B7-H3 is associated with the armored-cold phenotype and predicts poor immune checkpoint blockade response in melanoma.

Melanoma is the most suitable tumor type for immunotherapy, but not all melanoma patients could respond to immunotherapy. B7 homolog 3 (B7-H3) belongs to the B7 family and is overexpressed in a number of malignant tumors, but the expression pattern of B7-H3 in melanoma has not been well summarized. The expression of B7-H3 was investigated in melanoma and its correlations with features of the tumor microenvironment (TME) by using various public databases, including the Cancer Genome Atlas (TCGA), the GEPIA, and the Human Protein Atlas databases. In addition, the in-house melanoma tissue microarray was applied to validate the results from public databases. Based on the public and in-house cohorts, we found that B7-H3 was overexpressed in melanoma tumor tissues and high B7-H3 expression was related to poor clinical outcome. Moreover, B7-H3 was negatively correlated with levels of tumor-infiltrating lymphocytes (TILs) and positively correlated with collagen infiltration. With clinical translational value, the predictive value of B7-H3 for conventional immunotherapy was detected using the Kaplan-Meier plotter tool, and the results showed that melanoma patients with high B7-H3 expression were insensitive to anti-PD-1 and anti-CTLA-4 immunotherapy. In conclusion, we first investigate the expression of B7-H3 in melanoma and its correlations with the TME features, and indicate B7-H3 as a promising therapeutic target in melanoma patients that are insensitive to conventional immunotherapy.

Prognostic nomogram in patients with gastrointestinal stromal tumors: a SEER-based study.

Background: Gastrointestinal stromal tumor (GIST) is a common mesenchymal tumor of the gastrointestinal system. They originate from the interstitial cells of Cajal located within the muscle layer and are characterized by over-expression of the tyrosine kinase receptor KIT. Methods: Data from the Surveillance Epidemiology, and End Results (SEER) database of 1,213 patients diagnosed with GIST between 2010 and 2019 were dichotomized into a modeling set and a validation set at a 2:1 ratio. For the modeling set, both univariate and multivariate Cox regression analyses were used to identify independent prognostic factors. A nomogram was then constructed based on these determinants. Model efficacy was tested using receiver operating characteristic (ROC) curves, calibration curves, clinical decision curves, and risk stratification analysis in both subsets. Results: Identified prognostic determinants included age, sex, pathological differentiation level, tumor-node-metastasis (TNM) stage, surgical intervention, radiotherapy, and marital status. The constructed nomogram showed area under the ROC curve (AUC) values of 0.822, 0.793, and 0.779 for 1-, 3-, and 5-year overall survival (OS) in the modeling set, respectively, while in the validation set, the values were 0.796, 0.823, and 0.806, respectively. Calibration plots from both sets confirmed the concordance between predicted and observed survival. Decision curve analysis (DCA) indicated significant clinical utility for the nomogram. Risk stratification of the patient data revealed distinct survival differences between high-risk and low-risk cohorts in both sets (P<0.001). Conclusions: A novel and potent nomogram for the prognosis of GIST has been introduced. This model's precision offers crucial insights for clinical decisions, yet further external validation remains essential.

PSMC2 is a Novel Prognostic Biomarker and Predicts Immunotherapeutic Responses: From Pancreatic Cancer to Pan-Cancer.

Background: Proteasome 26S subunit ATPase 2 (PSMC2) is a part of the 19S regulatory complex, which catalyzes the unfolding and transport of substrates into the 20S proteasome. Our previous research demonstrated that PSMC2 participates in the tumorigenesis and progression of pancreatic cancer (PC). However, no systematic analysis has been conducted to conclude its expression pattern and correlation with tumor immunity. Aim: To investigate the expression level of PSMC2 in PC, its prognostic value and its relationship with tumor immunity. Methods: In numerous public and internal cohorts, the expression, prognostic significance, and immunological connections of PSMC2 in PC were investigated. Additionally, using data from The Cancer Genome Atlas (TCGA), a pan-cancer analysis was carried out to examine PSMC2's immunological assocaition, and the predictive power of PSMC2 for immunotherapy was also evaluated in numerous public cohorts. Results: PSMC2 was overexpressed in tumor tissues and linked to unfavorable prognosis in PC. PSMC2 was not only positively correlated with TIICs, also positively correlated with immune checkpoints in PC. In addition to PC, PSMC2 was expected to be an indicator of high immunogenicity in most cancer types. Importantly, PSMC2 could predict the immunotherapeutic responses in various cancer types, including urothelial carcinoma and breast cancer. Conclusion: From PC to pan-cancer analysis, we report that PSMC2 is a novel prognostic biomarker in multiple cancer types. PSMC2 is related to the immuno-hot phenotype and predicts the outcome of immunotherapy. Therefore, the current study emphasizes that cancer patients with high PMSC2 expression should actively receive immunotherapy to improve their prognosis.

