Identifying novel therapeutic targets in gastric cancer using genome-wide CRISPR-Cas9 screening.

Genome-scale CRISPR-Cas9 screening technology is a powerful tool to systematically identify genes essential for cancer cell survival. Herein, TKOv3, a genome-scale CRISPR-Cas9 knock-out library, was screened in the gastric cancer (GC) cells, and relevant validation experiments were performed. We obtained 854 essential genes for the AGS cell line, and 184 were novel essential genes. After knocking down essential genes: SPC25, DHX37, ABCE1, SNRPB, TOP3A, RUVBL1, CIT, TACC3 and MTBP, cell viability and proliferation were significantly decreased. Then, we analysed the detected essential genes at different time points and proved more characteristic genes might appear with the extension of selection. After progressive selection using a series of open datasets, 41 essential genes were identified as potential drug targets. Among them, methyltransferase 1 (METTL1) was over expressed in GC tissues. High METTL1 expression was associated with poor prognosis among 3 of 6 GC cohorts. Furthermore, GC cells growth was significantly inhibited after the down-regulation of METTL1 in vitro and in vivo. Function analysis revealed that METTL1 might play a role in the cell cycle through AKT/STAT3 pathways. In conclusion, compared with existing genome-scale screenings, we obtained 184 novel essential genes. Among them, METTL1 was validated as a potential therapeutic target of GC.

The Chinese Society of Hepatology position statement on the redefinition of fatty liver disease.

Fatty liver disease associated with metabolic dysfunction is of increasing concern in mainland China, the world's most populous country. The incidence of fatty liver disease is highest in China, surpassing the incidence in European countries and the USA. An international consensus panel recently published an influential report recommending a novel definition of fatty liver disease associated with metabolic dysfunction. This recommendation includes a switch in name from non-alcoholic fatty liver disease (NAFLD) to metabolic (dysfunction)-associated fatty liver disease (MAFLD) and adoption of a set of positive criteria for disease diagnosis that are independent of alcohol intake or other liver diseases. Given the unique importance of this proposal, the Chinese Society of Hepatology (CSH) invited leading hepatologists and gastroenterologists representing their respective provinces and cities to reach consensus on alternative definitions for fatty liver disease from a national perspective. The CSH endorses the proposed change from NAFLD to MAFLD (supported by 95.45% of participants). We expect that the new definition will result in substantial improvements in health care for patients and advance disease awareness, public health policy, and political, scientific and funding outcomes for MAFLD in China.

Effect of artificial liver blood purification treatment on the survival of critical ill COVID-19 patients.

Our aim was to investigate the effect of artificial liver blood purification treatment on the survival of severe/critical patients with coronavirus disease 2019 (COVID-19). A total of 101 severe and critical patients with coronavirus SARS-CoV-2 infection were enrolled in this open, case-control, multicenter, prospective study. According to the patients' and their families' willingness, they were divided into two groups. One was named the treatment group, in which the patients received artificial liver therapy plus comprehensive treatment (n = 50), while the other was named the control group, in which the patients received only comprehensive treatment (n = 51). Clinical data and laboratory examinations, as well as the 28-day mortality rate, were collected and analyzed. Baseline data comparisons on average age, sex, pre-treatment morbidity, initial symptoms, vital signs, pneumonia severity index score, blood routine examination and biochemistry indices etc. showed no difference between the two groups. Cytokine storm was detected, with a significant increase of serum interleukin-6 (IL-6) level. The serum IL-6 level decreased from 119.94 to 20.49 pg/mL in the treatment group and increased from 40.42 to 50.81 pg/mL in the control group (P < .05), indicating that artificial liver therapy significantly decreased serum IL-6. The median duration of viral nucleic acid persistence was 19 days in the treatment group (ranging from 6 to 67 days) and 17 days in the control group (ranging from 3 to 68 days), no significant difference was observed (P = .36). As of 28-day follow-up,17 patients in the treatment group experienced a median weaning time of 24 days, while 11 patients in the control group experienced a median weaning time of 35 days, with no significant difference between the two groups (P = .33). The 28-day mortality rates were 16% (8/50) in the treatment group and 50.98% (26/51) in the control group, with a significant difference (z = 3.70, P < .001). Cytokine storm is a key factor in the intensification of COVID-19 pneumonia. The artificial liver therapy blocks the cytokine storm by clearing inflammatory mediators, thus preventing severe cases from progressing to critically ill stages and markedly reducing short-term mortality.

