Consolidation tumor ratio less than or equal to 0.5, segment location, lepidic-predominant adenocarcinoma (LPA), negative hilar nodes (stations 10-12), and negative visceral pleural invasion (VPI) were used separately or in combination as predictors of negative LN status in the whole, superior, or inferior mediastinal zone. LPA, hilar node involvement, and VPI were diagnosed intraoperatively. All patients actually underwent systematic mediastinal LN dissection. The primary end point was the accuracy of the strategy in predicting LN involvement. If LN metastasis occurred in certain mediastinal zone that was predicted to be negative, it was considered as an "inaccurate" case. RESULTS: A total of 720 patients were enrolled. The median number of LN dissected was 15 (interquartile range: 11-20). All negative node status in certain mediastinal zone was correctly predicted by the strategy. Compared with final pathologic findings, the accuracy of frozen section to diagnose LPA, VPI, and hilar node metastasis was 94.0%, 98.9%, and 99.6%, respectively. Inaccurate intraoperative diagnosis of LPA, VPI, or hilar node metastasis did not lead to inaccurate prediction of node-negative status. CONCLUSIONS: This is the first prospective trial validating the specific mediastinal LN metastasis pattern in cT1N0 invasive NSCLC, which provides important evidence for clinical applications of selective LN dissection strategy.

Activation of brain lactate receptor GPR81 aggravates exercise-induced central fatigue.

Exercise-induced fatigue is a complex physiological phenomenon and is greatly influenced by central mechanisms in brain. As one of the most abundant circulating carbon metabolites, l-lactate in brain has been considered to be an important supplementary fuel during exercise; however, whether it plays a signaling role in fatigue remains largely obscure. In this study, our results initially revealed that brain l-lactate levels were increased after an exhaustive swimming session in several brain regions including motor cortex, hippocampus, and cerebellum. Then, we examined the specific role of brain lactate receptor, also known as hydroxycarboxylic acid receptor 1 (GPR81), in exercise-induced fatigue. We found that intracerebroventricular injection of either d-lactate (an enantiomer that could mediate activation of GPR81 as l-lactate) or a potent GPR81 agonist 3-chloro-5-hydroxybenzoic acid (CHBA), significantly decreased the swimming time to fatigue. After being subjected to the same weight-loaded swimming for 30 min, no obvious changes of blood lactate levels, gastrocnemius pAMPK/AMPK ratio, and glycogen contents were observed between intracerebroventricular CHBA-injected mice and vehicle-treated ones, which suggested a comparable degree of peripheral fatigue. Meanwhile, there were higher extracellular γ-aminobutyric acid (GABA) levels and lower extracellular glutamate levels and glutamate/GABA ratio in motor cortex of the intracerebroventricular CHBA-injected mice than that of vehicle-treated ones, indicating a greater extent of central fatigue in CHBA-injected mice than that in vehicle animals. Collectively, our results suggested that an increased level of brain l-lactate acts as a signaling molecule via activating GPR81, which in turn exacerbates central fatigue during exercise.

Prevalence and genotyping distribution of Enterocytozoon bieneusi in diarrheic pigs in Chongqing and Sichuan provinces, China.

The microsporidian fungal pathogen Enterocytozoon bieneusi is a unicellular parasite that infects humans and various animals, including pigs. Currently, there are few data on E. bieneusi infection a in diarrheic pigs in Chongqing and Sichuan Provinces, China. This study aims to determine the prevalence and genotype distribution of E. bieneusi in diarrheic pigs. In total, 514 fecal samples from diarrheic pigs were obtained from 14 large-scale farms in Chongqing and Sichuan Provinces (326 suckling pigs, 17 weaned pigs, 65 fattening pigs, and 106 sows). To identify the E. bieneusi genotypes, genomic DNA was isolated from the samples and tested by nested PCR, targeting the internal transcribed spacer region of the rRNA followed by DNA sequence analysis. The overall prevalence of E. bieneusi was 79.8% (410/514), with rates of 84.9% (90/106) in sows and 64.7% (11/17) in weaned pigs. We found 61 different genotypes, including seven known genotypes (E, F, CHG1, Peru8, CAF1, B, and BEB17) and 54 novel genotypes. These 54 new genotypes are variants of eight known genotypes (SDD2, A, B, HLJD-IV, PigSpEb1, O, JLD-I, and BEB17) based on their sequence similarities. Phylogenetically, all of the identified genotypes clustered with counterparts belonging to Group 1 and Group 2 of E. bieneusi. Therefore, we found a higher prevalence of E. bieneusi in sows than in preweaned and weaned pigs. These findings indicate that diarrheic pigs could be a potential reservoir host, which can contaminate the environment and be a source of microsporidia in humans and other animals.

Xingnao Kaiqiao Acupuncture Method Combined with Temporal Three-Needle in the Treatment of Acute Ischemic Stroke: A Randomized Controlled Trial.

