Objective: To construct a Family with sequence similarity 50 member A(FAM50A) gene knockout mouse insulinoma pancreatic β-cell line Beta-TC-6 using CRISPR/Cas9 gene editing technology and to prepare polyclonal antibodies specifically recognizing FAM50A. Methods: Two guide RNAs(sgRNAs) targeting the FAM50A gene were designed,and a recombinant plasmid expressing blue fluorescent protein(BFP) was constructed for gene knockout.The successfully constructed plasmid was transfected into Beta-TC-6 cells,and BFP-positive single cells were isolated for clonal expansion.The expanded monoclonal cell lines were genotyped by Sanger sequencing,and FAM50A protein expression was assessed by Western blot.Purified human recombinant FAM50A protein was used to immunize New Zealand rabbits for the preparation of a polyclonal antibody.The specificity of the prepared antibody was then validated using the successfully established FAM50A knockout cell line. Results: A monoclonal cell line with a successful knockout of the FAM50A gene was identified.Sanger sequencing confirmed base deletions at the target site.Western blot analysis showed a complete absence of FAM50A protein expression in this cell line.The prepared polyclonal antibody successfully recognized endogenous murine FAM50A protein in wild-type Beta-TC-6 cells and in hTERT-RPE1 cells overexpressing human FAM50A-GFP fusion protein,while no signal was detected in the FAM50A knockout cells. Conclusion This study successfully established a FAM50A gene knockout Beta-TC-6 cell model and generated a FAM50A polyclonal antibody,providing powerful tools for future research.

Monoclonal antibody drugs that inhibit programmed death 1 (PD-1) or programmed death ligand 1 (PD-L1) have been widely used in esophageal cancer (EC) and yielded significant therapeutic responses. However, only a few patients obtain lasting clinical benefits due to primary or acquired drug resistance, and new treatment schemes are urgently needed. The tumor immune microenvironment is the main factor that affects patients' response to immunosuppressive agents. This article will discuss the role of immunosuppressive cells and non-cellular components in the immune process to provide ideas for the next research direction of EC.

Chinese medicine self-assembly nano-strategies（CSAN） is to utilize the self-assembly property of Chinese medicine components， so that the Chinese medicine components can self-assemble to form structurally stable nano-preparations through non-covalent interactions. The formation of Chinese medicine self-assembly nano-preparations is often a synergistic result of a variety of non-covalent interactions， and many Chinese medicine monomers are susceptible to self-assembly due to their structural characteristics， and the phenomenon of self-assembly of Chinese medicine is also common in the decoction of single or compound Chinese medicine， which has attracted the attention of researchers. It is found that CSAN can improve the solubility and bioavailability of active components in Chinese medicine， which is of positive significance for the development and application of insoluble components of Chinese medicine. The self-assembly phenomenon of Chinese medicine decoction is closely related to the therapeutic efficacy， and the study of self-assembly phenomenon of Chinese medicine will bring a new perspective for the explanation of the mechanism of Chinese medicine decoction. At the same time， traditional Chinese medicine（TCM） has unique advantages in the field of anti-tumor. The application of CSAN in the field of oncology can not only exert the anti-tumor effect of the active components of Chinese medicine directly， but also act as a natural nano-carrier to carry chemotherapy drugs for combination chemotherapy， improve the targeting of drugs， enhance the anti-tumor efficacy， and reduce the side effects of chemotherapy， which has excellent anti-tumor potential. The preparation method of Chinese medicine self-assembly nano-preparations is simple， low cost， and has better safety than traditional nano-preparations， which is conducive to the promotion of the clinical transformation of nano-preparations， and also helps to provide new strategies and perspectives for promoting the modernization of TCM. Therefore， based on a large number of researches in this field in recent years， this paper reviewed the formation mechanism， different assembly forms， formation conditions and stability of Chinese medicine self-assembly nano-preparations by searching databases such as China national knowledge infrastructure（CNKI）， PubMed， WanFang data and VIP， and summarized the application of CSAN in different tumor therapies， providing a reference for further research on CSAN.