Highly biocompatible and recyclable biomimetic nanoparticles for antibiotic-resistant bacteria infection.

Increasing number of resistant bacteria have emerged with the overuse of antibiotics, which indicates that the bacterial infection has become a global challenge. Furthermore, the pollution of antibiotics to the environment has become a serious threat to public health. It is known that toxins produced by bacteria are the main cause of bacterial infections. Photothermal therapy is an effective antibacterial approach. However, the photothermal reagents cannot eliminate bacterial toxins, and even some anti-bacterial materials are toxic. Here, we synthesized a biomimetic recycled nanoparticle, red blood cell (RBC) membrane-coated Fe3O4 nanoparticles (RBC@Fe3O4), as an antibacterial agent. The RBC@Fe3O4 nanoparticles act as nano-sponges to trap toxins and then kill them all with a photothermal effect. We can describe this process simply as a battle between two armies. Our strategy is to disarm the "enemy" so that we can easily kill the "enemy" who has no power, which results in enhancing the bactericidal efficacy. The toxin of methicillin-resistant Staphylococcus aureus (MRSA) was absorbed by RBC@Fe3O4in vitro. In addition, in vivo studies proved that the RBC@Fe3O4 nanoparticles confer obvious survival benefits against toxin-induced lethality by absorbing the toxin of MRSA. Furthermore, using a mouse model of MRSA wound infection, the RBC@Fe3O4 nanoparticles with laser irradiation were found to have a superior wound-healing effect. Simultaneously, the RBC@Fe3O4 nanoparticles could be recycled in a simple way without affecting the bactericidal efficacy. The highly biocompatible and recyclable RBC@Fe3O4 biomimetic nanoparticles based on photothermal therapy and bacterial toxin adsorption strategy are promising for treating bacterial infections.

Morbidity and Mortality of Nosocomial Infection after Cardiovascular Surgery: A Report of 1606 Cases.

Nosocomial infection (NI) is one of the most significant complications arising after open heart surgery, and leads to increased mortality, hospitalization time and health resource allocation. This study investigated the morbidity, mortality, and independent risk factors associated with NI following open heart surgery. We retrospectively surveyed the records of 1606 consecutive cardiovascular surgical patients to identify those that developed NI. The NI selection criteria were based on the Centers for Disease Control and Prevention (CDC) guidelines. The term NI encompasses surgical site infection (SSI), central venous catheter-related infection (CVCRI), urinary tract infection (UTI), respiratory tract infection and pneumonia (RTIP), as well as other types of infections. Of 1606 cardiovascular surgery patients, 125 developed NI (7.8%, 125/1606). The rates of NI following surgery for congenital malformation, valve replacement, and coronary artery bypass graft were 2.6% (15/587), 5.5% (26/473) and 13.6% (32/236), respectively. The NI rate following surgical repair of aortic aneurysm or dissection was 16.8% (52/310). Increased risk of NI was detected for patients with a prior preoperative stay ≥3 days (OR=2.11, 95% CI=1.39-3.20), diabetes (OR=2.00, 95%=CI 1.26-3.20), length of surgery ≥6 h (OR=2.26, 95% CI=1.47-3.47), or postoperative cerebrovascular accident (OR=4.08, 95% CI=1.79-9.29). Greater attention should be paid toward compliance with ventilator and catheter regulations in order to decrease NI morbidity and mortality following cardiovascular procedures.

1810011o10 Rik Inhibits the Antitumor Effect of Intratumoral CD8+ T Cells through Suppression of Notch2 Pathway in a Murine Hepatocellular Carcinoma Model.

