Expansion of tumor-infiltrating lymphocytes in non-small cell lung cancer: Clinical potential and efficacy in EGFR mutation subsets.

Our study aimed to expand tumor-infiltrating lymphocytes (TILs) from primary non-small cell lung cancers (NSCLCs) and evaluate their reactivity against tumor cells. We expanded TILs from 103 primary NSCLCs using histopathological analysis, flow cytometry, IFN-γ release assays, cell-mediated cytotoxicity assays, and in vivo efficacy tests. TIL expansion was observed in all cases, regardless of EGFR mutation status. There was also an increase in the median CD4+/CD8+ ratio during expansion. In post-rapid expansion protocol (REP) TILs, 13 out of 16 cases, including all three cases with EGFR mutations, exhibited a two-fold or greater increase in IFN-γ secretion. The cytotoxicity assay revealed enhanced tumor cell death in three of the seven cases, two of which had EGFR mutations. In vivo functional testing in a patient-derived xenograft model showed a reduction in tumor volume. The anti-tumor activity of post-REP TILs underscores their potential as a therapeutic option for advanced NSCLC, irrespective of mutation status.

Proposal of New ypStage Grouping System for Esophageal Squamous Cell Carcinoma Patients who Underwent Neoadjuvant Chemoradiotherapy followed by Surgery.

OBJECTIVE: To propose a new ypTNM grouping system to address these limitations and improve prognostic relevance. SUMMARY BACKGROUND DATA: The current 8th edition of the American Joint Committee on Cancer (AJCC) ypStage system shows unsatisfactory prognostic relevance in patients with esophageal squamous cell carcinoma (ESCC) treated with neoadjuvant chemoradiotherapy (nCRT) followed by esophagectomy. METHODS: The study cohort included 501 ESCC patients who received nCRT followed by esophagectomy at the Samsung Medical Center in Korea between 1994 and 2018 (development cohort) and 422 patients treated at Asan Medical Center (validation cohort). Recursive partitioning with a tree-structured regression model was used to develop and validate a new ypStage grouping system. RESULTS: In the new ypStage grouping system, ypStage I includes ypT0N0 only; ypStage II includes ypTis-T2N0 or ypT0-T2N1; ypStage III includes ypT3N0-N1; and ypStage IV includes ypT4N0-N1 or ypTanyN2-3. This system adequately addressed the limitations of the existing AJCC classification system, including overlapping and reversal of survival rates. Moreover, the discrimination ability of the new system was higher than that of the existing system [concordance-index (C-index): 61.9%] in the development (C-index: 66.6%) and validation (C-index: 66.0%) cohorts. NRIe was 0.17 [95% confidence interval (CI): 0.09-0.26, P-<0.001) and 0.18 (95% CI: 0.10-0.27, P-<0.001)] in the development and validation cohorts, respectively. CONCLUSIONS: The current study proposes a clear revised version of the 8th edition of the AJCC ypStage grouping system that exhibits superior prognostic stratification in patients with ESCC treated with nCRT followed by esophagectomy.

Control of Memory Phenotype T Lymphocyte Homeostasis: Role of Costimulation.

Foxp3+ T regulatory cells (Tregs), CD4+Foxp3- T cells, and CD8+ T cells are composed of naive phenotype (NP) and memory phenotype (MP) subsets. Ten to 20% of each MP T cell population are cycling (Ki-67+) in vivo. We investigated the contribution of costimulatory (CD28) and coinhibitory (CTLA-4, PD-1) receptors on MP T cell homeostatic proliferation in vivo in the mouse. Blockade of CD28-CD80/CD86 signaling completely abolished MP Tregs and profoundly inhibited MP CD4+Foxp3- T cell proliferation, but it did not affect MP CD8+ T cell proliferation. Marked enhancement of homeostatic proliferation of MP Tregs and MP CD4+Foxp3- T cells was seen after blocking CTLA4-CD80/CD86 interactions and PD-1-PD-L1/2 interactions, and greater enhancement was seen with blockade of both pathways. The CD28 pathway also played an important role in the expansion of Tregs and MP T cells after treatment of mice with agonistic Abs to members of the TNF receptor superfamily, which can act directly (anti-GITR, anti-OX40, anti-4-1BB) or indirectly (anti-CD40) on T cells. Induction of a cytokine storm by blocking the interaction of NK inhibitory receptors with MHC class I had no effect on Treg homeostasis, enhanced MP CD4+ proliferation, and expansion in a CD28-dependent manner, but it enhanced MP CD8+ T cell proliferation in a CD28-independent manner. Because MP T cells exert potent biologic effects primarily before the induction of adaptive immune responses, these findings have important implications for the use of biologic agents designed to suppress autoimmune disease or enhance T effector function in cancer that may have negative effects on MP T cells.

