Abnormal global alternative RNA splicing in COVID-19 patients.

Viral infections can alter host transcriptomes by manipulating host splicing machinery. Despite intensive transcriptomic studies on SARS-CoV-2, a systematic analysis of alternative splicing (AS) in severe COVID-19 patients remains largely elusive. Here we integrated proteomic and transcriptomic sequencing data to study AS changes in COVID-19 patients. We discovered that RNA splicing is among the major down-regulated proteomic signatures in COVID-19 patients. The transcriptome analysis showed that SARS-CoV-2 infection induces widespread dysregulation of transcript usage and expression, affecting blood coagulation, neutrophil activation, and cytokine production. Notably, CD74 and LRRFIP1 had increased skipping of an exon in COVID-19 patients that disrupts a functional domain, which correlated with reduced antiviral immunity. Furthermore, the dysregulation of transcripts was strongly correlated with clinical severity of COVID-19, and splice-variants may contribute to unexpected therapeutic activity. In summary, our data highlight that a better understanding of the AS landscape may aid in COVID-19 diagnosis and therapy.

Transcriptional and proteomic insights into the host response in fatal COVID-19 cases.

Coronavirus disease 2019 (COVID-19), the global pandemic caused by SARS-CoV-2, has resulted thus far in greater than 933,000 deaths worldwide; yet disease pathogenesis remains unclear. Clinical and immunological features of patients with COVID-19 have highlighted a potential role for changes in immune activity in regulating disease severity. However, little is known about the responses in human lung tissue, the primary site of infection. Here we show that pathways related to neutrophil activation and pulmonary fibrosis are among the major up-regulated transcriptional signatures in lung tissue obtained from patients who died of COVID-19 in Wuhan, China. Strikingly, the viral burden was low in all samples, which suggests that the patient deaths may be related to the host response rather than an active fulminant infection. Examination of the colonic transcriptome of these patients suggested that SARS-CoV-2 impacted host responses even at a site with no obvious pathogenesis. Further proteomics analysis validated our transcriptome findings and identified several key proteins, such as the SARS-CoV-2 entry-associated protease cathepsins B and L and the inflammatory response modulator S100A8/A9, that are highly expressed in fatal cases, revealing potential drug targets for COVID-19.

Clinicopathological Features and Outcomes of Acute Kidney Injury in Critically Ill COVID-19 with Prolonged Disease Course: A Retrospective Cohort.

BACKGROUND: The incidence, severity, and outcomes of AKI in COVID-19 varied in different reports. In patients critically ill with COVID-19, the clinicopathologic characteristics of AKI have not been described in detail. METHODS: This is a retrospective cohort study of 81 patients critically ill with COVID-19 in an intensive care unit. The incidence, etiologies, and outcomes of AKI were analyzed. Pathologic studies were performed in kidney tissues from ten deceased patients with AKI. RESULTS: A total of 41 (50.6%) patients experienced AKI in this study. The median time from illness to AKI was 21.0 (IQR, 9.5-26.0) days. The proportion of Kidney Disease Improving Global Outcomes (KDIGO) stage 1, stage 2, and stage 3 AKI were 26.8%, 31.7%, and 41.5%, respectively. The leading causes of AKI included septic shock (25 of 41, 61.0%), volume insufficiency (eight of 41, 19.5%), and adverse drug effects (five of 41, 12.2%). The risk factors for AKI included age (per 10 years) (HR, 1.83; 95% CI, 1.24 to 2.69; P=0.002) and serum IL-6 level (HR, 1.83; 95% CI, 1.23 to 2.73; P=0.003). KDIGO stage 3 AKI predicted death. Other potential risk factors for death included male sex, elevated D-dimer, serum IL-6 level, and higher Sequential Organ Failure Assessment score. The predominant pathologic finding was acute tubular injury. Nucleic acid tests and immunohistochemistry failed to detect the virus in kidney tissues. CONCLUSIONS: AKI was a common and multifactorial complication in patients critically ill with COVID-19 at the late stage of the disease course. The predominant pathologic finding was acute tubular injury. Older age and higher serum IL-6 level were risk factors of AKI, and KDIGO stage 3 AKI independently predicted death.