The mechanisms by which tumor-responsive CD8+ T cells are regulated are important for understanding the tumor immunity and for developing new therapeutic strategies. In current study, we identified the expression of 1810011o10 Rik, which is the homolog of human thyroid cancer 1, in intratumoral activated CD8+ T cells in a murine hepatocellular carcinoma (HCC) implantation model. To investigate the role of 1810011o10 Rik in the regulation of antitumor activity of CD8+ T cells, normal CD8+ T cells were transduced with 1810011o10 Rik-expressing lentiviruses. Although 1810011o10 Rik overexpression did not influence agonistic antibody-induced CD8+ T cell activation in vitro, it inhibited the cytotoxic efficacy of CD8+ T cells on HCC cells in vivo. 1810011o10 Rik overexpression impeded CD8+ T cell-mediated HCC cell apoptosis and favored tumor cell growth in vivo. Further investigation revealed that 1810011o10 Rik blocked the nuclear translocation of Notch2 intracellular domain, which is crucial for CD8+ T cell activity. Furthermore, a brief in vitro experiment suggested that both antigen-presenting cells and TGF-ß might be necessary for the upregulation of Rik expression in activated CD8+ T cells. In general, our study disclosed a novel mechanism underlying the negative regulation of antitumor CD8+ T cells during HCC progression.

Mutation of rpoS gene decreased resistance to environmental stresses, synthesis of extracellular products and virulence of Vibrio anguillarum.

Vibrio anguillarum is a gram-negative halophilic bacterium that causes vibriosis in marine fish, freshwater fish and other aquatic animals. Bacteria have developed strategies to survive in harsh environments. The alternative sigma factor, RpoS (sigma(S)), plays a key role in surviving under stress conditions in some gram-negative bacteria. An rpoS mutant of pathogenic V. anguillarum W-1 was constructed by homologous recombination. The sensitivity of the rpoS mutant to osmotic stress [2.4 M NaCl in artificial seawater (ASW)] did not change obviously, but the sensitivity of the rpoS mutant to high temperature (45 degrees C in ASW), UV-irradiation and oxidative stress (5 mM H(2)O(2) in ASW) increased 33-fold, sixfold and 10-fold, respectively. The production of extracellular phospholipase, diastase, lipase, caseinase, hemolysin, catalase and protease of the rpoS mutant decreased markedly compared with those of the wild-type strain. Virulence of the rpoS mutant strain was also decreased when it was inoculated intraperitoneally into zebra fish; the lethal dose 50% of the wild type and the mutant was 8.66 x 10(4) and 2.55 x 10(6) CFU per fish, respectively. These results indicated that the RpoS of V. anguillarum plays important roles in bacterial adaptation to environmental stresses and its pathogenicity.

Effect of antisense oligodeoxynucleotide of telomerase RNA on telomerase activity and cell apoptosis in human colon cancer.

AIM: To explore the effect of antisense oligodeoxynucleotide (As-ODN) of telomerase RNA on telomerase activity and cell apoptosis in human colon cancer. METHODS: As-ODN was transfected into SW480 cells by liposomal transfection reagent. Telomerase activity of SW480 cells was examined by telomeric repeat amplification protocol (TRAP) and enzyme-linked immunosorbent assay (ELISA). Apoptosis was analyzed by morphology and flow cytometry. RESULTS: The telomerase activity in SW480 cells transfected with 1.0 micromol/L of As-ODN for 2-5 days, was significantly decreased in a time-dependent manner, and the cells underwent apoptosis. The missense ODN (Ms-ODN) and the control group transfected with SW480 cells did not show these changes. CONCLUSION: As-ODN can specifically inhibit the telomerase activity of SW480 cells and induce apoptosis.

Telomerase activity and cell apoptosis in colon cancer cell by human telomerase reverse transcriptase gene antisense oligodeoxynucleotide.

AIM: To evaluate the effect of human telomerase reverse transcriptase (hTERT) gene antisense oligodeoxynucleotide (As-ODN) on telomerase activity and cell apoptosis in colon cancer cell line SW480. METHODS: As-ODN was transfected into cells SW480 by liposomal transfection. Cultured cells were divided into three groups: ASODN (5'GGAGCGCGCGGCATCGCGGG-3'), sense oligodeoxynucleotide (5'-CCCGCGATGCCGCGCGCTCC-3', S-ODN) and control. The concentration of oligodeoxynucleotide and liposome was 10 micromol/L and 16 mg/L, respectively. The activity of telomerase was examined by telomeric repeat amplification protocol (TRAP)-enzyme-linked immunosorbent assay (ELISA), and cell apoptosis was observed by morphology and flow cytometry in each group. RESULTS: Telomerase activity began to be down-regulated or inhibited when cells SW480 were treated with As-ODN for 72 h, and cell apoptosis was induced. CONCLUSION: It is suggested that hTERT As-ODN might specially inhibit the activity of telomerase in colon cancer cells and it is further proved that the hTERT gene has a significant correlation with telomerase activity. Further evidence is needed to prove whether hTERT As-ODN is a potential tool for the treatment of colon cancer.