T cell-specific siRNA delivery suppresses HIV-1 infection in humanized mice.

Evaluation of the therapeutic potential of RNAi for HIV infection has been hampered by the challenges of siRNA delivery and lack of suitable animal models. Using a delivery method for T cells, we show that siRNA treatment can dramatically suppress HIV infection. A CD7-specific single-chain antibody was conjugated to oligo-9-arginine peptide (scFvCD7-9R) for T cell-specific siRNA delivery in NOD/SCIDIL2rgamma-/- mice reconstituted with human lymphocytes (Hu-PBL) or CD34+ hematopoietic stem cells (Hu-HSC). In HIV-infected Hu-PBL mice, treatment with anti-CCR5 (viral coreceptor) and antiviral siRNAs complexed to scFvCD7-9R controlled viral replication and prevented the disease-associated CD4 T cell loss. This treatment also suppressed endogenous virus and restored CD4 T cell counts in mice reconstituted with HIV+ peripheral blood mononuclear cells. Moreover, scFvCD7-9R could deliver antiviral siRNAs to naive T cells in Hu-HSC mice and effectively suppress viremia in infected mice. Thus, siRNA therapy for HIV infection appears to be feasible in a preclinical animal model.

Targeted delivery of CRISPR interference system against Fabp4 to white adipocytes ameliorates obesity, inflammation, hepatic steatosis, and insulin resistance.

Obesity is an increasing pathophysiological problem in developed societies. Despite all major progress in understanding molecular mechanisms of obesity, currently available anti-obesity drugs have shown limited efficacy with severe side effects. CRISPR interference (CRISPRi) mechanism based on catalytically dead Cas9 (dCas9) and single guide RNA (sgRNA) was combined with a targeted nonviral gene delivery system to treat obesity and obesity-induced type 2 diabetes. A fusion peptide targeting a vascular and cellular marker of adipose tissue, prohibitin, was developed by conjugation of adipocyte targeting sequence (CKGGRAKDC) to 9-mer arginine (ATS-9R). (dCas9/sgFabp4) + ATS-9R oligoplexes showed effective condensation and selective delivery into mature adipocytes. Targeted delivery of the CRISPRi system against Fabp4 to white adipocytes by ATS-9R induced effective silencing of Fabp4, resulting in reduction of body weight and inflammation and restoration of hepatic steatosis in obese mice. This RNA-guided DNA recognition platform provides a simple and safe approach to regress and treat obesity and obesity-induced metabolic syndromes.

Comparison of Long-Term Outcomes Between Minimally Invasive Pulmonary Resection With and Without Video-Assisted Mediastinoscopic Lymphadenectomy for Left-Sided Lung Cancer.

BACKGROUND: This study aimed to assess the long-term outcomes of video-assisted mediastinoscopic lymphadenectomy (VAMLA) combined with video-assisted thoracic surgery (VATS) for left-sided lung cancer pulmonary resection. PATIENTS AND METHODS: We retrospectively reviewed 1194 consecutive patients who underwent VATS anatomical resection for left-sided lung cancer between January 2007 and December 2016. Using propensity score-based inverse probability of treatment weighting (IPTW), perioperative outcomes and long-term survival outcomes were compared. RESULTS: Among 1194 patients, 295 (24.7%) underwent additional VAMLA (VATS + VAMLA group) and 899 patients (75.3%) underwent VATS only (VATS group). The proportion of patients with advanced N stage were higher in the VATS + VAMLA group (24.7%) than in the VATS group (18.3%). After IPTW adjustment, all baseline profiles between the two groups became similar. The long-term overall survival (OS) and recurrence-free survival (RFS) rates were similar between the VATS + VAMLA group and the VATS group (5-year OS, 77.8% versus 79.3%, p = 0.957; 5-year RFS, 69.6% versus 70.1%, p = 0.498). However, among patients with borderline pulmonary function (FEV1 ≤ 60% or DLCO ≤ 60%), the VATS + VAMLA group (n = 23) had a better prognosis than the VATS group (n = 36) (5-year OS, 67.4% versus 46.7%, respectively; p = 0.047; 5-year RFS, 74.6% versus 53.5%, respectively; p = 0.027). CONCLUSIONS: VAMLA might be a good complement to VATS for left-sided lung cancer, wherein optimal mediastinal lymph node dissection is not feasible under one-lung ventilation, such as when patients have borderline pulmonary function.