Total T Cell Density and Expression of T Memory Stem Cell Markers are Associated with Better Prognosis in Colon Cancer.

Background: Immune checkpoint inhibitors have achieved limited clinical effectiveness in colon cancer. Stem memory T cells (TSCMs) and in-situ cytotoxic T cells are dominant contributors to host immunity. Currently, data on the correlation between TSCM and T cell abundance and clinicopathological characteristics in colon cancer are largely unavailable. Methods: In-situ cytotoxic T cells are identified based on the quantification of CD3+ and CD8+ markers using immunohistochemistry (IHC) in the core of the tumor and the invasive margin of the tumor. The expression of representative markers of TSCMs, CD27 and CD95, was assayed using IHC in colon cancer tissues. Correlations between the levels of each marker and the clinicopathological characteristics as well as prognosis were evaluated. Results: High densities of CD3+ and CD8+ T cells correlated with stage I-II tumors, whereas a lower infiltration of cytotoxic T cells correlated with advanced-stage tumors. CD27 and CD95 were both expressed in the membrane of T cells present in the tumor stroma and their levels showed a negative correlation with the TNM stage. CD3, CD8, and CD27 were expressed at the same locations simultaneously, indicating their coordinated action against cancer. In addition, cytotoxic T cell densities and CD27 and CD95 expression remained independent prognostic factors for overall survival. Conclusion: In-situ cytotoxic T cells and TSCMs play important roles in colon cancer development. TSCMs marker CD27 and CD95 were both indicators of survival in patients with colon cancer. Thus, it is believed that TSCMs represent a desirable population for future use in combination immunotherapy.

STAT3/miR-130b-3p/MBNL1 feedback loop regulated by mTORC1 signaling promotes angiogenesis and tumor growth.

BACKGROUND: Aberrantly activated mammalian target of rapamycin complex 1 (mTORC1) plays a vital role in tumor angiogenesis, but its precise mechanisms are still unclear. METHODS: Micro-RNA-130b-3p (miR-130b-3p) expression in mTORC1-activated and control cells was examined by quantitative real-time PCR (qRT-PCR). MiR-130b-3p levels and their correlation with mTORC1 activity were evaluated by analyzing publicly available databases and in-house head and neck squamous cell carcinoma (HNSCC) tissues. The role of miR-130b-3p in mTORC1-mediated angiogenesis and tumor growth was examined using tube formation assay, chicken chorioallantoic membrane assay, cell line - derived xenograft models, and an HNSCC patient-derived xenograft (PDX) model. The regulatory mechanisms among signal transducer and activator of transcription 3 (STAT3), miR-130b-3p, and muscleblind-like protein 1 (MBNL1) were investigated via bioinformatics analyses, qRT-PCR, western blot, RNA immunoprecipitation, immunofluorescence, luciferase reporter assay, and chromatin immunoprecipitation assay. RESULTS: Elevated miR-130b-3p enhanced the angiogenic and tumorigenic abilities of mTORC1-activated cells both in vitro and in vivo. STAT3, a downstream effector of mTORC1, transactivated miR-130b-3p by direct binding promoter of the miR-130b gene. MBNL1 was identified as a direct target of miR-130b-3p. MBNL1 depletion rescued the compromised angiogenesis and tumor growth caused by miR-130b-3p inhibition. MiR-130b-3p levels were significantly upregulated and positively correlated with mTORC1 signaling in multiple cancers. MiR-130b-3p inhibition attenuated tumor angiogenesis and growth in an HNSCC PDX model. MBNL1 feedback inhibited STAT3 activation in mTORC1-activated cells. CONCLUSIONS: The STAT3/miR-130b-3p/MBNL1 feedback loop plays a vital role in mTORC1-mediated angiogenesis and tumor progression. This pathway could be targeted for therapeutic intervention of mTORC1-related cancers.

NEUROD1 predicts better prognosis in pancreatic cancer revealed by a TILs-based prognostic signature.

