Kindlin-2 Acts as a Key Mediator of Lung Fibroblast Activation and Pulmonary Fibrosis Progression.

Pulmonary fibrosis is a progressive and fatal lung disease characterized by activation of lung fibroblasts and excessive deposition of collagen matrix. We show here that the concentrations of kindlin-2 and its binding partner PYCR1, a key enzyme for proline synthesis, are significantly increased in the lung tissues of human patients with pulmonary fibrosis. Treatment of human lung fibroblasts with TGF-ß1 markedly increased the expression of kindlin-2 and PYCR1, resulting in increased kindlin-2 mitochondrial translocation, formation of the kindlin-2-PYCR1 complex, and proline synthesis. The concentrations of the kindlin-2-PYCR1 complex and proline synthesis were markedly reduced in response to pirfenidone or nintedanib, two clinically approved therapeutic drugs for pulmonary fibrosis. Furthermore, depletion of kindlin-2 alone was sufficient to suppress TGF-ß1-induced increases of PYCR1 expression, proline synthesis, and fibroblast activation. Finally, using a bleomycin mouse model of pulmonary fibrosis, we show that ablation of kindlin-2 effectively reduced the concentrations of PYCR1, proline, and collagen matrix and alleviate the progression of pulmonary fibrosis in vivo. Our results suggest that kindlin-2 is a key promoter of lung fibroblast activation, collagen matrix synthesis, and pulmonary fibrosis, underscoring the therapeutic potential of targeting the kindlin-2 signaling pathway for control of this deadly lung disease.

Distinct Disease Severity Between Children and Older Adults With Coronavirus Disease 2019 (COVID-19): Impacts of ACE2 Expression, Distribution, and Lung Progenitor Cells.

BACKGROUND: Children and older adults with coronavirus disease 2019 (COVID-19) display a distinct spectrum of disease severity yet the risk factors aren't well understood. We sought to examine the expression pattern of angiotensin-converting enzyme 2 (ACE2), the cell-entry receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and the role of lung progenitor cells in children and older patients. METHODS: We retrospectively analyzed clinical features in a cohort of 299 patients with COVID-19. The expression and distribution of ACE2 and lung progenitor cells were systematically examined using a combination of public single-cell RNA-seq data sets, lung biopsies, and ex vivo infection of lung tissues with SARS-CoV-2 pseudovirus in children and older adults. We also followed up patients who had recovered from COVID-19. RESULTS: Compared with children, older patients (>50 years.) were more likely to develop into serious pneumonia with reduced lymphocytes and aberrant inflammatory response (P = .001). The expression level of ACE2 and lung progenitor cell markers were generally decreased in older patients. Notably, ACE2 positive cells were mainly distributed in the alveolar region, including SFTPC positive cells, but rarely in airway regions in the older adults (P < .01). The follow-up of discharged patients revealed a prolonged recovery from pneumonia in the older (P < .025). CONCLUSIONS: Compared to children, ACE2 positive cells are generally decreased in older adults and mainly presented in the lower pulmonary tract. The lung progenitor cells are also decreased. These risk factors may impact disease severity and recovery from pneumonia caused by SARS-Cov-2 infection in older patients.

Computer-Aided Pathologic Diagnosis of Nasopharyngeal Carcinoma Based on Deep Learning.

The pathologic diagnosis of nasopharyngeal carcinoma (NPC) by different pathologists is often inefficient and inconsistent. We have therefore introduced a deep learning algorithm into this process and compared the performance of the model with that of three pathologists with different levels of experience to demonstrate its clinical value. In this retrospective study, a total of 1970 whole slide images of 731 cases were collected and divided into training, validation, and testing sets. Inception-v3, which is a state-of-the-art convolutional neural network, was trained to classify images into three categories: chronic nasopharyngeal inflammation, lymphoid hyperplasia, and NPC. The mean area under the curve (AUC) of the deep learning model is 0.936 based on the testing set, and its AUCs for the three image categories are 0.905, 0.972, and 0.930, respectively. In the comparison with the three pathologists, the model outperforms the junior and intermediate pathologists, and has only a slightly lower performance than the senior pathologist when considered in terms of accuracy, specificity, sensitivity, AUC, and consistency. To our knowledge, this is the first study about the application of deep learning to NPC pathologic diagnosis. In clinical practice, the deep learning model can potentially assist pathologists by providing a second opinion on their NPC diagnoses.