Influence of Preoperative Nutritional Status on Patients Who Undergo Upfront Surgery for Esophageal Squamous Cell Carcinoma.

Few studies have focused on preoperative nutritional status of esophageal cancer patients eligible for upfront surgery. We aimed to investigate the association of preoperative nutritional status with prognosis of patients who undergo upfront surgery for esophageal cancer. A total of 274 patients who underwent upfront surgery for esophageal squamous cell carcinoma between January 2012 and December 2016 were eligible. Preoperative nutritional status was evaluated using prognostic nutritional index (PNI) scoring system, nutritional risk screening 2002 (NRS 2002), and controlling nutritional status. The median age was 63 years (interquartile range, 58-70) and 94.7% of patients were male. The pathological stages were Stage I-74.5% (204/274), Stage II-20.4% (56/274), and Stage III-5.1% (14/274). Multivariate analysis revealed that advanced stage, a low PNI, and a high NRS 2002 were independent predictors of overall survival. During median follow-up period of 55 mo, overall survival rates were lower in the high NRS 2002 group (P < 0.001). A high NRS 2002 score was associated with frequent postoperative complications, especially pneumonia and anastomosis site leakage (P = 0.003). The poor preoperative nutritional status with a high NRS 2002 is associated with postoperative complications as well as poor overall survival in patients with upfront surgery for esophageal cancer.Supplemental data for this article is available online at https://doi.org/10.1080/01635581.2022.2042573.

Diagnostic and prognostic values of 2-[18F]FDG PET/CT in resectable thymic epithelial tumour.

OBJECTIVES: We aimed to evaluate the diagnostic ability for the prediction of histologic grades and prognostic values on recurrence and death of pretreatment 2-[18F]FDG PET/CT in patients with resectable thymic epithelial tumours (TETs). METHODS: One hundred and fourteen patients with TETs who underwent pretreatment 2-[18F]FDG PET/CT between 2012 and 2018 were retrospectively evaluated. TETs were classified into three histologic subtypes: low-risk thymoma (LRT, WHO classification A/AB/B1), high-risk thymoma (HRT, B2/B3), and thymic carcinoma (TC). Area under the receiver operating characteristics curve (AUC) was used to assess the diagnostic performance of PET/CT variables (maximum standardised uptake value [SUVmax], metabolic tumour volume [MTV], total lesion glycolysis [TLG], maximum diameter). Cox proportional hazards models were built using PET/CT and clinical variables. RESULTS: The tumours included 52 LRT, 33 HRT, and 29 TC. SUVmax showed good diagnostic ability for differentiating HRT/TC from LRT (AUC 0.84, 95% confidence interval [CI] 0.76 - 0.92) and excellent ability for differentiating TC from LRT/HRT (AUC 0.94, 95% CI 0.90 - 0.98), with significantly higher values than MTV, TLG, and maximum diameter. With an optimal cut-off value of 6.4, the sensitivity, specificity, and accuracy for differentiating TC from LRT/HRT were 69%, 96%, and 89%, respectively. In the multivariable Cox proportional hazards analyses for freedom-from-recurrence, SUVmax was an independent prognostic factor (p < 0.001), whereas MTV and TLG were not. SUVmax was a significant predictor for overall survival in conjunction with clinical stage and resection margin. CONCLUSION: SUVmax showed excellent diagnostic performance for prediction of TC and significant prognostic value in terms of recurrence and survival. KEY POINTS: • Maximum standardised uptake value (SUVmax) shows excellent performance in the differentiation of thymic carcinoma from low- and high-risk thymoma. • SUVmax is an independent prognostic factor for freedom-from-recurrence in the multivariable Cox proportional hazard model and a significant predictor for overall survival. • 2-[18F]FDG PET/CT can provide a useful diagnostic and prognostic imaging biomarker in conjunction with histologic classification and stage and help choose appropriate management for thymic epithelial tumours.