Objective: The objective of this study was to investigate the difference between the clinical effectiveness of two acupuncture methods in the treatment of acute ischemic stroke (AIS) and provide more evidence-based medical evidence of acupuncture's effectiveness in stroke rehabilitation. Methods: We conducted an outcome assessor-and data analyst-blinded, randomized, and controlled trial. Seventy-two participants were randomly allocated to the observation group and control group with a 1 : 1 allocation ratio by the generating of a random number table.The observation group received the "Xingnao kaiqiao" acupuncture method combined with "Temporal three needles," and the control group received conventional acupuncture "Scalp acupuncture" combined with the traditional "body acupuncture" method. The acupuncture treatment was performed once per day for one week by trained acupuncturists. Both groups underwent secondary prevention of cerebral infarction and received a 3-months' followup. After a 1-week acupuncture intervention, the changes of NIHSS (National Institutes of Health Stroke Scale) scores, Percent Change and Absolute Change of NIHSS scores, MBI (Modified Barthel Index), and the rate of MBI ≥ 80 in two groups were observed. After 3 months' followup, the mRS (Modified Ranking Scale) and the clinical efficacy of the two groups were compared. Results: The apparent efficiency rate of the observation group was 63.9%, higher than 19.4% of the control group, and the difference was significant (P < 0.05). After treatment, NIHSS scores, Percent Change, and Absolute Change of NIHSS scores in the observation group had a significant reduction than the control group (all P < 0.05). MBI in the observation group increased significantly more than in the control group (P < 0.05), but the rate of MBI ≥ 80 in the two groups was not significantly different (P > 0.05). After 3 months' of followup, the mRS score frequencies of the observation group were not statistically different from the control group (P > 0.05). The rate of mRS scores of 0-1 in the observation and control group were 55.6% and 38.9%, and there was no significant difference either (P > 0.05). Conclusion: Compared with "Scalp acupuncture" combined with "body acupuncture," "Xingnao kaiqiao" acupuncture method combined with "Temporal three-needle" had superiority in the improvement of neurological deficit, potential functional disability, and score of basic activities of daily living. As to the independent rate to basic activities of daily living and good prognosis of 3 months, there were no statistical differences.

Loss of TSC1/TSC2 sensitizes immune checkpoint blockade in non-small cell lung cancer.

Tuberous sclerosis complex subunit 1 (TSC1) and 2 (TSC2) are frequently mutated in non-small cell lung cancer (NSCLC), however, their effects on antitumor immunity remained unexplored. A CRISPR screening in murine KrasG12D/Trp53-/- (KP) model identified Tsc1 and Tsc2 as potent regulators of programmed cell death ligand 1 (Pd-l1) expression in vitro and sensitivity to anti-programmed cell death receptor 1 (PD-1) treatment in vivo. TSC1 or TSC2 knockout (KO) promoted the transcriptional and membrane expression of PD-L1 in cell lines. TSC2-deficient tumors manifested an inflamed microenvironment in patient samples and The Cancer Genome Atlas dataset. In syngeneic murine models, KP-Tsc2-KO tumors showed notable response to anti-PD-1 antibody treatment, but Tsc2-wild-type tumors did not. Patients with TSC1/TSC2-mutant NSCLC receiving immune checkpoint blockade (ICB) had increased durable clinical benefit and survival. Collectively, TSC1/TSC2 loss defines a distinct subtype of NSCLC characterized as inflamed tumor microenvironment and superior sensitivity to ICB.

Prognostic implication of tumor spread through air spaces in patients with pathologic N0 lung adenocarcinoma.

OBJECTIVES: Tumor spread through air spaces (STAS) has been identified as an invasive pattern in lung adenocarcinoma (ADC), but the prognostic implication of STAS has not been well studied in patients with pathologic N0 lung ADC. The purpose of this study was to evaluate the prognostic implication of STAS in pathologic N0 lung ADC patients after radical surgery. MATERIALS AND METHODS: Between January 2017 and December 2018, 796 patients with completely resected pathologic N0 lung ADC were reviewed. Pearson's chi-square test or Fisher exact test was used for comparing the relationship between STAS and clinicopathological features. The log-rank test and multivariate Cox regression models were used to explore prognostic factors. RESULTS: Among the 796 patients, STAS was positive in 201 patients (25.3%). The presence of STAS was significantly associated with patients with solid nodules (P < 0.001), micropapillary pattern-predominant adenocarcinoma/solid pattern-predominant adenocarcinoma (P < 0.001), larger tumor size (P < 0.001), visceral pleural invasion (P < 0.001) and lymphovascular invasion (P < 0.001). Multivariable analysis showed that STAS was an independent prognostic factor for recurrence-free survival (RFS) in pathologic N0 lung ADC patients (P = 0.014). For patients with acinar pattern-predominant adenocarcinoma (APA) / papillary pattern-predominant adenocarcinoma (PPA) / invasive mucinous adenocarcinoma (IMA) and patients who underwent lobectomy, STAS was an independent prognostic factor for RFS (P = 0.015, P = 0.011; respectively) and overall survival (OS) (P = 0.038, P = 0.020; respectively). CONCLUSION: In this study, STAS was an independent prognostic factor for RFS in pathologic N0 lung adenocarcinomas, and it was also an independent prognostic factor for RFS and OS in patients with APA/PPA/IMA and those who received lobectomy.