Wet- versus dry-suction techniques for endoscopic ultrasound-guided fine-needle aspiration of solid lesions: a multicenter randomized controlled trial.

BACKGROUND: The optimal sampling techniques for endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) remain unclear and have not been standardized. The aim of this study was to compare the wet-suction and dry-suction techniques for sampling solid lesions in the pancreas, mediastinum, and abdomen. METHODS: This was a multicenter, crossover, randomized controlled trial with randomized order of sampling techniques. The 296 consecutive patients underwent EUS-FNA with 22G needles and were randomized in a ratio of 1:1 into two separate groups that received the dry-suction and wet-suction techniques in a different order. The primary outcome was to compare the histological diagnostic accuracy of dry suction and wet suction for malignancy. The secondary outcomes were to compare the cytological diagnostic accuracy and specimen quality. RESULTS: Among the 269 patients with pancreatic (n = 161) and non-pancreatic (n = 108) lesions analyzed, the wet-suction technique had a significantly better histological diagnostic accuracy (84.9 % [95 % confidence interval (CI) 79.9 % - 89.0 %] vs. 73.2 % [95 %CI 67.1 % - 78.7 %]; P = 0.001), higher specimen adequacy (94.8 % vs. 78.8 %; P < 0.001), and less blood contamination (P < 0.001) than the dry-suction technique. In addition, sampling non-pancreatic lesions with two passes of wet suction provided a histological diagnostic accuracy of 91.6 %. CONCLUSIONS: The wet-suction technique in EUS-FNA generates better histological diagnostic accuracy and specimen quality than the dry-suction technique. Furthermore, sampling non-pancreatic lesions with two passes of EUS-FNA with wet suction may provide a definitive histological diagnosis when rapid on-site evaluation is not routinely available.

SOX11: a potentially useful marker in surgical pathology: a systematic analysis of SOX11 expression in epithelial and non-epithelial tumours.

AIMS: SOX11 is known as an essential transcription factor for regulating neurogenesis. Recently, SOX11 has been suggested to be a diagnostic marker and oncogene because of its significant expression in mantle cell lymphoma (MCL). However, SOX11 expression in other tumour types has not yet been extensively studied. METHODS AND RESULTS: Herein, we examined SOX11 expression in 2026 cases of neuroectodermal, germ cell, mesenchymal and epithelial tumours by immunohistochemistry. SOX11 was consistently expressed in all neuroectodermal tumours with neural differentiation, as well as in immature teratomas revealing neurogenesis. Less frequently, SOX11 expression was observed in only 50% of astrocytomas and 24% of malignant peripheral nerve sheath tumours, and was mainly sporadic and weak/intermediate. In epithelial tumours, significant SOX11 expression was identified in 97% of salivary ductal carcinomas (SDCs) and a high percentage of high-grade neuroendocrine carcinomas (NECs), especially the small-cell lung carcinomas (68%), and was absent in most other carcinomas, except for less and/or focal and weak expression in adenocarcinomas from the lung, genital tract and breast, and salivary adenoid cystic carcinomas and epithelial-myoepithelial carcinomas. In mesenchymal tumours, in addition to MCLs, prominent SOX11 expression was observed in 90% of rhabdomyosarcomas and all myxoid/round cell liposarcomas (MRCLs). Less frequent and/or focal and weak expression was observed in lymphoblastic, Burkitt and follicular lymphomas, synovial sarcoma and angiosarcoma. CONCLUSION: SOX11 showed prominent expression in neuroectodermal tumours with neural differentiation, high grade-NEC, SDC, rhabdomyosarcoma and MRCL. The high sensitivity and specificity of SOX11 in SDC and MRCL make it a useful diagnostic marker.

miR-148b-3p functions as a tumor suppressor in GISTs by directly targeting KIT.