Objective:To investigate the effect and mechanism of longikaurin A (LK-A) on glioma proliferation.Methods:Resuscitated frozen human glioma cell lines U87 and SNB19 were in vitro sub-cultured; CCK-8 assay was used to detect the cell activity changes 24 and 48 h after 0, 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 4.0, 6.0, 8.0 μmol/L LK-A. Edu proliferation assay was used to detect the changes in number of Edu positive cells after 0, 1, 2 and 3 μmol/L LK-A, and cell clonal formation assay was used to detect the changes in number of cell colony formation after 0, 0.1, 0.2 and 0.3 μmol/L LK-A. Flow cytometry was used to detect the changes in cell cycle proportion after 0, 1, 2 and 3 μmol/L LK-A. Expression changes of proliferation-related proteins (Ki-67 and c-Myc) and cell cycle-related proteins (Cyclin B1, Cyclin D1, cyclin dependent kinase 1 [CDKl]) and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway-related proteins were detected by Western blotting after 0, 1, 2, and 3 μmol/L LK-A. Results:U87 and SNB19 cell viabilities decreased gradually after being treated with 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 4.0, 6.0, and 8.0 μmol/L LK-A for 24 and 48 h compared with those with 0 μmol/L LK-A, with significant differences ( P<0.05); cell viability was dose-dependent. The number of Edu positive cells in U87 and SNB19 after being treated with 2, and 3 μmol/L LK-A was significantly decreased compared with that with 0 μmol/L LK-A ( P<0.05). The colony formation number of U87 and SNB19 cells after being treated with 0.1, 0.2 and 0.3 μmol/L LK-A decreased significantly compared with that with 0 μmol/L LK-A ( P<0.05). The proportion of U87 and SNB19 cells at G2/M phase after being treated with 2 and 3 μmol/L LK-A were increased significantly compared with that with 0 μmol/L LK-A ( P<0.05). The Ki-67, c-Myc, Cyclin B1, CDK1, p-PI3K and p-AKT expressions were significantly decreased, while Cyclin D1 expression was significantly increased in U87 and SNB19 cells after being treated with 2 and 3 μmol/L LK-A compared with those with 0 μmol/L LK-A ( P<0.05). Conclusion:LK-A can inhibit the glioma proliferation and arrest glioma at G2/M phase, with dose-dependent manner; the mechanism is related to inhibition of PI3K/AKT signaling pathway by LK-A.

The outbreak and spread of coronavirus disease 2019 (COVID-19) highlighted the importance and urgency of the research and development of therapeutic drugs. Very early into the COVID-19 pandemic, China has begun developing drugs, with some notable progress. Herein, we summarizes the anti-COVID-19 drugs and promising drug candidates originally developed and researched in China. Furthermore, we discussed the developmental prospects, mechanisms of action, and advantages and disadvantages of the anti-COVID-19 drugs in development, with the aim to contribute to the rational use of drugs in COVID-19 treatment and more effective development of new drugs against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the variants. Neutralizing antibody is an effective approach to overcome COVID-19. However, drug resistance induced by rapid virus mutation will likely to challenge neutralizing antibodies. Taking into account current epidemic trends, small molecule drugs have a crucial role in fighting COVID-19 due to their significant advantage of convenient administration and affordable and broad-spectrum. Traditional Chinese medicines, including natural products and traditional Chinese medicine prescriptions, contribute to the treatment of COVID-19 due to their unique mechanism of action. Currently, the research and development of Chinese anti-COVID-19 drugs have led to some promising achievements, thus prompting us to expect even more rapidly available solutions.