Effects of Multimodal Inpatient Rehabilitation vs Conventional Pulmonary Rehabilitation on Physical Recovery After Esophageal Cancer Surgery.

OBJECTIVE: To determine the effects of multimodal rehabilitation initiated immediately after esophageal cancer surgery on physical recovery compared with conventional pulmonary rehabilitation. DESIGN: Retrospective study. SETTING: Private quaternary care hospital. PARTICIPANTS: Fifty-nine inpatients (N=59) who participated in either conventional pulmonary rehabilitation (n=30) or in multimodal rehabilitation (n=29) after esophageal cancer surgery were included. INTERVENTIONS: Both groups performed pulmonary exercises, including deep breathing, chest expansion, inspiratory muscle training, coughing, and manual vibration. In the conventional pulmonary rehabilitation group, light-intensity mat exercise, stretching, and walking were performed. The multimodal rehabilitation group performed resistance exercises and moderate- to high-intensity aerobic interval exercises using a bicycle. MAIN OUTCOME MEASURES: The European Organization for Research and Treatment of Cancer Core Quality of Life Questionnaire C30 (EORTC QLQ-C30), pain, 6-minute walk test (6MWT), 30-second chair stand test, and grip strengths were assessed before and after the rehabilitation programs. RESULTS: Symptom scales of pain, dyspnea, and insomnia in the EORTC QLQ-C30 as well as 6MWT improved significantly after each program (P<.05). 6MWT (73.1±52.6 vs 28.4±14.3, P<.001, d=1.15), 30-second chair stand test (3.5±3.9 vs 0.35±2.0, P<.001, d=1.06), and left grip strength (1.2±1.3 vs 0.0±1.5, P=.002, d=0.42) improved significantly in the multimodal rehabilitation group compared with the pulmonary rehabilitation group. While right grip strength also showed more improvement for those undergoing the multimodal program, the mean strength difference was not clinically meaningful. CONCLUSIONS: A multimodal inpatient rehabilitation program instituted early after esophageal cancer surgery improved endurance for walking more than conventional pulmonary rehabilitation as measured by the 6MWT and the 30-second chair stand test.

Predicting Postoperative Complications and Long-Term Survival After Lung Cancer Surgery Using Eurolung Risk Score.

BACKGROUND: This study aimed to assess the clinical relevance of the parsimonious Eurolung risk scoring system for predicting postoperative morbidity, mortality, and long-term survival in Korean patients with surgically resected non-small cell lung cancer. METHODS: This retrospective analysis used the data of patients who underwent anatomical resection for non-small cell lung cancer between 2004 and 2018 at a single institution. The parsimonious aggregate Eurolung score was calculated for each patient. The Cox regression model was used to determine the ability of the Eurolung scoring system for predicting long-term outcomes. RESULTS: Of the 7,278 patients in the study, cardiopulmonary complications and mortality occurred in 687 (9.4%) and 53 (0.7%) patients, respectively. The rate of cardiopulmonary complications and mortality gradually increased with the increase in the Eurolung risk scores (all P < 0.001). When risk scores were grouped into four categories, the Eurolung scoring system showed a stepwise deterioration of overall survival with the increase in risk scores, and this association was statistically significant (P < 0.001). Multivariate Cox analysis showed that the Eurolung scoring system, classified into four categories, was a significant prognostic factor of overall survival even after adjusting for covariates such as tumor histology and pathological stage (P < 0.001). CONCLUSION: Stratification based on the parsimonious Eurolung scoring system showed good discriminatory ability for predicting postoperative morbidity, mortality, and long-term survival in South Korean patients with surgically resected non-small cell lung cancer. This might help clinicians to provide a detailed prognosis and decide the appropriate treatment option for high-risk patients with non-small cell lung cancer.