BACKGROUND: Gain-of-function mutations and overexpression of KIT are characteristic features of gastrointestinal stromal tumor (GIST). Dysregulation in miRNA expression may lead to KIT overexpression and tumorigenesis. METHODS: miRNA microarray analysis and real-time PCR were used to determine the miRNA expression profiles in a cohort of 69 clinical samples including 50 CD117IHC+/KITmutation GISTs and 19 CD117IHC-/wild-type GISTs. GO enrichment and KEGG pathway analyses were performed to reveal the predicted targets of the dysregulated miRNAs. Of the dysregulated miRNAs whose expression was inversely correlated with that of KIT miRNAs were predicted by bioinformatics analysis and confirmed by luciferase reporter assay. Cell counting kit-8 (CCK-8) and flow cytometry were used to measure the cell proliferation, cycle arrest and apoptosis. Wound healing and transwell assays were used to evaluate migration and invasion. A xenograft BALB/c nude mouse model was applied to investigate the tumorigenesis in vivo. Western blot and qRT-PCR were used to investigate the protein and mRNA levels of KIT and its downstream effectors including ERK, AKT and STAT3. RESULTS: Of the six miRNAs whose expression was inversely correlated with that of KIT, we found that miR-148b-3p was significantly downregulated in the CD117IHC+/KITmutation GIST cohort. This miRNA was subsequently found to inhibit proliferation, migration and invasion of GIST882 cells. Mechanistically, miR-148b-3p was shown to regulate KIT expression through directly binding to the 3'-UTR of the KIT mRNA. Restoration of miR-148b-3p expression in GIST882 cells led to reduced expression of KIT and the downstream effectors proteins ERK, AKT and STAT3. However, overexpression of KIT reversed the inhibitory effect of miR-148b-3p on cell proliferation, migration and invasion. Furthermore, we found that reduced miR-148b-3p expression correlated with poor overall survival (OS) and disease-free survival (DFS) in GIST patients. CONCLUSION: miR-148b-3p functions as an important regulator of KIT expression and a potential prognostic biomarker for GISTs.

FGFR3 induces degradation of BMP type I receptor to regulate skeletal development.

Fibroblast growth factors (FGFs) and their receptors (FGFRs) play significant roles in vertebrate organogenesis and morphogenesis. FGFR3 is a negative regulator of chondrogenesis and multiple mutations with constitutive activity of FGFR3 result in achondroplasia, one of the most common dwarfisms in humans, but the molecular mechanism remains elusive. In this study, we found that chondrocyte-specific deletion of BMP type I receptor a (Bmpr1a) rescued the bone overgrowth phenotype observed in Fgfr3 deficient mice by reducing chondrocyte differentiation. Consistently, using in vitro chondrogenic differentiation assay system, we demonstrated that FGFR3 inhibited BMPR1a-mediated chondrogenic differentiation. Furthermore, we showed that FGFR3 hyper-activation resulted in impaired BMP signaling in chondrocytes of mouse growth plates. We also found that FGFR3 inhibited BMP-2- or constitutively activated BMPR1-induced phosphorylation of Smads through a mechanism independent of its tyrosine kinase activity. We found that FGFR3 facilitates BMPR1a to degradation through Smurf1-mediated ubiquitination pathway. We demonstrated that down-regulation of BMP signaling by BMPR1 inhibitor dorsomorphin led to the retardation of chondrogenic differentiation, which mimics the effect of FGF-2 on chondrocytes and BMP-2 treatment partially rescued the retarded growth of cultured bone rudiments from thanatophoric dysplasia type II mice. Our findings reveal that FGFR3 promotes the degradation of BMPR1a, which plays an important role in the pathogenesis of FGFR3-related skeletal dysplasia.

Hyperhomocysteinemia regulated SCF expression in cultured cardiomyocytes via modulation of NF-κB activities.