FLOT2 promotes nasopharyngeal carcinoma progression through suppression of TGF-ß pathway via facilitating CD109 expression.

In nasopharyngeal carcinoma (NPC), the TGF-ß/Smad pathway genes are altered with inactive TGF-ß signal, but the mechanisms remain unclear. RNA-sequencing results showed that FLOT2 negatively regulated the TGF-ß signaling pathway via up-regulating CD109 expression. qRT-PCR, western blot, ChIP, and dual-luciferase assays were used to identify whether STAT3 is the activating transcription factor of CD109. Co-IP immunofluorescence staining assays were used to demonstrate the connection between FLOT2 and STAT3. In vitro and in vivo experiments were used to detect whether CD109 could rescue the functional changes of NPC cells resulting from FLOT2 alteration. IHC and Spearman correlation coefficients were used to assay the correlation between FLOT2 and CD109 expression in NPC tissues. Our results found that FLOT2 promotes the development of NPC by inhibiting TGF-ß signaling pathway via stimulating the expression of CD109 by stabilizing STAT3, which provides a potential therapeutic strategy for NPC treatment.

A Potential "Anti-Warburg Effect" in Circulating Tumor Cell-mediated Metastatic Progression?

Metabolic reprogramming is a defining hallmark of cancer metastasis, warranting thorough exploration. The tumor-promoting function of the "Warburg Effect", marked by escalated glycolysis and restrained mitochondrial activity, is widely acknowledged. Yet, the functional significance of mitochondria-mediated oxidative phosphorylation (OXPHOS) during metastasis remains controversial. Circulating tumor cells (CTCs) are considered metastatic precursors that detach from primary or secondary sites and harbor the potential to seed distant metastases through hematogenous dissemination. A comprehensive metabolic characterization of CTCs faces formidable obstacles, including the isolation of these rare cells from billions of blood cells, coupled with the complexities of ex vivo-culturing of CTC lines or the establishment of CTC-derived xenograft models (CDX). This review summarized the role of the "Warburg Effect" in both tumorigenesis and CTC-mediated metastasis. Intriguingly, bioinformatic analysis of single-CTC transcriptomic studies unveils a potential OXPHOS dominance over Glycolysis signature genes across several important cancer types. From these observations, we postulate a potential "Anti-Warburg Effect" (AWE) in CTCs-a metabolic shift bridging primary tumors and metastases. The observed AWE could be clinically important as they are significantly correlated with therapeutic response in melanoma and prostate patients. Thus, unraveling dynamic metabolic regulations within CTC populations might reveal an additional layer of regulatory complexities of cancer metastasis, providing an avenue for innovative anti-metastasis therapies.

ThermomiR-377-3p-induced suppression of Cirbp expression is required for effective elimination of cancer cells and cancer stem-like cells by hyperthermia.