Spinal nerve signal intensity on Dixon T2-weighted water-only sequence: an important outcome predictor after lumbar transforaminal epidural injection.

OBJECTIVES: To evaluate the role of Dixon T2-weighted water-fat separation technique in predicting the outcome of lumbar transforaminal epidural injections (TFESIs). METHODS: Patients who underwent TFESI of a single spinal nerve within 3 months after magnetic resonance imaging (MRI) exam between August 2018 and April 2020 were identified. The patients were classified into positive or negative outcome groups based on the response to the TFESI procedure. Two musculoskeletal radiologists measured the signal intensity of the injected side spinal nerves, contralateral side spinal nerves, and subcutaneous fat on axial Dixon T2-weighted water-only images, and the diameter of spinal nerve on axial Dixon T2-weighted in-phase images of the pre-procedural MRI. The measured values of the injected side spinal nerves were compared between the two groups and with the contralateral side spinal nerve. RESULTS: A total of 94 patients were included, 76 in the positive outcome group and 18 in the negative outcome group. The mean signal intensity and the nerve-to-fat signal ratio of the injected side spinal nerve were significantly higher in the positive outcome group than in the negative outcome group (793.78 vs. 679.19, p = 0.016; 4.21 vs. 3.28, p = 0.003). In the positive outcome group, the diameter of the spinal nerve was significantly higher on the injected side than on the contralateral side (6.91 mm vs. 6.37 mm, p = 0.016). CONCLUSIONS: The mean signal intensity and the nerve-to-fat signal ratio of the spinal nerve on axial Dixon T2-weighted water-only images can help predict patient response to the TFESI. KEY POINTS: • Applying the Dixon technique to lumbar spine MRI can help predict patient response to the TFESI procedure. • An increased nerve-to-fat signal ratio and mean spinal nerve signal intensity on axial Dixon T2-weighted water-only images predicted favorable TFESI outcomes.

Sublobar Resection in Stage IA Non-Small Cell Lung Cancer: Role of Preoperative CT Features in Predicting Pathologic Lymphovascular Invasion and Postoperative Recurrence.

BACKGROUND. Prognostic factors on preoperative CT in stage IA non-small cell lung cancer (NSCLC) may help select patients for sublobar resection or lobectomy. OBJECTIVE. The purpose of this study was to identify CT features predictive of pathologic lymphovascular invasion (LVI) in stage IA NSCLC and to evaluate the features' prognostic value in patients who undergo sublobar resection. METHODS. This retrospective study included 904 patients (mean age, 62.0 years; 453 men, 451 women) who underwent lobectomy (n = 574) or sublobar resection (n = 330) for stage IA NSCLC. Two thoracic radiologists independently evaluated findings on pre-operative chest CT and then resolved discrepancies. Recurrences were identified from medical record review. Multivariable logistic regression was used to identify independent predictors of pathologic LVI. Multivariable Cox proportional hazards models were used to identify prognostic features. Interreader agreement was assessed. RESULTS. Pathologic LVI was present in 10.2% (92/904) of patients. It was present only in solid-dominant part-solid nodules (PSNs) and solid nodules and only in nodules with a solid portion diameter over 10 mm. Among solid-dominant PSNs and solid nodules with a solid portion diameter over 10 mm, independent (p < .05) predictors of pathologic LVI were peritumoral interstitial thickening (odds ratio [OR], 13.22) and pleural contact (defined as pleural contact measuring over one-quarter of the circumference of the nodule's solid portion) (OR, 2.45). Also among such nodules, peritumoral interstitial thickening achieved 80.4% sensitivity, 76.7% specificity, and 77.4% accuracy; pleural contact achieved 35.9% sensitivity, 82.5% specificity, and 74.3% accuracy; and presence of either feature achieved 90.2% sensitivity, 64.3% specificity, and 68.9% accuracy for predicting pathologic LVI. In patients undergoing sublobar resection, after adjusting for T category and operative type, recurrence-free survival (RFS) was independently (p < .05) predicted by solid-dominant PSN or solid nodule with a solid portion diameter over 10 mm also showing peritumoral interstitial thickening (hazard ratio [HR], 5.37) or also showing either peritumoral interstitial thickening or pleural contact (HR, 6.05). The interreader agreement kappa values were 0.67 for peritumoral interstitial thickening and 0.77 for pleural contact. CONCLUSION. Pathologic LVI occurred only in solid-dominant PSNs and solid nodules with solid portion over 10 mm. Among such nodules, peritumoral interstitial thickening and pleural contact independently predicted pathologic LVI and RFS. CLINICAL IMPACT. CT features may help select patients with stage IA NSCLC for sublobar resection rather than more extensive surgery.