Hyperhomocysteinemia (HHcy) is an important, independent risk factor for coronary artery disease, especially for the myocardial infarction. Our previous study has shown that myocardial stem cell factor (SCF) mediated cardiac stem cells migration, which was involved in cardiac repair. However, it is not clear regarding the action of HHcy on the expression of SCF in cardiomyocytes. In the present study, cultured neonatal rat cardiomyocytes were treated with 20, 50, or 100 µM homocysteine (Hcy) for 5 h. Results showed an significantly increase of SCF expression with 20-50 µM Hcy incubation, which matched with elevated nuclear factor-kappaB (NF-κB) activities. Treatment with NF-κB inhibitor N-acetylcysteine significantly inhibited the increase of SCF. Nevertheless, 100 µM Hcy markedly decreased the expression of SCF, which was in accordance with the suppression of NF-κB activities. The present study indicated that HHcy regulated the expression of SCF in a concentration-dependent manner via modulation of NF-κB activities. Thus, HHcy may increase the risk for cardiovascular diseases not only by causing endothelial dysfunction but also by directly exerting detrimental effects on cardiomyocytes.

The effect of erythropoiesis­stimulating agents on lung cancer patients: a meta­analysis.

Previous studies have demonstrated that erythropoiesis-stimulating agents (ESAs) can reduce anemia and improve quality of life in cancer patients, but ESAs may increase mortality. Therefore, we conducted a meta-analysis of randomized controlled trials (RCT) comparing the effect and risk of ESAs about the prevention or treatment of anemia in cancer patients. Four databases including PubMed, Embase, Web of science and Cochrane Library were searched for published RCTS on ESAs in the treatment of anemia in lung cancer patients from 2000 to 2023. Endpoints including mortality, incidence of thrombotic vascular events, blood transfusion requirement, and incidence of adverse events. Our meta-analysis included 8 studies, with a sample size of 4240 patients, including 2548 patients in the ESAs group and 1692 patients in the control group. The risk of mortality was lower in patients using ESAs than control group (RR 0.96, 95% CI 0.92-0.99, P = 0.02). But there was no significant difference in the risk of mortality between the patients using ESAs and controls (RR 0.99, 95% CI 0.92-1.06, P = 0.69) after removing Pere 2020. Subgroup analysis found that patients diagnosed with small cell lung cancer (SCLC) (RR 1.00, 95% CI 0.92-1.08, P = 0.16) or non-small cell lung cancer (NSCLC) (RR 1.01, 95% CI 0.87-1.17, P = 0.13) were no significant difference in mortality rate. The thrombotic vascular events increase in patients using ESAs than control group (RR 1.40, 95% CI 1.13-1.72, P = 0.002). The blood transfusion requirement of ESAs group was lower than control group (RR 0.56, 95% CI 0.44-0.72, P < 0.00001). And the subgroups of Darbepoetin alfa (RR 0.57, 95% CI 0.41-0.79, P = 0.003) and Epoetin alfa (RR 0.68, 95% CI 0.47-0.99, P = 0.01) had lower transfusion requirements than the control group. In the SCLC subgroup (RR 0.51, 95% CI 0.40-0.65, P = 0.34), blood transfusion requirements were lower in the ESAs group, but there was no significant difference between the subgroup of patients with NSCLC (RR 0.61, 95% CI 0.36-1.04, P = 0.009). There was no statistically significant difference between the two groups in the incidence of adverse reactions (RR 0.98, 95% CI 0.95-1.00, P = 0.10). In conclusion, ESAs does not increase the mortality of lung cancer patients or may reduce the risk of death, and can reduce the need for blood transfusion, although ESA can increase the incidence of thrombotic vascular adverse events.Registration PROSPERO CRD42023463582.

Proteomic Stratification of Prognosis and Treatment Options for Small Cell Lung Cancer.