BACKGROUND: In recent years, the development of adjunctive therapeutic hyperthermia for cancer therapy has received considerable attention. However, the mechanisms underlying hyperthermia resistance are still poorly understood. In this study, we investigated the roles of cold­inducible RNA binding protein (Cirbp) in regulating hyperthermia resistance and underlying mechanisms in nasopharyngeal carcinoma (NPC). METHODS: CCK-8 assay, colony formation assay, tumor sphere formation assay, qRT-PCR, Western blot were employed to examine the effects of hyperthermia (HT), HT + oridonin(Ori) or HT + radiotherapy (RT) on the proliferation and stemness of NPC cells. RNA sequencing was applied to gain differentially expressed genes upon hyperthermia. Gain-of-function and loss-of-function experiments were used to evaluate the effects of RNAi-mediated Cirbp silencing or Cirbp overexpression on the sensitivity or resistance of NPC cells and cancer stem-like cells to hyperthermia by CCK-8 assay, colony formation assay, tumorsphere formation assay and apoptosis assay, and in subcutaneous xenograft animal model. miRNA transient transfection and luciferase reporter assay were used to demonstrate that Cirbp is a direct target of miR-377-3p. The phosphorylation levels of key members in ATM-Chk2 and ATR-Chk1 pathways were detected by Western blot. RESULTS: Our results firstly revealed that hyperthermia significantly attenuated the stemness of NPC cells, while combination treatment of hyperthermia and oridonin dramatically increased the killing effect on NPC cells and cancer stem cell (CSC)­like population. Moreover, hyperthermia substantially improved the sensitivity of radiation­resistant NPC cells and CSC­like cells to radiotherapy. Hyperthermia noticeably suppressed Cirbp expression in NPC cells and xenograft tumor tissues. Furthermore, Cirbp inhibition remarkably boosted anti­tumor­killing activity of hyperthermia against NPC cells and CSC­like cells, whereas ectopic expression of Cirbp compromised tumor­killing effect of hyperthermia on these cells, indicating that Cirbp overexpression induces hyperthermia resistance. ThermomiR-377-3p improved the sensitivity of NPC cells and CSC­like cells to hyperthermia in vitro by directly suppressing Cirbp expression. More importantly, our results displayed the significantly boosted sensitization of tumor xenografts to hyperthermia by Cirbp silencing in vivo, but ectopic expression of Cirbp almost completely counteracted hyperthermia-mediated tumor cell-killing effect against tumor xenografts in vivo. Mechanistically, Cirbp silencing-induced inhibition of DNA damage repair by inactivating ATM-Chk2 and ATR-Chk1 pathways, decrease in stemness and increase in cell death contributed to hyperthermic sensitization; conversely, Cirbp overexpression-induced promotion of DNA damage repair, increase in stemness and decrease in cell apoptosis contributed to hyperthermia resistance. CONCLUSION: Taken together, these findings reveal a previously unrecognized role for Cirbp in positively regulating hyperthermia resistance and suggest that thermomiR-377-3p and its target gene Cirbp represent promising targets for therapeutic hyperthermia.

Loss of cytoplasmic KLF4 expression is correlated with the progression and poor prognosis of nasopharyngeal carcinoma.

AIMS: To examine, in nasopharyngeal carcinoma (NPC), the correlation of Krüppel-like factor 4 (KLF4) expression with clinicopathological features including patient prognosis. METHODS AND RESULTS: Using real-time PCR and immunohistochemistry, expression of KLF4 mRNA and protein was examined in NPC and nasopharyngeal tissues. The relationship of KLF4 expression levels with clinical features and prognosis of NPC patients was analysed. mRNA expression was markedly lower in NPC than in the nasopharyngeal tissues. Using immunohistochemistry, staining for KLF4 protein was found in the nuclei and cytoplasm of nasopharyngeal and malignant epithelial cells, but decreased cytoplasmic expression was observed in atypical hyperplasia and NPC samples compared to normal and squamous epithelium samples (P < 0.001). In addition, levels of cytoplasmic KLF4 protein were correlated inversely with the nodal (N) status (TNM classification; P = 0.002) and overall clinical stage (P < 0.001) of NPC patients. Patients with NPC showing lower cytoplasmic KLF4 expression had a significantly shorter overall survival time than those with high NPC KLF4 expression. Multivariate analysis suggested that the level of KLF4 expression was an independent prognostic indicator (P = 0.008) for NPC survival. CONCLUSION: Low levels of cytoplasmic KLF4 expression are a potentially unfavourable prognostic factor for patients with NPC.