It has been well-defined that tumor-infiltrating lymphocytes (TILs) play critical roles in pancreatic cancer (PaCa) progression. This research aimed to comprehensively explore the composition of TILs in PaCa and their potential clinical significance. A total of 178 samples from the TCGA and 63 samples from the GSE57495 dataset were enrolled in our study. ImmuCellAI was applied to calculate the infiltrating abundance of 24 immune cell types in PaCa and further survival analysis revealed the prognostic values of TILs in PaCa. Moreover, the Hallmark enticement analysis of differentially expressed genes (DEGs) between low- and high-risk groups was performed as well. Immunohistochemistry staining was used to evaluate NEUROD1 expression. As result, different kinds of TILs had distinct infiltrating features. In addition, Specific TILs subsets had notable prognostic values in PaCa. We further established a 6-TILs signature to assess the prognosis of PaCa patients. Kaplan-Meier and Cox regression analyses both suggested the significant prognostic value of the signature in PaCa. Based on the prognostic signature, we screened a great deal of potential prognostic biomarkers and successfully validated NEUROD1 as a novel prognostic biomarker in PaCa. Overall, the current study illuminated the immune cells infiltrating the landscape in PaCa and identified a TILs-dependent signature and NEUROD1 for prognostic prediction in PaCa patients.

Microfluidic devices: The application in TME modeling and the potential in immunotherapy optimization.

With continued advances in cancer research, the crucial role of the tumor microenvironment (TME) in regulating tumor progression and influencing immunotherapy outcomes has been realized over the years. A series of studies devoted to enhancing the response to immunotherapies through exploring efficient predictive biomarkers and new combination approaches. The microfluidic technology not only promoted the development of multi-omics analyses but also enabled the recapitulation of TME in vitro microfluidic system, which made these devices attractive across studies for optimization of immunotherapy. Here, we reviewed the application of microfluidic systems in modeling TME and the potential of these devices in predicting and monitoring immunotherapy effects.

The Clinical Significance of Deglycosylated PD-L1 Level Detection Using 28-8 Monoclonal Antibody in Lung Adenocarcinoma.

Purpose: The aim of this study was to explore the clinical significance of deglycosylated PD-L1 level and its correlation with EGFR and ALK mutation in lung adenocarcinoma. Materials and Methods: We estimated the intensity of both native and deglycosylated PD-L1 signals using a 28-8 antibody on lung adenocarcinoma tissue microarray sections. We analyzed the difference in the H-score between tumor and paratumor tissues, as well as that before and after deglycosylation. Correlations between EGFR or ALK status and PD-L1 expression were analyzed. We also evaluated the differences among survival curves. Results: The expression level of PD-L1 in lung adenocarcinoma tissues was significantly higher than that in paratumor tissues (P<0.0001). Deglycosylation significantly enhanced the detection of PD-L1 in tumor tissues (P<0.0001). There was no statistical significance between the signal intensity of deglycosylated PD-L1 and the survival of patients (P=0.9099). However, the response to deglycosylation of PD-L1 was significantly correlated with the survival of patients with stage N1-N3 (P=0.0435) and stage T3-T4 (P=0.0366) and male patients (P=0.0258). A statistical trend was found in the correlation between the response to deglycosylation of PD-L1 and the survival of patients with grade II-III plus grade III (P=0.0973). Correlation between EGFR or ALK status and the expression of PD-L1 was not found (P>0.05). Conclusion: PD-L1 deglycosylation enhances the detection of PD-L1 when utilizing a 28-8 antibody. Moreover, the response to deglycosylation of PD-L1 may predict the survival of certain patients with lung adenocarcinoma.

Case Report: Successful Immunotherapy Improved the Prognosis of the Unfavorable Subset of Cancer of Unknown Primary.