Small cell lung cancer (SCLC) is a highly malignant and heterogeneous cancer with limited therapeutic options and prognosis prediction models. Here, we analyzed formalin-fixed, paraffin-embedded (FFPE) samples of surgical resections by proteomic profiling, and stratified SCLC into three proteomic subtypes (S-I, S-II, and S-III) with distinct clinical outcomes and chemotherapy responses. The proteomic subtyping was an independent prognostic factor and performed better than current tumor-node-metastasis or Veterans Administration Lung Study Group staging methods. The subtyping results could be further validated using FFPE biopsy samples from an independent cohort, extending the analysis to both surgical and biopsy samples. The signatures of the S-II subtype in particular suggested potential benefits from immunotherapy. Differentially overexpressed proteins in S-III, the worst prognostic subtype, allowed us to nominate potential therapeutic targets, indicating that patient selection may bring new hope for previously failed clinical trials. Finally, analysis of an independent cohort of SCLC patients who had received immunotherapy validated the prediction that the S-II patients had better progression-free survival and overall survival after first-line immunotherapy. Collectively, our study provides the rationale for future clinical investigations to validate the current findings for more accurate prognosis prediction and precise treatments.

Roles of I(f) and intracellular Ca2+ release in spontaneous activity of ventricular cardiomyocytes during murine embryonic development.

In recent years, the contribution of I(f), an important pacemaker current, and intracellular Ca(2+) release (ICR) from sarcoplasmic reticulum to pacemaking and arrhythmia has been intensively studied. However, their functional roles in embryonic heart remain uncertain. Using patch clamp, Ca(2+) imaging, and RT-PCR, we found that I(f) regulated the firing rate in early and late stage embryonic ventricular cells, as ivabradine (30 µM), a specific blocker of I(f), slowed down action potential (AP) frequency. This inhibitory effect was even stronger in late stage cells, though I(f) was down-regulated. In contrast to I(f), ICR was found to be indispensable for the occurrence of APs in ventricular cells of different stages, because abolishment of ICR with ryanodine and 2-aminoethoxydiphenyl borate (2-APB), specific blockers of ryanodine receptors (RyRs) and inositol trisphosphate receptors (IP3Rs), completely abolished APs. In addition, we noticed that RyR- and IP3R-mediated ICR coexisted in early-stage ventricular cells and RyRs functionally dominated. While at late stage RyRs, but not IP3Rs, mediated ICR. In both early and late stage ventricular cells, Na-Ca exchanger current (I(Na/Ca)) mediated ICR-triggered depolarization of membrane potential and resulted in the initiation of APs. We also observed that different from I(f), which presented as the substantial component of the earlier diastolic depolarization current, application of ryanodine, and/or 2-APB slowed the late phase of diastolic depolarization. Thus, we conclude that in murine embryonic ventricular cells I(f) regulates firing rate, while RyRs and IP3Rs (early stage) or RyRs (late stage)-mediated ICR determines the occurrence of APs.

Thermal ablation for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism.

Many studies have been conducted on splenic thermal ablation for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism. In this article, we review the evolution and current status of radiofrequency and microwave ablation in the treatment of spleen diseases. All publications from 1990 to 2011 on radiofrequency and microwave ablation for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism were retrieved by searching PubMed. Thermal ablation in the spleen for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism can preserve part of the spleen and maintain splenic immunologic function. Thermal ablation for assisting hemostasis in partial splenectomy minimizes blood loss during operation. Thermal ablation for spleen trauma reduces the number of splenectomy and the amount of blood transfusion. Thermal ablation for splenic metastasis is minimally invasive and can be done under the guidance of an ultrasound, which helps shorten the recovery time. Thermal ablation for hypersplenism increases platelet (PLT) and white blood cell (WBC) counts and improves liver function. It also helps to maintain splenic immunologic function and even improves splenic immunologic function in the short-term. In conclusion, thermal ablative approaches are promising for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism. In order to improve therapeutic effects, directions for future studies may include standardized therapeutic indications, prolonged observation periods and enlarged sample sizes.