Nasopharyngeal carcinoma ecology theory: cancer as multidimensional spatiotemporal "unity of ecology and evolution" pathological ecosystem.

Nasopharyngeal carcinoma (NPC) is a particular entity of head neck cancer that is generally regarded as a genetic disease with diverse intertumor and intratumor heterogeneity. This perspective review mainly outlines the up-to-date knowledge of cancer ecology and NPC progression, and presents a number of conceptual stepping-stones. At the beginning, I explicitly advocate that the nature of NPC (cancer) is not a genetic disease but an ecological disease: a multidimensional spatiotemporal "unity of ecology and evolution" pathological ecosystem. The hallmarks of cancer is proposed to act as ecological factors of population fitness. Subsequently, NPC cells are described as invasive species and its metastasis as a multidirectional ecological dispersal. The foundational ecological principles include intraspecific relationship (e.g. communication) and interspecific relationship (e.g. competition, predation, parasitism and mutualism) are interpreted to understand NPC progression. "Mulberry-fish-ponds" model can well illustrate the dynamic reciprocity of cancer ecosystem. Tumor-host interface is the ecological transition zone of cancer, and tumor buddings should be recognized as ecological islands separated from the mainland. It should be noted that tumor-host interface has a significantly molecular and functional edge effect because of its curvature and irregularity. Selection driving factors and ecological therapy including hyperthermia for NPC patients, and future perspectives in such field as "ecological pathology", "multidimensional tumoriecology" are also discussed. I advance that "nothing in cancer evolution or ecology makes sense except in the light of the other". The cancer ecology tree is constructed to comprehensively point out the future research direction. Taken together, the establishment of NPC ecology theory and cancer ecology tree might provide a novel conceptual framework and paradigm for our understanding of cancer complex causal process and potential preventive and therapeutic applications for patients.

Deep Multi-Magnification Similarity Learning for Histopathological Image Classification.

Precise classification of histopathological images is crucial to computer-aided diagnosis in clinical practice. Magnification-based learning networks have attracted considerable attention for their ability to improve performance in histopathological classification. However, the fusion of pyramids of histopathological images at different magnifications is an under-explored area. In this paper, we proposed a novel deep multi-magnification similarity learning (DSML) approach that can be useful for the interpretation of multi-magnification learning framework and easy to visualize feature representation from low-dimension (e.g., cell-level) to high-dimension (e.g., tissue-level), which has overcome the difficulty of understanding cross-magnification information propagation. It uses a similarity cross entropy loss function designation to simultaneously learn the similarity of the information among cross-magnifications. In order to verify the effectiveness of DMSL, experiments with different network backbones and different magnification combinations were designed, and its ability to interpret was also investigated through visualization. Our experiments were performed on two different histopathological datasets: a clinical nasopharyngeal carcinoma and a public breast cancer BCSS2021 dataset. The results show that our method achieved outstanding performance in classification with a higher value of area under curve, accuracy, and F-score than other comparable methods. Moreover, the reasons behind multi-magnification effectiveness were discussed.

ESRG is critical to maintain the cell survival and self-renewal/pluripotency of hPSCs by collaborating with MCM2 to suppress p53 pathway.

The mechanisms of self-renewal and pluripotency maintenance of human pluripotent stem cells (hPSCs) have not been fully elucidated, especially for the role of those poorly characterized long noncoding RNAs (lncRNAs). ESRG is a lncRNA highly expressed in hPSCs, and its functional roles are being extensively explored in the field. Here, we identified that the transcription of ESRG can be directly regulated by OCT4, a key self-renewal factor in hPSCs. Knockdown of ESRG induces hPSC differentiation, cell cycle arrest, and apoptosis. ESRG binds to MCM2, a replication-licensing factor, to sustain its steady-state level and nuclear location, safeguarding error-free DNA replication. Further study showed that ESRG knockdown leads to MCM2 abnormalities, resulting in DNA damage and activation of the p53 pathway, ultimately impairs hPSC self-renewal and pluripotency, and induces cell apoptosis. In summary, our study suggests that ESRG, as a novel target of OCT4, plays an essential role in maintaining the cell survival and self-renewal/pluripotency of hPSCs in collaboration with MCM2 to suppress p53 signaling. These findings provide critical insights into the mechanisms underlying the maintenance of self-renewal and pluripotency in hPSCs by lncRNAs.