Comprehensive genomic profiles of small cell lung cancer.

We have sequenced the genomes of 110 small cell lung cancers (SCLC), one of the deadliest human cancers. In nearly all the tumours analysed we found bi-allelic inactivation of TP53 and RB1, sometimes by complex genomic rearrangements. Two tumours with wild-type RB1 had evidence of chromothripsis leading to overexpression of cyclin D1 (encoded by the CCND1 gene), revealing an alternative mechanism of Rb1 deregulation. Thus, loss of the tumour suppressors TP53 and RB1 is obligatory in SCLC. We discovered somatic genomic rearrangements of TP73 that create an oncogenic version of this gene, TP73Δex2/3. In rare cases, SCLC tumours exhibited kinase gene mutations, providing a possible therapeutic opportunity for individual patients. Finally, we observed inactivating mutations in NOTCH family genes in 25% of human SCLC. Accordingly, activation of Notch signalling in a pre-clinical SCLC mouse model strikingly reduced the number of tumours and extended the survival of the mutant mice. Furthermore, neuroendocrine gene expression was abrogated by Notch activity in SCLC cells. This first comprehensive study of somatic genome alterations in SCLC uncovers several key biological processes and identifies candidate therapeutic targets in this highly lethal form of cancer.

Close Observation versus Additional Surgery after Noncurative Endoscopic Resection of Esophageal Squamous Cell Carcinoma.

INTRODUCTION: After noncurative endoscopic submucosal dissection (ESD) of superficial esophageal squamous cell carcinoma (SESCC), additional esophagectomy is generally recommended. However, considering its high mortality and morbidity, it is uncertain if additional surgery improves the clinical outcomes. This study aimed to compare the clinical outcomes between patients who were observed without additional treatment and those who underwent radical esophagectomy. METHODS: A total of 52 patients with SESCC who underwent complete but noncurative ESD from January 2008 to December 2016 at the Samsung Medical Center and Asan Medical Center in Korea were retrospectively analyzed. Clinicopathologic characteristics and oncologic outcomes were compared between the observation group (n = 23) and the additional surgery group (n = 29). RESULTS: During a mean follow-up of 34.4 and 41.7 months, respectively, the rates of death (observation vs. surgery, 17.4 vs. 10.3%; p = 0.686), recurrence (observation vs. surgery, 13 vs. 17.2%; p = 1.000), and disease-specific death (observation vs. surgery, 4.3 vs. 6.9%; p = 1.000) did not significantly differ between the 2 groups. The 3-year overall survival was 86.3 and 96.4%, respectively (p = 0.776). The 3-year recurrence-free survival (observation vs. surgery, 85.0 vs. 88.7%; p = 0.960) and disease-specific survival (observation vs. surgery, 95.2 vs. 96.4%; p = 0.564) also did not significantly differ. CONCLUSIONS: The clinical outcomes of close observation of noncuratively resected SESCC are comparable to those of additional surgery, at least in the midterm. The wait-and-see strategy could be a feasible management option after noncurative ESD of SESCC in selected patients.

Surgical outcomes of pulmonary metastasectomy in hepatocellular carcinoma patients according to approach method: thoracoscopic versus open approach.