A novel TPD52L2-ROS1 gene fusion expanding the molecular alterations in inflammatory myofibroblastic tumor: case report and literature review.

BACKGROUND: Inflammatory myofibroblastic tumor (IMT) is a distinctive tumor composed of spindle cells accompanied by mixed inflammatory cells, and immunohistochemical positivity for ALK (anaplastic lymphoma kinase protein) can be detected in half of IMTs. The diagnosis of ALK-negative IMT could be a challenge. Recently, the fusions of some kinase genes, such as RET, NTRK1, ROS1, etc., are revealed in ALK-negative IMT. CASE PRESENTATION: A 19-year-old woman presented with swelling of the left upper arm. Magnetic resonance imaging (MRI) scan revealed a tumor in the left postbrachium extended to the left axillary, serratus anterior muscle, and latissimus dorsi muscle. Histopathologically, the irregular-circumscribed tumor was composed of dense spindle-shaped cells with eosinophilic abundant cytoplasm and hyalinized mesenchyme in an inflammatory background. Immunohistochemically (IHC), tumor cells were positive for SMA, MDM2, and p16; the cells were negative for desmin, MyoD1, Myogenin, pan-cytokeratin, S100, SOX10, HMB45, Malen-A, CD34, CD31, CD99, and ALK. By RNA-based NGS, a novel fusion between TPD52L2 3' end of exon 1-4 and ROS1 5' end of exon 36-43 was revealed. ROS1 IHC staining was negative. The final diagnosis of IMT with TPD52L2-ROS1 fusion was made. Subsequently, the patient experienced a good clinical response to Crizotinib, and clinical follow-up showed stable disease after 9 months. CONCLUSION: This report expands the spectrum of ROS1 gene rearrangements in the IMT and highlights the importance of molecular analysis of IMT for getting a diagnostic clue and determining potential therapeutic strategies.

PU-Flow: A Point Cloud Upsampling Network With Normalizing Flows.

Point cloud upsampling aims to generate dense point clouds from given sparse ones, which is a challenging task due to the irregular and unordered nature of point sets. To address this issue, we present a novel deep learning-based model, called PU-Flow, which incorporates normalizing flows and weight prediction techniques to produce dense points uniformly distributed on the underlying surface. Specifically, we exploit the invertible characteristics of normalizing flows to transform points between euclidean and latent spaces and formulate the upsampling process as ensemble of neighbouring points in a latent space, where the ensemble weights are adaptively learned from local geometric context. Extensive experiments show that our method is competitive and, in most test cases, it outperforms state-of-the-art methods in terms of reconstruction quality, proximity-to-surface accuracy, and computation efficiency. The source code will be publicly available at https://github.com/unknownue/puflow.

Knowledge structure and emerging trends of cognitive impairment induced by sleep deprivation: A bibliometric analysis based on CiteSpace and VOSviewer from 2000 to 2022.

This paper implements a bibliometric approach to investigate the research hotspots and future research directions in the relevant field literature. It also offers research ideas and methods for preventing and treating cognitive impairment induced by sleep deprivation in the clinical setting. The evolution of various clusters in the field is summarized through Citespace's projection function for keywords in the literature. CiteSpace and Vosviewer are utilized to analyze and visualize the attributes of the articles, including number of publications, citation frequency, country/region, institution, journal, authors, keywords, and references, from the 2280 publications obtained. A total of 2280 publications were collected, with the number of papers and citations in the field continuously increasing year by year. The most influential country in this field is the United States, and the University of Washington is the most influential institution. The most authoritative journal in the field is identified as SLEEP. Sleep deprivation, prefrontal cortex, and performance are the current topics of interest. The article with the strongest citation burst, lasting from 2015 to 2018, is "Sleep Drives Metabolite Clearance from the Adult Brain." The most influential article and co-cited reference, "Neurocognitive Consequences of Sleep Deprivation," highlights that sleep deprivation from various causes may lead to cognitive impairment. Future research should investigate all forms of cognitive impairment resulting from sleep deprivation. The findings of this study will assist researchers in improving their knowledge structure, identifying research hotspots, and revealing future directions in the field.