Aberrant expression of nuclear vimentin and related epithelial-mesenchymal transition markers in nasopharyngeal carcinoma.

Expression of vimentin and the epithelial to mesenchymal transition (EMT) markers E-cadherin, ß-catenin is essential for the progression of various human cancers. Our study aimed to investigate the aberrant localization E-cadherin, ß-catenin and vimentin, and their prognostic significance in 122 nasopharyngeal carcinoma (NPC) patients by immunohistochemistry and immunofluorescence. Our results showed that both membranous and cytoplasmic localization of E-cadherin staining were associated with lymph node metastasis (p = 0.000 and 0.005, respectively) and clinical stage (p = 0.000 and 0.007, respectively). High cytoplasmic ß-catenin correlated significantly with larger tumor size (p = 0.020), lymph node metastasis (p = 0.000) and advanced clinical stage (p = 0.036). However, no significant difference was observed between membranous ß-catenin and clinicopathologic features (p ≥ 0.05). High nuclear vimentin expression correlated significantly with positive lymph node metastasis (p = 0.000) and advanced clinical stage (p = 0.000). Multivariate analysis showed that nuclear vimentin and cytoplasmic E-cadherin were independent prognostic factors (p = 0.016 and 0.001, respectively), as well as M classification (p = 0.001). More importantly, patients with high coexpression of nuclear vimentin and cytoplasmic E-cadherin had shorter survival time (p = 0.000). Furthermore, high coexpression of these two proteins was closely associated with lymph node metastasis (p = 0.000) and advanced clinical stage (p = 0.000). Our studies provide convincing evidence that EMT may play an important role in the biological progression of NPC, and nuclear vimentin and cytoplasmic E-cadherin might have independent prognostic value in NPC patients and serve as novel targets for prognostic therapeutics.

High expression of nuclear Snail, but not cytoplasmic staining, predicts poor survival in nasopharyngeal carcinoma.

BACKGROUND: Transcription factor Snail has been shown to promote tumor progression and metastasis in various cancers. However, its clinical significance in nasopharyngeal carcinoma (NPC) is still scanty. We have explored the clinical significance of Snail expression and its association with patient outcome in NPC. METHODS: Immunohistochemistry was used to examine the expression levels of Snail in 122 patients with NPC. RESULTS: Cytoplasmic Snail was detected in 37.7 %, and nuclear staining was detected in 49.2 % of primary tumors, respectively. No significant associations were found between cytoplasmic Snail and the clinicopathologic variables except lymph node metastasis (P = 0.042). However, nuclear Snail was significantly associated with tumor stage (P = 0.003), T classification (P = 0.045), lymph node metastasis (P = 0.019), distant metastasis (P = 0.003), and reduced E-cadherin expression (P = 0.021). Patients with high nuclear Snail expression, but not cytoplasmic staining, had significantly shorter survival than those with low expression (P < 0.001). Significantly, nuclear Snail was an independent prognostic predictor for NPC (P < 0.001). Furthermore, the prognostic impact was largely limited to stage III-IV patients. CONCLUSIONS: We demonstrated first that nuclear Snail, but not cytoplasmic staining, predicts worse outcome. In addition, the prognostic value in stage III-IV suggests that nuclear Snail could be a potential therapeutic target for late stage of NPC patients.