Background: Cancer of unknown primary (CUP) is heterogeneous and has a wide variety of clinical presentations and a poor prognosis in most patients, with a median overall survival of only 6 months. The development of molecular profiling contributes to precision therapy, and targeted drugs and immune checkpoint inhibitors (ICIs) greatly promote individualized treatment. Case presentation: Here, we reported a case of an unfavorable subset of CUP who had a long time of survival after the immunotherapy-prominent comprehensive treatment. A 48-year-old man presented with back pain and a cough. A diagnostic work-up showed bone marrow, multiple bones, and lymph node metastasis. Lymph node pathology implies metastatic poorly differentiated cancer. Next-generation sequencing (NGS) showed no special targets, but the tumor proportion score (TPS) of programmed death-ligand 1 (PD-L1) was 80% and the tumor mutation burden (TMB) was 16.7 per million bases. After two cycles of pembrolizumab 200 mg D1 plus nanoparticle albumin-bound (nab)-paclitaxel 200 mg D1&8 (q3w), PET-CT and bone marrow aspiration cytology showed a complete response (CR). Subsequently, pembrolizumab alone was used for three months. The left inguinal lymph nodes showed new metastasis. After two cycles of the combination treatment of pembrolizumab and (nab)-paclitaxel, a partial response (PR) was achieved. After seven months, retroperitoneal lymph nodes showed new metastasis, and the sequential treatment with radiotherapy and pembrolizumab exhibited encouraging efficacy. To date, the patient has survived nearly 40 months with the combination therapy. Conclusions: The ICI-prominent comprehensive treatment provided clinical benefit for the reported case of CUP. Thus, CUP patients with markers of benefiting from immunotherapy should be actively treated with immunotherapy to improve their prognosis.

miR-224 Regulates the Aggressiveness of Hepatoma Cells Through the IL-6/STAT3/SMAD4 Pathway.

BACKGROUND: Previous studies have shown that miR-224 regulates the progression of liver cancer. The aim of this study was to investigate the underlying mechanisms. METHODS: The miR-224, p-STAT3 and SMAD4 expression levels were checked with tissue or/and serum samples of HCC patients by qRT-PCR or IHC methods. The regulatory role of IL-6 in p-STAT3 and SMAD4 was investigated by Western-blot. The targeted gene of miR-224 was verified by both Western-blot and luciferase reporter assay. Furthermore, the carcinogenesis of miR-224 in HCC was investigated by cell experiments in vitro and mouse xenograft model and in vivo imaging in vivo. RESULTS: It was found miR-224 was elevated in both tissue and serum of HCC patients. The p-STAT3 expression was higher but the SMAD4 was lower in the HCC tumor tissues. Moreover, IL-6 can induce the p-STAT3/STAT3 and miR-224 expression in HCC cells and STAT3 played the bridge role between IL-6 and miR-224. Target gene studies found miR-224 targeted the 3'UTR of SMAD4. Finally, the promoting roles of miR-224in the growth, proliferation, invasion and migration of HCC were discovered by in vitro and in vivo studies. CONCLUSION: It implies that miR-224 may potentially represent a new target for developing novel anti-HCC therapeutics.

Removal of N-Linked Glycosylation Enhances PD-L1 Detection in Colon Cancer: Validation Research Based on Immunohistochemistry Analysis.

In recent years, immunotherapies have emerged as effective therapeutic strategies for treating human cancers. However, accumulating evidence has revealed an inconsistency between the response to immune checkpoint inhibitors and programmed death ligand 1 (PD-L1) expression status detected by immunohistochemistry staining. Recent research has revealed that the removal of N-Linked glycosylation significantly enhanced PD-L1 detection, resulting in both more accurate PD-L1 quantification and clinical outcome prediction. In the present study, we evaluated natural and deglycosylated PD-L1 expression in colon cancer using the PD-L1 28-8 antibody. The results of the present study validated the hypothesis that PD-L1 had a higher expression in colon cancer tissues compared with normal tissues. Additionally, colon tumors with defective mismatch repair tended to express higher PD-L1 than those without. Most importantly, the results of the present study indicated that the removal of N-linked glycosylation remarkably enhanced PD-L1 detection. Moreover, the PD-L1 signal intensity of samples with a low natural PD-L1 signal was enhanced more remarkably than that of samples with high signal intensity. Overall, our research provides an improved strategy for patient stratification for anti-PD-1/PD-L1 therapy, which deepens the clinical significance of this established strategy for treatment of colon cancer.

MicroRNA-23b-3p promotes pancreatic cancer cell tumorigenesis and metastasis via the JAK/PI3K and Akt/NF-κB signaling pathways.