Engraftment of engineered ES cell-derived cardiomyocytes but not BM cells restores contractile function to the infarcted myocardium.

Cellular cardiomyoplasty is an attractive option for the treatment of severe heart failure. It is, however, still unclear and controversial which is the most promising cell source. Therefore, we investigated and examined the fate and functional impact of bone marrow (BM) cells and embryonic stem cell (ES cell)-derived cardiomyocytes after transplantation into the infarcted mouse heart. This proved particularly challenging for the ES cells, as their enrichment into cardiomyocytes and their long-term engraftment and tumorigenicity are still poorly understood. We generated transgenic ES cells expressing puromycin resistance and enhanced green fluorescent protein cassettes under control of a cardiac-specific promoter. Puromycin selection resulted in a highly purified (>99%) cardiomyocyte population, and the yield of cardiomyocytes increased 6-10-fold because of induction of proliferation on purification. Long-term engraftment (4-5 months) was observed when co-transplanting selected ES cell-derived cardiomyocytes and fibroblasts into the injured heart of syngeneic mice, and no teratoma formation was found (n = 60). Although transplantation of ES cell-derived cardiomyocytes improved heart function, BM cells had no positive effects. Furthermore, no contribution of BM cells to cardiac, endothelial, or smooth muscle neogenesis was detected. Hence, our results demonstrate that ES-based cell therapy is a promising approach for the treatment of impaired myocardial function and provides better results than BM-derived cells.

Prognostic value and non-neuroendocrine role of INSM1 in small cell lung cancer.

BACKGROUND: Small cell lung cancer (SCLC) is a malignant lung neuroendocrine tumor with early metastasis, rapid progression, and poor outcomes. Insulinoma-associated protein 1 (INSM1) has been an excellent marker for neuroendocrine (NE) differentiation and widely used in the diagnosis of NE neoplasms, including SCLC. However, its role beyond NE diagnostic marker remained little reported. METHODS: We examined immunohistochemical expression of INSM1 in 73 surgically resected SCLC, analyzed its prognostic value by Kaplan-Meier method, and investigated clinical-pathological features of INSM1 high SCLC. In vitro, We assessed INSM1 function on glucose intake, tumor migration, and Cisplatin resistance by 2-NBDG glucose uptake fluorescent assay, transwell assay, and ANNEXIN V/PI assay, respectively. In vivo, we evaluated the therapeutic value of metformin on reversing INSM1 induced chemoresistance by BALB/c nude mice xenograft tumor model. RESULTS: High INSM1 expression was correlated with lymph node metastasis (LNM) (p = 0.0005), later TNM stages (p = 0.0003), and predicted poor survival (Log-rank p = 0.038). Multivariate Cox analysis confirmed INSM1 as an independent prognostic factor in SCLC (p = 0.012, HR:3.195, 95%CI:1.288-7.927). Interestingly, LNM was correlated with worse prognosis only in patients received chemotherapy (Log-rank p = 0.027) rather than the others (Log-rank p = 0.40). In patients having LNM and treated with chemotherapy, high INSM1 was correlated with worse clinic outcome (Log-rank p = 0.009). In vitro, overexpression of INSM1 decreased AMPK-α expression as well as glucose intake, promoted tumor cell migration, and limited the apoptosis induced by Cisplatin, which all could be reversed by Metformin. In vivo, INSM1 overexpression also contributed to tumor growth beyond inducing Cisplatin resistance. CONCLUSION: Our finding suggested INSM1 played more role than a NE marker, partly through down-regulating AMPK signal. INSM1 may serve as a novel prognostic marker and therapeutic target in SCLC.

SOX11 is a sensitive and specific marker for pulmonary high-grade neuroendocrine tumors.