Tumour budding and the expression of cancer stem cell marker aldehyde dehydrogenase 1 in nasopharyngeal carcinoma.

AIMS: To detect the prognostic significance of tumour budding and its expression of aldehyde dehydrogenase 1 (ALDH1) in nasopharyngeal carcinoma (NPC). METHODS AND RESULTS: Tumour budding was investigated in 105 patients with NPC by immunohistochemistry for pan-cytokeratin (AE1/AE3). The intensity of budding correlated strongly with T classification (P=0.008), lymphatic invasion (P<0.001), vascular invasion (P=0.029), lymph node metastasis (P < 0.001), and clinical stage (P=0.010). Univariate analysis revealed that patients with high budding grade had poorer survival than those with low grade (P=0.002). Multivariate analysis showed that tumour budding was an independent predictor of survival (P=0.001). Furthermore, budding cells showed high-level expression of the cancer stem cell (CSC) marker ALDH1. Budding cells with high-level ALDH1 expression contributed to several aggressive behaviours and poor survival (P=0.000). CONCLUSIONS: We describe, for the first time, the presence of tumour budding and its correlation with aggressive tumour behaviour and poor patient survival in NPC. The degree of tumour budding could be a valuable predictive factor in NPC. In addition, we show, also for the first time, that budding cells in NPC might possess the invasive and metastatic properties of CSCs.

Nuclear expression of N-cadherin correlates with poor prognosis of nasopharyngeal carcinoma.

AIMS: To investigate the aberrant expression of N-cadherin in nasopharyngeal carcinoma (NPC) and its prognostic significance. METHODS AND RESULTS: Immunohistochemical staining for N-cadherin protein was performed on tissue microarray (TMA) from 122 NPC patients. Cytoplasmic N-cadherin was observed in 42.6% and nuclear N-cadherin in 45.1% of NPC tissues. High expression of cytoplasmic and nuclear N-cadherin was associated with a majority of the clinicopathological variables, including lymph node metastasis, distant metastasis and clinical stage. Cytoplasmic N-cadherin was associated positively with nuclear N-cadherin expression (P = 0.000). In univariate analysis, cytoplasmic N-cadherin showed no significant impact on patient prognosis. In contrast, the overall survival was significantly shorter in patients with high nuclear N-cadherin than those with low levels of staining (P = 0.002). A high expression of nuclear N-cadherin predicted poorer survival in patients with late stage disease (P = 0.033), but not those with early tumour stage. In addition, multivariate analysis showed nuclear N-cadherin to bean independent prognostic marker for NPC patients (P = 0.024). CONCLUSIONS: Nuclear N-cadherin expression may represent a valuable prognostic marker in NPC patients, especially those with late stage disease.

Neoplastic spindle cells in nasopharyngeal carcinoma show features of epithelial-mesenchymal transition.

AIM: To investigate whether the neoplastic spindle cells in nasopharyngeal carcinoma (NPC) are associated with the process of epithelial-mesenchymal transition (EMT). METHODS AND RESULTS: We used immunohistochemistry to analyse the expression of cytokeratin, E-cadherin, ß-catenin, vimentin, fibronectin, Snail1, Slug and aldehyde dehydrogenase 1 (ALDH1) in 115 cases of NPC in which there were neoplastic spindle cells; in 47 cases a neoplastic squamous cell component was also present. There was no significant difference in the expression of cytokeratin observed in the neoplastic spindle cells (P = 0.644), compared to the squamous component whereas E-cadherin expression was reduced. By contrast, the expression of ß-catenin, vimentin, fibronectin, Snail1, Slug and ALDH1 was up-regulated in the spindle cells (all P = 0.000). Furthermore, E-cadherin expression was associated negatively with ß-catenin (P < 0.001), vimentin (P < 0.001), fibronectin (P < 0.001), Slug (P < 0.001) and ALDH1 (P < 0.001) in neoplastic spindle cells, but did not correlate with Snail1 expression (P = 0.093). CONCLUSIONS: Our findings demonstrate for the first time that EMT might play an important role in the development of neoplastic spindle cells in NPC.