BACKGROUND: The role of surgical intervention as a treatment for pulmonary metastasis (PM) from hepatocellular carcinoma (HCC) has not been established. In this study, we investigated the clinical outcomes of pulmonary metastasectomy. Using propensity score matching (PSM) analysis, we compared the results according to the surgical approach: video-assisted thoracic surgery (VATS) versus the open method. METHODS: A total of 134 patients (115 men) underwent pulmonary metastasectomy for isolated PM of HCC between January 1998 and December 2010 at Seoul Asan Medical Center. Of these, 84 underwent VATS (VATS group) and 50 underwent thoracotomy or sternotomy (open group). PSM analysis between the groups was used to match them based on the baseline characteristics of the patients. RESULTS: During the median follow-up period of 33.4 months (range, 1.8-112.0), 113 patients (84.3%) experienced recurrence, and 100 patients (74.6%) died of disease progression. There were no overall survival rate, disease-free survival rate, and pulmonary-specific disease-free survival rate differences between the VATS and the open groups (p = 0.521, 0.702, and 0.668, respectively). Multivariate analysis revealed local recurrence of HCC, history of liver cirrhosis, and preoperative alpha-fetoprotein level as independent prognostic factors for overall survival (hazard ratio, 1.729/2.495/2.632, 95% confidence interval 1.142-2.619/1.571-3.963/1.554-4.456; p = 0.010/< 0.001/< 0.001, respectively). CONCLUSIONS: Metastasectomy can be considered a potential alternative for selected patients. VATS metastasectomy had outcomes comparable to those of open metastasectomy.

Natural Progression of Ground-glass Nodules after Curative Resection for Non-small Cell Lung Cancer.

BACKGROUND: This retrospective study investigated the natural course of synchronous ground-glass nodules (GGNs) that remained after curative resection for non-small-cell lung cancer (NSCLC). METHODS: Prospectively collected retrospective data were reviewed concerning 2,276 patients who underwent curative resection for NSCLC between 2008 and 2017. High-resolution computed tomography or thin-section computed tomography data of 82 patients were included in the study. Growth in size was considered the most valuable outcome, and patients were grouped according to GGN size change. Patient demographic data (e.g., age, sex, and smoking history), perioperative data (e.g., GGN characteristics, histopathology and pathological stage of the resected tumours), and other medical history were evaluated in a risk factor analysis concerning GGN size change. RESULTS: The median duration of follow-up was 36.0 months (interquartile range, 23.0-59.3 months). GGN size decreased in 6 patients (7.3%), was stationary in 43 patients (52.4%), and increased in 33 patients (40.2%). In univariate analysis, male sex, the GGN size on initial CT, part-solid GGN and smoking history (≥ 10 pack-years) were significant risk factors. Among them, multivariate analysis revealed that lager GGN size, part-solid GGN and smoking history were independent risk factors. CONCLUSION: During follow-up, 40.2% of GGNs increased in size, emphasising that patients with larger GGNs, part-solid GGN or with a smoking history should be observed.

Effective cryopreservation protocol for preservation of male primate (Macaca fascicularis) prepubertal fertility.

RESEARCH QUESTION: Can specimen types (cells versus tissues) and additive cryoprotectant agents contribute to efficient cryopreservation of primate spermatogonial stem cells (SSC)? DESIGN: Testicular tissues or cells from four prepubertal monkeys were used in this study. The freezing efficacy of testicular tissue was compared with cell suspensions using conventional freezing media (1.4 mol/l dimethyl sulfoxide [DMSO]) and the efficacy of cryoprotectant additives (1.4 mol/l DMSO combined with trehalose 200 mmol/l, hypotaurine 14 mmol/l, necrostatin-1 50 µmol/l or melatonin 100 µmol/l) was evaluated in testicular tissue freezing. RESULTS: The survival rate (46.0 ± 4.8% versus 33.7 ± 6.0%; P = 0.0286) and number of recovered cells (5.0 ± 1.5 × 106 cells/g versus 0.7 ± 0.8 × 106 cells/g; P = 0.0286) were significantly higher in frozen tissues than in frozen cell suspensions. After tissue freezing, a higher number of recovered PGP9.5+ cells were observed with 200 mmol/l trehalose treatment than in DMSO controls (2.4 ± 0.6 × 106 cells/g versus 1.1 ± 0.3 × 106 cells/g; P = 0.0164). Normal establishment of donor-derived colony was observed in SSC after tissue freezing with 200 mmol/l trehalose. CONCLUSIONS: Testicular tissue freezing is more effective than single cell suspension freezing for higher recovery of undifferentiated spermatogonia. Moreover, it was verified that slow freezing using 200 mmol/l trehalose, 1.4 mol/l DMSO and 10% KnockOut™ Serum Replacement in Dulbecco's phosphate-buffered saline is an effective cryopreservation protocol for primate testicular tissue.

