Objective To establish an Apak gene stable and permanent knockout cell line using CRISPR/Cas9 system in human colon cancer cells ( HCT116 cells), and study the effect of Apak knock-out on p53 activity and apoptosis. Methods The lentiCRISPR v2-sgRNA Apak expression plasmid was co-transfected with lentivirus coated plasmids pSPAX2 and pMD2.G.The supernatant was collected, filtered, and used to infect HCT116 cells.The positive clones were screened out by puromycin culture and Western blot was used to detect Apak knockout cell lines.Luciferase reporter gene assay, flow cytometry analysis and colony formation assay were used to examine p53 activity and apoptosis of Apak knockout cells, respectively.Results Apak knockout HCT116 cell lines were generated in which p53 activity and apoptosis were increased,but the colony formation was decreased.Conclusion The Apak stable knockout cell lines of HCT116 are successfully generated by CRISPR/Cas9 system for further functional study.

Objective To evaluate the efficacy of stereotactic body radiation therapy (SBRT) for hepatic metastases from colorectal cancer,and to collect data for the application of this technique.Methods A total of 28 patients from No.306 Hospital of PLA,including 17 male and 11 female with median age of 63.8 (range from 31 to 86),were treated with SBRT for colorectal cancer with liver metastases with 54 lesions in total.The GTV,CTV and PTV were delineated above the enhanced CT scans acquired during normal quiet respiration.CTV was obtained by adding 5 mm isotropic margin from GTV,and PTV was obtained by adding 5 to 10 mm isotropic margin from CTV.Prescription dose line covered 50％-60％ of isodose curve at 3-6 Gy/fraction.The total dose was 39-45 Gy and the biologically equivalent doses(BED)was 50.7-65.3 Gy.The patients were followed-up beginning at 3 months after SBRT.The change in size of the lesion based on enhanced CT or MR scans was evaluated.Toxicity was evaluated and scored according to the RTOG criteria.Local control rate and survival rate were analysed.Results All patients completed the treatment.With median follow-up of 15.1 months (range frome 3 to 30 months),7 patients survived at the end of follow-up.The local control rate (LC) was 79.2％,and 1-and 2-year overall survival rate(OS) were 82.7％ and 48.6％,respectively.There was a close corelation between the size of lesion and the LC.The LC (PR + CR) was much better at the size of lesion less than 14 cm3 than that at the size more than 65 cm3(x2 =4.17,P＜0.05).When the size was more than 180 cm3,the LC was zero.Toxicity included fatigue (60.7％),grade 1 and 2 digestive system toxicity (28.6％),a transient grade 1 and 2 bone marrow suppression (46.4％) and a transient increase in transaminase(17.8％).No grade 3 toxicity and above and late toxicity were observed.Conclusions Stereotactic body radiation therapy could be suggested as the first choice for the selected patients who suffer form colorectal liver metastases,especially for those who cannot undergo surgery.

Objective To measure the organ weights , blood physiological and biochemical parameters , and immune function of Specific Pathogen Free ( SPF) Rag2 knockout ( KO) mice.Methods Rag2 knockout mice were selected at five and ten weeks , and the organ weights , blood physiological and biochemical parameters were observed .The percentages of CD3+, CD4+, CD8+, CD19+, B220+, NK1.1+, and CD11b+were checked by FCM in Rag2 KO mice at six week of age in terms of its T , B lymphocyte function and NK cell activity .Results Among the same sex group of NOD/SCID mice, the weights of brain, lung, spleen, liver, heart, kidney, and the levels of TBIL, WBC in 10 weeks of age are higher than 5 weeks.At the same age, the weights of heart, kidney, liver, spleen, and the levels of AST, ALP, A/G, GLU, PLT, PCT, WBC, and LYM%in male mice are higher than females.The Rag2 KO mice lacks T cells(0.36 ±0.15)%、CD4+T cells(0.21 ±0.06)%、CD8+T cells(0.23 ±0.07)%、CD19+B cells(0.28 ±0.04)%、B220+B cells(2.03 ±0.42)%).The percentage of NK cells is(24.13 ±3.62)%, and the percentage of granulocytes is (57.20 ±3.85)%.Conclusion The study suggests that the organ weights , blood physiological and biochemical parameters are affected by age and gender in Rag2 KO mice, which main biological characteristics are similar with C 57BL/6J mice.The Rag2 KO mice show the deficiency of T , B cells function .