Expression and significance of PTEN, hypoxia-inducible factor-1 alpha in colorectal adenoma and adenocarcinoma.

AIM: To investigate the expression and significance of PTEN, hypoxia-inducible factor-1 alpha (HIF-1alpha), and targeting gene VEGF during colorectal carciogenesis. METHODS: Total 71 cases colorectal neoplasms (9 cases of colorectal adenoma and 62 colorectal adenocarcinoma) were formalin fixed and paraffin-embedded, and all specimens were evaluated for PTEN mRNA, HIF-1alpha mRNA and VEGF protein expression. PTEN mRNA, HIF-1alpha mRNA were detected by in situ hybridization. VEGF protein was identified by citrate-microwave SP immunohistochemical method. RESULTS: There were significant differences in PTEN, HIF-1alpha and VEGF expression between colorectal adenomas and colorectal adenocarcinoma (P<0.05). The level of PTEN expression decreased as the pathologic stage increased. Conversely, HIF-1alpha and VEGF expression increased with the Dukes stage as follows: stage A (0.1029+/-0.0457: 0.1207+/-0.0436), stage B (0.1656+/-0.0329: 0.1572+/-0.0514), and stage C+D (0.2335+/-0.0748: 0.2219+/-0.0803). For PTEN expression, there was a significant difference among Dukes stage A, B, and C+D, and the level of PTEN expression was found to be significant higher in Dukes stage A or B than that of Dukes stage C or D. For HIF-1alpha expression, there was a significant difference between Dukes stage A and B, and the level of HIF-1alpha expression was found to be significantly higher in Dukes stage C+D than that of Dukes stage A or B. The VEGF expression had similar results as HIF-1alpha expression. In colorectal adenocarcinoma, decreased levels of PTEN were significantly associated with increased expression of HIF-1alpha mRNA (r=-0.36, P<0.05) and VEGF protein (r=-0.48, P<0.05) respectively. The levels of HIF-1 were positively correlated with VEGF expression (r=0.71, P<0.01). CONCLUSION: Loss of PTEN expression and increased levels of HIF-1alpha and VEGF may play an important role in carcinogenesis and progression of colorectal adenocarcinoma.

Mast cell density and the context of clinicopathological parameters and expression of p185, estrogen receptor, and proliferating cell nuclear antigen in gastric carcinoma.

AIM: To investigate the relationship between the mast cell density (MCD) and the context of clinicopathological parameters and expression of p185, estrogen receptor (ER), and proliferating cell nuclear antigen (PCNA) in gastric carcinoma. METHODS: Mast cell, p185, ER, and PCNA were detected using immunohistochemical S-P labeling method. Mast cell was counted in tissue of gastric carcinoma and regional lymph nodes respectively, and involved lymph nodes (ILN) were examined as usual. RESULTS: MCD was significantly related to both age and depth of penetration (chi(2)=4.688,P<0.05 for age and chi(2)=9.350, P<0.01 for depth of penetration) between MCD>21/0.03 mm(2) and MCD< or =21/0.03 mm(2) in 100 patients; MCD in 1-6 ILN group patients was significantly higher than that in 7-15 ILN or >15 ILN group patients (u=6.881, 8.055, P<0.01); There were significant differences intergroup in positive expression rate of p185, ER and PCNA between MCD>21/0.03 mm(2) and MCD< or =21/0.03 mm(2) in 100 patients. CONCLUSION: Mast cell may have effect on inhibiting invasive growth of tumor, especially in the aged patients; The number of mast cells, in certain degree, may predicate the number of involved lymph nodes, which is valuable for assessment of prognosis; MCD was related to the expression of p185, ER, and PCNA in gastric carcinoma. It suggests that mast cell accumulation may inhibit the proliferation and the dissemination of the gastric carcinoma.