Objective:Clinicopathological features, treatment and prognosis of urinary and male reproductive system soft tissue sarcoma (STS) and sarcomatoid carcinoma in adults were compared.Methods:A retrospective analysis was performed on the clinical data of 73 patients with STS and 15 patients with sarcomatoid carcinoma in adult urinary and male reproductive system in the First Affiliated Hospital of Zhengzhou University. There were 59 males and 14 females in STS group, with a median age of 41 (18-78)years old. The maximum tumor diameter ranged from 0.5 to 19.0 cm. The primary tumors were located in testis and peritesticular (23 cases), kidney (23 cases), prostate (15 cases), bladder (8 cases), ureter(3 cases), other parts(1 case). There were 18 cases of lymph node metastasis and 8 cases of distant metastasis. Among 73 patients with STS, 66 patients underwent surgical resection, of which 31 patients underwent radical resection. Among the 66 patients who underwent surgery, 3 patients received neoadjuvant chemotherapy; 22 patients received adjuvant chemotherapy; 5 patients were treated with adjuvant radiotherapy. Among 7 patients with STS did not receive surgical treatment, 2 patients received radiotherapy combined with chemotherapy, 2 patients received chemotherapy alone, and 3 patients received symptomatic support treatment.There were 11 males and 4 females in sarcomatoid carcinoma group, with a median age of 65 (23 - 84)years old. The measurable tumor diameter ranged from 0.4 to 16.9 cm. The primary tumors were located in kidney (6 cases), bladder (5 cases), ureter(2 cases) and prostate(2 cases). There were 2 patients of lymph node metastasis and 4 patients of distant metastasis. Of the 15 patients with sarcomatoid carcinoma, 12 patients underwent surgical resection, of which 5 patients underwent radical resection. 2 patients were treated with adjuvant therapy after operation. Among the 12 patients who received surgical treatment, 2 patients had distant metastasis before operation, all of which originated from the kidney. Among the 3 patients without surgical treatment, 1 patients received systemic chemotherapy and 2 patients received symptomatic supportive treatment. There was no significant difference in gender, tumor maximum diameter, distant metastasis and operation, chemotherapy, radiotherapy and operation combined with chemotherapy ( P>0.05) and there were significant differences in age, tumor primary location and lymph node metastasis ( P<0.05) between STS and sarcomatoid carcinoma patients.The categorical variables of the two groups were compared by χ2.With Kaplan-Meier method for univariate survival analysis, the Cox was used for multivariate analysis. Results:The median follow-up time was 18.3(0.3-90.4) months.In STS group, there were 14 patients of synovial sarcoma, 11 patients of liposarcoma, 15 patients of rhabdomyosarcoma, 16 patients of leiomyosarcoma, 10 patients of other types, and 7 patients of spindle cell sarcoma without specific classification. Among 66 patients with STS, 8 patients recurred, 14 patients metastasized after operation, 4 patients recurred and metastasized after operation. The 7 patients without surgical treatment all progressed. Among the 10 patients of sarcomatoid carcinoma without distant metastasis before operation, 3 patients recurred and 3 patients metastasized after operation. Two patients of renal sarcomatoid carcinoma with distant metastasis were treated with nephrectomy and chemotherapy. One of them had overall survival (OS) up to 2 years, and one recurred 2 months after operation. The 3 patients without surgical treatment all progressed without remission. The median OS of STS patients were 59.3 (95% CI 24.1-94.5) months and that of sarcomatoid carcinoma patients were 8.7 (95% CI 6.1-11.2) months. The OS of STS patients were better than those of sarcomatoid carcinoma patients ( HR=2.874, 95% CI 1.118-7.386, P=0.022). Conclusions:The onset age of STS in adult urinary and male reproductive system was lower than that in sarcomatoid carcinoma. The primary lesions of STS were mainly in testis, peritesticular and kidney. The primary lesions of sarcomatoid carcinoma were mainly in kidney. Among STS, leiomyosarcoma was the most common type.STS and sarcomatoid carcinoma should be diagnosed and treated with surgery quickly, and systemic therapy should be performed for patients who cannot be treated with surgery.