ObjectiveTo investigate the role of platelet-derived growth factor(PDGF) and its receptor in the development of hypertrophic scar. MethodsThe expression of PDGF and its receptor were detected in biopsy specimens of 9 pieces of normal skin, 7 pieces of granulation tissue of burn wound and 34 pieces of hypertrophic scar by immunohistochemical staining using specific polyclonal antibodies.ResultsPDGF and its receptor markedly increased in granulation tissue and hypertrophic scars, reaching the peak in the hypertrophic scars within 6 months and then decreased after the peak, whereas PDGF and its receptor expressed weakly in only a few normal skin specimens, and the differences were significant(P<0.05).ConclusionsThe increasing expression of PDGF and its receptor may be related to the development of hypertrophic scar.

Alzheimer's disease (AD) is characterized byβ-amyloid (Aβ) deposition and tauprotein hyperphosphorylation, whereas its pathogenesis has not been fully known so far. The metabolism of Aβand tau protein is critically affected by autophagy. In the early phase of AD, Aβand tau protein can induce themselves to be eliminated via mTOR-dependent and independent autophagy pathways. In addition, transcription factors EB and apolipoprotein E4 also regulate autophagy and thus participate in the metabolism of Aβand tau protein, affecting AD progression. This review summarized the roles of autophagy in the metabolism of Aβand tau protein and the autophagy regulators closely related to AD in the recent studies.

WHO Guideline on Country Pharmaceutical Pricing Policies,published in 2020,outlines 10 commonly used pricing methods internationally.However,due to the unique composition of costs for traditional Chinese medicine(TCM),directly applying pricing methods designed for chemical and biological drugs may lead to discrepancies.Currently,in China,drug pricing primarily in-volves internal reference pricing,tender negotiation pricing,promoting the use of quality-assured generic and biosimilar drugs,and centralized procurement.It systematically analyzes various pricing methods and identifies their applicability and underlying reasons concerning the pricing of TCMwithin the medical insurance system.The method of value-based pricing and increasing price transparency has advantages for the pricing regulation of traditional Chinese medicine under medical insurance.

BACKGROUNDInsulin resistance (IR) plays an important pathophysiological role in the development of diabetes, dyslipidemia, hypertension, and cardiovascular disease. Moreover, IR can occur even in non-obese people without diabetes. However, direct detection of IR is complicated. In order to find a simple surrogate marker of IR early in non-obese people, we investigate the association of commonly-used biochemical markers (liver enzymes and lipid profiles) with IR in urban middle-aged and older non-obese Chinese without diabetes.

METHODSThis cross-sectional study included 1 987 subjects (1 473 women). Fasting blood samples were collected for measurement of glucose, insulin, liver enzymes, lipid profiles and creatinine. Subjects whose homeostasis model of assessment-IR (HOMA-IR) index values exceeded the 75th percentile (2.67 for women and 2.48 for men) of the population were considered to have IR. The area under the receiver operating characteristic curve (ROC) was used to compare the power of potential markers in identifying IR.

RESULTSTriglycerides (TG) and ratio of TG to high-density lipoprotein cholesterol (TG/HDL-C) discriminated IR better than other indexes for both sexes; areas under the receiver operating characteristic (ROC) curves (AUC) values were 0.770 (95% confidence interval 0.733-0.807) and 0.772 (0.736-0.809), respectively, for women and 0.754 (0.664-0.844) and 0.756 (0.672-0.840), respectively, for men. To identify IR, the optimal cut-offs for TG and TG/HDL-C ratio were 1.315 mmol/L (sensitivity 74.3%, specificity 71.0%) and 0.873 (sensitivity 70.1%, specificity 73.4%), respectively, for women, and 1.275 mmol/L (sensitivity 66.7%, specificity 74.4%) and 0.812 (sensitivity 75.8%, specificity 69.2%), respectively, for men.

CONCLUSIONTG and TG/HDL-C ratio could be used to identify IR in urban middle-aged and older non-obese Chinese without diabetes.

Aged ; Alanine Transaminase ; blood ; Aspartate Aminotransferases ; blood ; Cholesterol, HDL ; blood ; Cross-Sectional Studies ; Diabetes Mellitus ; blood ; Female ; Humans ; Insulin Resistance ; physiology ; Liver ; enzymology ; Male ; Middle Aged ; Triglycerides ; blood