BACKGROUND: Synaptophysin (SYN), chromogranin A (CGA), CD56 and insulinoma-associated protein 1 (INSM1) are proposed neuroendocrine (NE) markers used for diagnosis of pulmonary NE tumors. These NE markers have been identified in subsets of non-NE tumors requiring differential diagnosis, thus we sought to explore new NE markers. METHODS: We evaluated the immunohistochemical expression of SOX11, a transcription factor involved in neurogenesis, in pulmonary NE tumors and large cell carcinomas (LCCs). RESULTS: We found that SOX11 showed a sensitivity similar to INSM1 and CGA, and less than SYN and CD56 in small cell lung carcinomas (SCLCs) and large cell neuroendocrine carcinomas (LCNECs). While SOX11 is more specific than the other four markers for diagnosis of high-grade neuroendocrine carcinomas (HG-NECs) because 1) None of LCCs (0/63), the most challenging non-NE tumor type for differential diagnosis due to overlapped morphology with LCNECs displayed SOX11 positivity. While expression of at least one of SYN, CGA, CD56 or INSM1 was identified in approximately 60% (18/30) of LCCs. 2) SOX11 was only expressed in 1 of 37 carcinoid tumors in contrast to diffuse expression of SYN, CGA, CD56 and INSM1. In HG-NECs, we noticed that SOX11 was a good complementary marker for SCLC diagnosis as it was positive in 7 of 18 SYN-/CGA-/CD56- SCLCs and 3 of 8 SYN-/CGA-/CD56-/INSM1- SCLCs, and SOX11 positivity in 4 of 6 SYN-/CGA-/CD56- cases previously diagnosed as LCCs with NE morphology provides additional evidence of NE differentiation for reclassification into LCNECs, which was further confirmed by electromicroscopical identification of neurosecretory granules. We also found SOX11 expression cannot predict the prognosis in patients with HG-NECs. CONCLUSIONS: Therefore, SOX11 is a useful complementary transcriptional NE marker for diagnosis and differential diagnosis of SCLC and LCNEC.

INSM1 Expression in Mesenchymal Tumors and Its Clinicopathological Significance.

Background: Insulinoma-associated protein 1 (INSM1) has been identified as a nuclear marker of neuroendocrine tumors. Although INSM1 appears to be a subtle and specific biomarker for neuroendocrine tumor, its expression and clinicopathological significance in mesenchymal tumors remain unclear. Methods: We analyzed INSM1 mRNA level in GEO database and conducted immunohistological staining to detect the expression of INSM1 on 576 mesenchymal tumors from pathology department of Tongji Hospital. Results: At transcription level, INSM1 expression in AITL (angioimmunoblastic T-cell lymphoma) was higher than their adjacent normal tissues as well as Hodgkin's lymphoma. Moreover, INSM1 expression in well-differentiated liposarcoma (WDLPS) was significantly higher than normal fat (P = 0.014) and dedifferentiated liposarcoma (DDLPS) (P = 0.0248). At protein level, the positive rate of INSM1 in AITL was 18/48 (47.4%), while in DDLPS was 9/20 (45%). INSM1 expression in AITL was significantly higher than Hodgkin's lymphoma (P = 0.008). And INSM1 expression in WDLPS was significantly lower than DDLPS (P = 0.015). Conclusion: The combination of GEO data and immunohistochemistry data indicated that the expression level of INSM1 was higher in AITL compared with normal control, suggesting that INSM1 may be involved in pathogenesis of AITL. The abnormal expression of INSM1 was found in WDLPS, and the positive rate of INSM1 was higher in DDLPS than in WDLPS. INSM1 may be involved in the regulation of liposarcoma development. There were significant differences in the expression of INSM1 between AITL and Hodgkin's lymphoma and WDLPS and DDLPS. These findings may assist in the differential diagnosis of these tumors when common markers are difficult to identify, enriching the diagnostic index system of mesenchymal tumors.