Overexpressed DNA-binding protein inhibitor 2 as an unfavorable prognosis factor promotes cell proliferation in nasopharyngeal carcinoma.

The aim of the present study was to analyze the expression of DNA-binding protein inhibitor 2 (ID2) in nasopharyngeal carcinoma (NPC) and its correlation with clinicopathological features. It was found that the expression of ID2 was significantly increased in NPC cells when compared with that in NP69 cell line. Similar level of ID2 cytoplasmic expression was observed in NPC when compared with that in non-cancerous nasopharynx tissues. However, the level of ID2 in nucleus was increased in NPC when compared with that in normal nasopharynx tissues. Furthermore, the higher expression level of nuclear ID2 was significantly associated with tumor size (T classification), lymph node metastasis (N classification), and clinical stage. Patients with increased ID2 expression level had poorer overall survival rates than those with low ID2 levels. The inhibition of ID2 expression in NPC cell line SUNE1 by lentiviral-mediated short hairpin RNA could suppress cell proliferation and colony formation, but did not disrupt cell migration. Knocking down the expression of ID2 by RNA interference could down-regulate the expression of Snail, suggesting that ID2-promoted cell growth, partially attributing to the regulation of Snail activity in NPC. Our study demonstrated that over-expression of ID2 protein is an unfavorable prognostic factor which promotes cell proliferation in NPC.

TBL1X and Flot2 form a positive feedback loop to promote metastasis in nasopharyngeal carcinoma.

Metastasis is the main cause of death in patients with nasopharyngeal carcinoma (NPC). The molecular mechanisms underlying the metastasis of NPC remain to be elucidated. TBL1X has been shown abnormally expressed in diverse cancers. However, the role and mechanism of TBL1X in NPC remain unknown. Here, we showed TBL1X expression was significantly higher in metastatic NPC tissues compared to non-metastatic tissues and significantly correlated with TNM stage and metastasis of NPC patients. In addition, NPC patients with high TBL1X expression had a poor prognosis. TBL1X interacted with TCF4 to trans-activate Flot2 expression. TBL1X promoted NPC cell migration and invasion in vitro and in vivo through Flot2. Moreover, Flot2 increased the expression of TBL1X by upregulating c-myc, which was identified to be a positively regulatory transcription factor of TBL1X. TBL1X could restore the functional changes of NPC cells resulting from Flot2 alteration. TBL1X and Flot2 were positively correlated in NPC. Patients with high expression of both TBL1X and Flot2 possessed poorer overall survival (OS) and disease-free survival (DFS) compared to patients with high expression of any single one of the two proteins. Our findings demonstrate that TBL1X and Flot2 positively regulate each other to promote NPC metastasis, which provides novel potential molecular targets for NPC treatment.

Differential effects of macrophage subtypes on SARS-CoV-2 infection in a human pluripotent stem cell-derived model.

Dysfunctional immune responses contribute critically to the progression of Coronavirus Disease-2019 (COVID-19), with macrophages as one of the main cell types involved. It is urgent to understand the interactions among permissive cells, macrophages, and the SARS-CoV-2 virus, thereby offering important insights into effective therapeutic strategies. Here, we establish a lung and macrophage co-culture system derived from human pluripotent stem cells (hPSCs), modeling the host-pathogen interaction in SARS-CoV-2 infection. We find that both classically polarized macrophages (M1) and alternatively polarized macrophages (M2) have inhibitory effects on SARS-CoV-2 infection. However, M1 and non-activated (M0) macrophages, but not M2 macrophages, significantly up-regulate inflammatory factors upon viral infection. Moreover, M1 macrophages suppress the growth and enhance apoptosis of lung cells. Inhibition of viral entry using an ACE2 blocking antibody substantially enhances the activity of M2 macrophages. Our studies indicate differential immune response patterns in distinct macrophage phenotypes, which could lead to a range of COVID-19 disease severity.