Objective:To find the biomarkers that accurately predict the survival of patients with esophageal squamous cell carcinoma (ESCC).Methods:The immune related genes that were significantly related to the overall survival (OS) of patients with ESCC were screened from The Cancer Genome Atlas (TCGA) database to construct a prognostic risk score model. The prognoses of the high-risk and low-risk groups were compared by Kaplan-Meier method. The accuracy of the model was evaluated by the receiver operating characteristic (ROC) curve. Tumor tissue samples of 83 patients with pathological diagnosis of ESCC were collected from Anyang Cancer Hospital for external verification. Cox regression analysis was used to comprehensively evaluate the effects of prognostic risk score and various clinical characteristics on OS of patients with ESCC.Results:Seven immune-related genes that were significantly related to survival prognosis were selected from the TCGA database and included in the prognostic risk score model, which were S100A12, SLC40A1, FABP9, TNFSF10, IGHA2, IL1F10, and STC2. The 1- and 2-year survival rates of the low-risk group (40 cases) were 94.3% and 82.5%, respectively, while those of the high-risk group (40 cases) were 75.9% and 32.9%, respectively.The prognosis of the high-risk group was worse than that of the low-risk group ( P<0.001). The 83 external validation samples obtained consistent results by using the prognostic risk score model. The prognostic risk score was positively correlated with the content of CD4 + T lymphocytes in ESCC ( rs=0.259, P=0.020), but not correlated with the content of B lymphocytes, CD8 + T lymphocytes, neutrophils, macrophages or dendritic cells ( P>0.05). Conclusions:S100A12, SLC40A1, FABP9, TNFSF10, IGHA2, IL1F10, and STC2 were risk genes significantly associated with OS of patients with ESCC. The prognostic risk score was an independent prognostic factor for the OS of patients with ESCC, and it was correlated with the content of CD4 + T lymphocytes in ESCC tissue.

Objective:To find the biomarkers that accurately predict the survival of patients with esophageal squamous cell carcinoma (ESCC).Methods:The immune related genes that were significantly related to the overall survival (OS) of patients with ESCC were screened from The Cancer Genome Atlas (TCGA) database to construct a prognostic risk score model. The prognoses of the high-risk and low-risk groups were compared by Kaplan-Meier method. The accuracy of the model was evaluated by the receiver operating characteristic (ROC) curve. Tumor tissue samples of 83 patients with pathological diagnosis of ESCC were collected from Anyang Cancer Hospital for external verification. Cox regression analysis was used to comprehensively evaluate the effects of prognostic risk score and various clinical characteristics on OS of patients with ESCC.Results:Seven immune-related genes that were significantly related to survival prognosis were selected from the TCGA database and included in the prognostic risk score model, which were S100A12, SLC40A1, FABP9, TNFSF10, IGHA2, IL1F10, and STC2. The 1- and 2-year survival rates of the low-risk group (40 cases) were 94.3% and 82.5%, respectively, while those of the high-risk group (40 cases) were 75.9% and 32.9%, respectively.The prognosis of the high-risk group was worse than that of the low-risk group ( P<0.001). The 83 external validation samples obtained consistent results by using the prognostic risk score model. The prognostic risk score was positively correlated with the content of CD4 + T lymphocytes in ESCC ( rs=0.259, P=0.020), but not correlated with the content of B lymphocytes, CD8 + T lymphocytes, neutrophils, macrophages or dendritic cells ( P>0.05). Conclusions:S100A12, SLC40A1, FABP9, TNFSF10, IGHA2, IL1F10, and STC2 were risk genes significantly associated with OS of patients with ESCC. The prognostic risk score was an independent prognostic factor for the OS of patients with ESCC, and it was correlated with the content of CD4 + T lymphocytes in ESCC tissue.