MicroRNA (miR)-23b-3p plays an important role in tumor growth, proliferation, invasion and migration in pancreatic cancer (PC). However, the function and mechanistic role of miR-23b-3p in the development of PC remains largely unknown. In the present study, the miR-23b-3p levels in the serum of patients with PC were found to be elevated, and the phosphorylation levels of Janus kinase (JAK)2, PI3K, Akt and NF-κВ were found to be upregulated. In addition, miR-23b-3p was induced in response to interleukin-6 (IL-6), which is known to be involved in the progression of PC. Overexpression of miR-23b-3p, on the other hand, activated the JAK/PI3K and Akt/NF-κB signaling pathways in PC cells, as evidenced by miR-23b-3p-induced upregulation of phosphorylated (p-)JAK2, p-PI3K, p-Akt and p-NF-κВ, as well as the downregulation of PTEN; and these effects were found to be reversible by miR-23b-3p inhibition. Furthermore, miR-23b-3p was found to downregulate PTEN by directly targeting the 3'-untranslated region of PTEN mRNA. Notably, in an in vivo xenograft mouse model, overexpression of miR-23b-3p accelerated PC cell-derived tumor growth, activated the JAK/Akt/NF-κВ signaling pathway and promoted liver metastasis. In contrast, knockdown of miR-23b-3p suppressed tumor growth and metastasis as well as JAK/Akt/NF-κВ signaling activity. In vivo imaging of the mice further confirmed the metastasis promoting role of miR-23b-3p in PC. These results suggested that miR-23b-3p enhances PC cell tumorigenesis and metastasis, at least, partially via the JAK/PI3K and Akt/NF-κB signaling pathways. Therefore, targeting miR-23b-3p or the JAK/PI3K and Akt/NF-κB signalings may be potential therapeutic strategy against PC.

RNA-Seq Profiling of Circular RNAs and the Oncogenic Role of circPVT1 in Cutaneous Squamous Cell Carcinoma.

BACKGROUND: Cutaneous squamous cell carcinoma (CSCC) is associated with a poor 5-year survival rate. circRNAs have an important role in a number of physiological and pathological processes. However, the relationship between circRNAs and cutaneous squamous cell carcinoma (CSCC) is unclear. PURPOSE: The aim of the present study was to investigate the expression of circRNAs in cutaneous squamous cell carcinoma (CSCC) and its effect on CSCC proliferation and metastasis. METHODS: We used high-throughput sequencing (RNA-seq) to identify circRNAs that were differentially in CSCC tissue and their paracarcinoma tissue. Quantitative real-time PCR results confirm deep-sequencing findings in CSCC tissue and cell lines. CCK-8 assay and flow cytometry were used to detect the effect of circPVT1 on the proliferation and migration of CSCC cells. RESULTS: We identified 449 circRNAs that were differentially expressed between CSCC and normal adjacent tissue samples. circPVT1 (hsa_circ_0001821) was further researched to confirm its oncogene role in CSCC. CONCLUSION: Differentially expressed circular RNA plays an important role in the development of CSCC, and circPVT1 may be an important target for the treatment of CSCC.

Efficacy of fiberoptic bronchoscopy and bronchoalveolar lavage in childhood-onset, complicated plastic bronchitis.

BACKGROUND: Plastic bronchitis (PB) is a rare, variable, and potentially fatal disease. This study aimed to assess the efficacy of fiberoptic bronchoscopy (FOB) and bronchoalveolar lavage (BAL) in treating children with PB. METHODS: In total, 15 children with PB, between 2012 and 2020, were enrolled in our study. Within 12 hours of admission, FOB and BAL were performed and reviewed under local anesthesia and sedation. Before and after FOB, clinical findings and chest imaging were evaluated. RESULTS: Regarding the onset of symptoms before FOB, all cases had prominent cough for 7.00 ± 4.55 days, and 14 had persistent high fever. In total, 13 cases had complete obstruction from bronchial casts, consistent with consolidated lesions; 2 had partial airway obstruction. Within 3 days, complete resolution was revealed in nine cases. Overall, six cases underwent repeated FOB (range, 2-3 times) for persistent atelectasis and airway obstruction. Except for two cases with type 2 PB, cast histology confirmed type 1 PB for all cases. Only eight children had minor intra- and post-procedure complications. Reverse transcription-polymerase chain reaction for Mycoplasma pneumoniae in sputum and BAL samples were positive in 13 cases. Next-generation sequencing of the BAL samples was positive for adenovirus and Human parainfluenza virus in one case, respectively. During 1 month to 7 years of follow-up, no patient developed PB recurrence, asthmatic attacks, or chronic cough. CONCLUSIONS: Early FOB and BAL were effective in alleviating clinical findings, atelectasis, and airway obstruction. Serial FOB could be performed in patients with recurrent symptoms.