Decreasing Prevalence of Benign Etiology in Resected Lung Nodules Suspicious for Lung Cancer over the Last Decade.

This study investigated treatment strategy for suspicious lung cancer with postoperatively proven benign etiology. In this retrospective study, we collected patients who underwent pulmonary resection for radiologically suspected lung cancer from 2010 to 2019 at Department of Thoracic Surgery, Fudan University Shanghai Cancer Center (FUSCC). Radiological features, preoperative follow-up time, preoperative pathology and postoperative pathology of these patients were documented. We classified resected benign lesions based on paraffin section and compared the therapy management performed on indeterminate lung nodules of 2 time periods (2010-2014 vs 2015-2019). 17,188 patients were included in this cohort and 1,381 (8.03%) cases were postoperatively proved to be benign lesions. Resected benign lesions proportion significantly decreased by years, from 14.5-6.2%. The respective resected benign lesions proportions for pure GGO nodules, part solid nodules and solid nodules were 5.3%, 3.0% and 11.7%. The resected benign lesions rate for patients with longer preoperative follow-up time was much lower (p < 0.001). Among the benign lesions, 14.2% were benign tumors, 25.7% were granulomatous, 30.2% were pneumonia, 18.0% were fibrosis and 11.9% were other types. If we consider that resections for granulomatous and pneumonia radiologically featured as solid nodules exceeding 2 cm, benign tumor and inflammatory pseudotumor are therapeutic, the nontherapeutic pulmonary resection rate is 4.26%. For patients with GGO nodules, the median preoperative follow-up time increased with the time being and the resected benign rate in period 2 (2015-2019) was significantly lower than that in period 1 (2010-2014). Wedge resection was the most common surgery strategy especially for small nodules and no matter for small or large nodules, the frequency of sublobar resection in period 2 was higher than that in period 1. The resected benign lesions rate at our department was relatively low and decreasing over the last decade. Meanwhile, our follow-up and surgical strategy improved over time. For patients with GGO nodules, 4-6months preoperative follow-up is recommended to avoid surgical intervention for benign lesions. For solid nodules with inconclusive diagnosis, limited resection should be first considered to maintain the balance between reducing the risk of cancer progressing and minimizing the resection for benign lesions.

Novel HIF-1-target gene isthmin1 contributes to hypoxia-induced hyperpermeability of pulmonary microvascular endothelial cells monolayers.

Hypoxia-induced pulmonary microvascular endothelial cell (PMVEC) monolayers hyperpermeability is vital for vascular leakage, which participates in vascular diseases, such as acute lung injury (ALI) and high-altitude pulmonary edema (HAPE). We previously observed that PMVEC permeability was markedly elevated in hypoxia when cocultured with primary type II alveolar epithelial cells (AECII) in which isthmin1 (ISM1) was highly upregulated. However, whether the upregulation of ISM1 plays a role in hypoxia-induced PMVEC hyperpermeability is unclear. In this study, we assessed the role of AECII-derived ISM1 in hypoxia-induced PMVEC hyperpermeability with an AECII/PMVEC coculture system and uncovered the underlying mechanism whereby hypoxia stimulates ISM1 gene expression. We found that ISM1 gene expression was upregulated in cultured AECII cells exposed to hypoxia (3% O2) and that AECII-derived ISM1 participated in hypoxia-induced hyperpermeability of PMVEC monolayers, as small interference RNA (siRNA)-mediated knockdown of ISM1 in AECII markedly attenuated the increase in PMVEC permeability in coculture system under hypoxia. In addition, we confirmed that ISM1 was regulated by hypoxia-inducible factor-1α (HIF1α) according to the evidence that silencing of HIF1α inhibited the hypoxia-mediated upregulation of ISM1. Mechanismly, overexpression of HIF1α transcriptionally activated ISM1 gene expression by directly binding to the conserved regulatory elements upstream of the ism1 locus. We identified a novel HIF-1-target gene ISM1, which involves in hyperpermeability of pulmonary microvascular endothelial cell monolayers under hypoxia. Our in vitro cell experiments implied that the upregulated ISM1 derived from alveolar epithelium might be a vital modulator in hypoxia-induced endothelial hyperpermeability and thereby implicates with hypoxic pulmonary-related diseases.