Elevated expression of CDK4 in lung cancer.

BACKGROUND: The aim of the present study was to analyze the expression of Cyclin-dependent kinase 4 (CDK4) in lung cancer and its correlation with clinicopathologic features. Furthermore, the involvement of CDK4-mediated cell cycle progression and its molecular basis were investigated in the pathogenesis of lung cancer. METHODS: Using immunohistochemistry analysis, we analyzed CDK4 protein expression in 89 clinicopathologically characterized lung cancer patients (59 males and 30 females) with ages ranging from 36 to 78 years and compared them to 23 normal lung tissues. Cases with cytoplasmic and nuclear CDK4 immunostaining score values greater than or equal to 7 were regarded as high expression while scores less than 7 were considered low expression. The correlation between the expression level of CDK4 and clinical features was analyzed. Furthermore, we used lentiviral-mediated shRNA to suppress the expression of CDK4 and investigate its function and molecular mechanism for mediating cell cycle progression. RESULTS: The expression level of CDK4 protein was significantly increased in lung cancer tissues compared to normal tissues (P < 0.001). In addition, high levels of CDK4 protein were positively correlated with the status of pathology classification (P = 0.047), lymph node metastasis (P = 0.007), and clinical stage (P = 0.004) of lung cancer patients. Patients with higher CDK4 expression had a markedly shorter overall survival time than patients with low CDK4 expression. Multivariate analysis suggested the level of CDK4 expression was an independent prognostic indicator (P < 0.001) for the survival of patients with lung cancer. Use of lentiviral-mediated shRNA to inhibit the expression of CDK4 in lung cancer cell line A549 not only inhibited cell cycle progression, but also dramatically suppressed cell proliferation, colony formation, and migration. Furthermore, suppressing CDK4 expression also significantly elevated the expression of cell cycle regulator p21 CONCLUSION: Overexpressed CDK4 is a potential unfavorable prognostic factor and mediates cell cycle progression by regulating the expression of p21 in lung cancer.

Increased expression of MMP9 is correlated with poor prognosis of nasopharyngeal carcinoma.

INTRODUCTION: The aim of the present study was to analyze the expression of matrix metalloproteinase 9 (MMP9) in nasopharyngeal carcinoma (NPC) and its correlation with clinicopathologic features, including the survival of patients with NPC. METHODS: Using real-time PCR, we detected the mRNA expression of MMP9 in normal nasopharyngeal tissues and nasopharyngeal carcinoma (NPC) tissues. Using immunohistochemistry analysis, we analyzed MMP9 protein expression in clinicopathologically characterized 164 NPC cases (116 male and 48 female) with age ranging from 17 to 80 years (median = 48.4 years) and 32 normal nasopharyngeal tissues. Cases with greater than or equal to 6 and less than 6 of the score value of cytoplasmic MMP9 immunostaining were regarded as high expression and low expression, respectively. The relationship between the expression levels of MMP9 and clinical features was analyzed. RESULTS: The expression level of MMP9 mRNA was markedly greater in NPC tissues than that in the nasopharyngeal tissues. Immunohistochemical analysis revealed that the protein expression of MMP9 detected in NPC tissues was higher than that in the nasopharyngeal tissues (P = 0.004). In addition, high levels of MMP9 protein were positively correlated with the status of lymph node metastasis (N classification) (P = 0.002) and clinical stage (P < 0.001) of NPC patients. Patients with higher MMP9 expression had a significantly shorter overall survival time than did patients with low MMP9 expression. Multivariate analysis suggested that the level of MMP9 expression was an independent prognostic indicator (P = 0.008) for the survival of patients with NPC. CONCLUSION: High level of MMP9 expression is a potential unfavorable prognostic factor for patients with NPC.