Myosin light chain kinase inhibitor ML7 improves vascular endothelial dysfunction and permeability via the mitogen-activated protein kinase pathway in a rabbit model of atherosclerosis.

Endothelial dysfunction (ED) and hyperpermeability are considered as the initiating steps in early atherosclerosis. Phosphorylation of myosin light chain (MLC) is key to cause vascular hyperpermeability via endothelial cell contraction. However, it is unclear whether MLC phosphorylation can also regulate the balance between contraction and relaxation of endothelial cells, thereby affecting endothelium-dependent diastolic function and leading to ED. The present study investigated relationships between ED and MLC phosphorylation and underlying mechanisms. Twenty-four male New Zealand white rabbits were randomly divided into three groups: control, AS, and ML7 (MLCK inhibitor) groups, and fed with normal diet, high-fat diet (HFD), and HFD plus oral ML7 (1 mg/kg daily) respectively. HFD-fed rabbits showed typical atheromatous lesions and endothelial hyperpermeability, and these lesions could be partly reversed following ML7 therapy. Western blotting revealed significant increased expression of myosin light chain kinase (MLCK) and phosphorylation of MLC, JNK, and ERK in the arterial wall of rabbits in the AS group compared with those of the control group (p < 0.05), whereas the ML7 group showed markedly decreased levels of these proteins compared with the AS group (p < 0.05). The endothelium-dependent relaxation rate was significantly reduced both in vitro and in vivo in AS group, and was improved using ML7 therapy. Taken together, these results indicate that MLCK expression and subsequent MLC phosphorylation increase vascular endothelial permeability and endothelium-dependent diastolic dysfunction by promoting endothelial cell contraction, which may be initiated by the activation of the MAP/ERK (MEK) and MAP/JNK (MEK) pathways.

The emerging roles of circular RNAs in ovarian cancer.

Circular RNA (circRNA) is a novel class of regulatory noncoding RNA (ncRNA) molecules with a unique covalently closed loop structure. Next-generation sequencing shows that thousands of circRNAs are widely and stably expressed in multiple eukaryotes. As novel regulatory ncRNAs, circRNAs possess several specific molecular functions, including regulating gene transcription and translation, acting as miRNA sponges, and interacting with functional proteins. Ovarian cancer (OvCa) is one of the most aggressive malignant diseases affecting the lives of thousands of women worldwide, and the majority of OvCa cases are diagnosed at advanced stages. Accumulating evidence has revealed the significant roles of circRNAs in the occurrence and progression of OvCa, indicating the function of circRNAs as promising biomarkers and their therapeutic relevance in this disease. This review aims to summarize the mechanisms by which circRNAs mediate OvCa progression as well as their diagnostic and prognostic values in OvCa.

Novel insights into Dhh signaling in antler chondrocyte proliferation and differentiation: Involvement of Foxa.

The desert hedgehog (Dhh) is crucial for spermatogenesis and Leydig cell differentiation, but little is known regarding its physiological function in cartilage. In this study, Dhh mRNA was abundant in antler chondrocytes, where it advanced cell proliferation concomitant with accelerated transition from the G1 to the S phase and induced elevation of the hypertrophic chondrocyte markers, Col X and Runx2. Silencing of Ptch1 resulted in appreciable Smo accumulation and enhanced rDhh stimulation of Smo, whose impediment by cyclopamine obscured the proliferative function of Dhh and alleviated its guidance of chondrocyte differentiation. Further analysis evidenced the noteworthy positive action of Smo in the bridging between Dhh and Gli transcription factors. Obstruction of Gli1 by GANT58 caused the failed stimulation of Col X and Runx2 by rDhh. Analogously, siRNA against Gli1-3 hindered chondrocyte differentiation in the context of rDhh. Simultaneously, Gli transcription factors mediated the regulation of Dhh on Foxa1, Foxa2, and Foxa3, whose knockdown impaired chondrocyte differentiation. Attenuation of Foxa antagonized the augmentation of Col X and Runx2 generated by rDhh. Collectively, Dhh signaling through its target Foxa appears to induce antler chondrocyte proliferation and differentiation.

PSMC2 is Up-regulated in Pancreatic Cancer and Promotes Cancer Cell Proliferation and Inhibits Apoptosis.