Gestational Exposure to Bisphenol A Affects Testicular Morphology, Germ Cell Associations, and Functions of Spermatogonial Stem Cells in Male Offspring.

Exposure to bisphenol A (BPA) in the gestational period damages the reproductive health of offspring; detailed evidence regarding BPA-induced damage in testicular germ cells of offspring is still limited. In this study, pregnant mice (F0) were gavaged with three BPA doses (50 µg, 5 mg, and 50 mg/kg body weight (bw)/day; tolerable daily intake (TDI), no-observed-adverse-effect-level (NOAEL), and lowest-observed-adverse-effect level (LOAEL), respectively) on embryonic days 7 to 14, followed by investigation of the transgenerational effects of such exposure in male offspring. We observed that the NOAEL- and LOAEL-exposed F1 offspring had abnormalities in anogenital distance, nipple retention, and pubertal onset (days), together with differences in seminiferous epithelial stages and testis morphology. These effects were eradicated in the next F2 and F3 generations. Moreover, there was an alteration in the ratio of germ cell population and the apoptosis rate in germ cells increased in F1 offspring at the LOAEL dose. However, the total number of spermatogonia remained unchanged. Finally, a reduction in the stemness properties of spermatogonial stem cells in F1 offspring was observed upon LOAEL exposure. Therefore, we provide evidence of BPA-induced disruption of physiology and functions in male germ cells during the gestational period. This may lead to several reproductive health issues and infertility in offspring.

Paternal Exposure to Bisphenol-A Transgenerationally Impairs Testis Morphology, Germ Cell Associations, and Stemness Properties of Mouse Spermatogonial Stem Cells.

Bisphenol-A (BPA) exposure in an adult male can affect the reproductive system, which may also adversely affect the next generation. However, there is a lack of comprehensive data on the BPA-induced disruption of the association and functional characteristics of the testicular germ cells, which the present study sought to investigate. Adult male mice were administered BPA doses by gavage for six consecutive weeks and allowed to breed, producing generations F1-F4. Testis samples from each generation were evaluated for several parameters, including abnormal structure, alterations in germ cell proportions, apoptosis, and loss of functional properties of spermatogonial stem cells (SSCs). We observed that at the lowest-observed-adverse-effect level (LOAEL) dose, the testicular abnormalities and alterations in seminiferous epithelium staging persisted in F0-F2 generations, although a reduced total spermatogonia count was found only in F0. However, abnormalities in the proportions of germ cells were observed until F2. Exposure of the male mice (F0) to BPA alters the morphology of the testis along with the association of germ cells and stemness properties of SSCs, with the effects persisting up to F2. Therefore, we conclude that BPA induces physiological and functional disruption in male germ cells, which may lead to reproductive health issues in the next generation.

Testicular endothelial cells promote self-renewal of spermatogonial stem cells in rats†.

Spermatogonial stem cells (SSCs) are the basis of spermatogenesis in male due to their capability to multiply in numbers by self-renewal and subsequent meiotic processes. However, as SSCs are present in a very small proportion in the testis, in vitro proliferation of undifferentiated SSCs will facilitate the study of germ cell biology. In this study, we investigated the effectiveness of various cell lines as a feeder layer for rat SSCs. Germ cells enriched for SSCs were cultured on feeder layers including SIM mouse embryo-derived thioguanine and ouabain-resistant cells, C166 cells, and mouse and rat testicular endothelial cells (TECs) and their stem cell potential for generating donor-derived colonies and offspring was assessed by transplantation into recipient testes. Rat germ cells cultured on TECs showed increased mRNA and protein levels of undifferentiated spermatogonial markers. Rat SSCs derived from these germ cells underwent spermatogenesis and generated offspring when transplanted into recipients. Collectively, TECs can serve as an effective feeder layer that enhances the proliferative and self-renewal capacity of cultured rat SSCs while preserving their stemness properties.