Objective: To explore the expression of microRNA-17-5p (miR-17-5p) in esophageal squamous cell carcinoma (ESCC) and its effects on cell proliferation and invasion ability. Methods: Real-time quantitative PCR (RT-qPCR) was used to detect the miR-17-5p level in ESCC tissues and cells. MiR-17-5p inhibitor and negative control (NC) were transfected into EC9706 and TE1 cells, and miR-17-5p expression was examined by using RT-qPCR. Cell counting kit-8 (CCK-8) and EdU were conducted to detect cell proliferation and Transwell chamber was used to investigate cell invasion ability. Dual-luciferase reporter assay was used to detect the direct interaction of miR-17-5p and retinoblastoma-like protein-2 (RBL2). Western blot and RT-qPCR were used to detect the expression of RBL2 in ESCC tissues, respectively. Finally, the correlation between RBL2 and miR-17-5p was analyzed. Results: The miR-17-5p level in ESCC tissues was 4.222±0.392, significantly higher than 1.081±0.046 in normal esophageal epithelial tissues (P<0.001). The expressions of miR-17-5p in ESCC cells, including EC9706, Eca109, TE1, KYSE450, KYSE70 and KYSE520, were 13.84±1.266, 6.453±0.293, 11.41±0.520, 2.613±0.548, 5.251±0.239 and 4.251±0.195, respectively, all obviously higher than (1.007±0.079) in normal esophageal epithelial cell Het-1A (P<0.05). The miR-17-5p level in patients with ESCC Ⅲ~Ⅳ was 5.094±0.562, markedly higher than 2.934±0.364 in patients with ESCCⅠ~Ⅱ(P<0.01). Moreover, the miR-17-5p level in ESCC patients with lymph node metastasis was 5.523±0.634, markedly higher than 3.533±0.461 in ESCC patients without lymph node metastasis (P<0.05). The survival ratio of ESCC patients with higher miR-17-5p level was evidently lower than that of ESCC patients with lower miR-17-5p level (P<0.05). MiR-17-5p inhibitor significantly downregulated the miR-17-5p expression in EC9706 and TE1, which suppressed cell proliferation and invasion ability. Dual-luciferase reporter assay revealed that co-transfection of 3′ untranslated region-wild type (3′UTR-WT) of RBL2 and miR-17-5p mimic significantly reduced luciferase activity in EC9706 and TE1 cells (P<0.01), which implicated that RBL2 was the direct target of miR-17-5p. The result of western blot revealed that RBL2 protein expressions in miR-17-5p group of EC9706 and TE1 cells were 0.936±0.055 and 0.923±0.048, obviously higher than 0.087±0.019 and 0.102±0.010 in control group (P<0.001). The expression of RBL2 in ESCC tissues was 0.219±0.510, markedly lower than 0.983±0.324 in normal esophageal epithelial tissues (P<0.001). The miR-17-5p level exhibited a negative correlation with RBL2 level in ESCC tissues (r=-0.462, P<0.001). Downregulation of RBL2 reversed the miR-17-5p inhibitor induced suppression of cell proliferation and invasion ability. Conclusion MiR-17-5p participates in the carcinogenesis and development of ESCC, thus may be a potential therapeutic target for ESCC patients.

Objective To analyze the feasibility of totally no tube (TNT) in da Vinci robotic mediastinal mass surgery and its significance for fast track surgery. Methods A total of 79 patients receiving robotic mediastinal TNT surgery in the General Hospital of Shenyang Military Command from January 2016 to December 2017 were enrolled as a TNT group; 35 patients receiving robotic mediastinal surgery in General Hospital of Shenyang Military Command from January 2014 to December 2017 and 54 patients receiving thoracoscopic mediastinal surgery during the same period were enrolled as a non-TNT group and a video-assisted thoracoscopic surgery (VATS) group. The muscle relaxation and tracheal intubation/laryngeal masking time, operation time, intraoperative blood loss, postoperative ICU stay, postoperative hospital stay, postoperative visual analogue scale (VAS), hospitalization costs and postoperative complications and other related indicators were retrospectively analyzed among the three groups. Results Surgeries were successfully completed in 168 patients with no transfer to thoracotomy, serious complications (postoperative complications in 9 patients) or death during the perioperative period. All patients were discharged. Compared with the non-TNT group, the TNT group had significantly less muscle relaxation-tracheal intubation/laryngeal masking time, operation time, intraoperative blood loss, VAS pain score, ICU stay, postoperative hospital stay (P<0.01); there was no significant difference in the total cost of hospitalization between the two groups (P>0.05). Between the non-TNT group and the VATS group, there was no significant difference in time of muscle relaxation and tracheal intubation, operation time and ICU stay (P>0.05). The non-TNT group was superior to the VATS group in terms of intraoperative blood loss, VAS pain scores on the following day after operation, chest drainage volume 1-3 days postoperatively, postoperative catheterization time and postoperative hospital stay (P<0.05); but the cost of hospitalization in the non-TNT group was significantly higher (P=0.000). Conclusion The da Vinci robot is safe and feasible for the treatment of mediastinal masses. At the same time, TNT is also safe and reliable on the basis of robotic surgery which has many advantages such as better comfort, less pain, ICU stay and hospital stay as well as faster recovery.