Objective: The lack of effective therapeutic targets poses a leading challenge to prolong survival and improve the quality of life for pancreatic cancer patients. Proteasome 26S subunit ATPase 2 (PSMC2), a recently discovered gene, has been implicated in certain human carcinogenesis. However, limited data is available concerning the functional significance of PSMC2 in cancer cell growth, and whether it plays a role in pancreatic carcinogenesis remains unknown. Materials and Methods: Quantitative RT-PCR (qRT-PCR) was performed to assess mRNA expression levels of PSMC2 in different pancreatic cancer cell lines. Knockdown of PSMC2 was achieved by using short hairpin RNA (shRNA). The effects of PSMC2 silencing on pancreatic cancer cell proliferation and apoptosis were evaluated by the MTT cell proliferation assay, Celigoassays, Annexin V fluorescence-activated cell sorting (FACS) assay and Caspase-3/7 array. Results: High expression of PSMC2 was detected in three pancreatic cancer cell lines (SW1990, PANC-1, and AsPC-1). Knockdown of PSMC2 in SW1990 cells inhibited proliferation and enhanced apoptosis. Conclusions: Our primary study suggests that PSMC2 might be involved in the progression of pancreatic cancer and may serve as a potential therapeutic target.

The Importance of a Conjoint Analysis of Tumor-Associated Macrophages and Immune Checkpoints in Pancreatic Cancer.

OBJECTIVES: Tumor-associated macrophages are dominant players in establishing the inmmunosuppressive microenvironment in pancreatic ductal adenocarcinoma (PDAC). Immune checkpoint inhibitor monotherapy has achieved limited clinical effectiveness. To date, the interaction of macrophages and checkpoint regulators and their correlation with clinicopathologic characteristics in PDAC have been largely unavailable. METHODS: Macrophages and immune checkpoint expression were assessed by immunohistochemistry from 80 PDAC samples. Clinicopathologic features and the prognostic value of each marker were evaluated. In vitro changes in the expression of immune markers in cocultured macrophages and PDAC cells were detected by Western blot and immunosorbance assays. RESULTS: The macrophages marker CD163 and the checkpoint marker programmed death-ligand 1 (PD-L1) remained as the independent prognostic factors for overall survival (hazard ratio, 2.543; P = 0.017 and hazard ratio, 2.389; P = 0.021). Furthermore, integrated analysis of CD163 and PD-L1 served as more optimal indicators of survival (P = 0.000). In vitro coculture of macrophages and PDAC cells significantly increased the expression of CD163 and PD-L1, compared with monocultured counterpart (P < 0.05). CONCLUSIONS: Combined analysis of CD163 and PD-L1 was enhanced indicators of survival in PDAC patients. The interaction of macrophages and immune checkpoints implied the value of the combination therapy.

Function and regulation of transforming growth factor ß1 signalling in antler chondrocyte proliferation and differentiation.

OBJECTIVES: Chondrocyte proliferation and differentiation are crucial for endochondral ossification, but their regulatory mechanism remains unclear. The present study aimed to determine the physiological function of TGFß1 signalling in the proliferation and differentiation of antler chondrocytes and explore its relationship with Notch, Shh signalling and Foxa. MATERIALS AND METHODS: Immunofluorescence, Western blot, MTS assay, flow cytometry, RNA interference and real-time PCR were used to analyse the function and regulatory mechanisms of TGFß1 signalling in antler chondrocyte proliferation and differentiation. RESULTS: TGFß1, TGFBR1 and TGFBR2 were highly expressed in antler cartilage. TGFß1 promoted chondrocyte proliferation, increased the proportion of S-phase cells and induced the expression of hypertrophic chondrocyte markers Col X, Runx2 and Alpl. However, this induction was weakened by TGFß receptor inhibitor SB431542 and Smad3 inhibitor SIS3. Simultaneously, TGFß1 activated Notch and Shh signalling whose blockage attenuated the above effects of rTGFß1, whereas addition of rShh rescued the defects in chondrocyte proliferation and differentiation elicited by SB431542 and SIS3. Further analysis revealed that inhibition of Notch signalling impeded TGFß1 activation of the Shh pathway. Knockdown of Foxa1, Foxa2 and Foxa3 abrogated the effects of TGFß1 on chondrocyte differentiation. Notch and Shh signalling mediated the regulation of Foxa transcription factors by TGFß1. CONCLUSIONS: TGFß1 signalling could induce the proliferation and differentiation of antler chondrocytes through Notch-Shh-Foxa pathway.