Objective Hyperglycaemia and hyperlipid are common in clinical cardiovascular disease .The article was to ob-serve the change of endothelial and the expression of Rho-kinase in the aorta of rats with hyperglycaemia and hyperlipid and the rela-tionship between them .Methods Healthy male Wistar rats were randomly divided into 3 groups:5 as normal control group , 10 as model group and 10 as drug intervention group .The latter two groups were given high-fat diets for 5 weeks after tail vein injection of streptozocin (STZ) to establish hyperglycaemia and hyperlipid models , then fasudil (inhibitor of Rho-kinases) or saline were respec-tively given intraperitoneally for 4 weeks.Venous blood glucose (VBG), blood lipid levels, ET-1 and eNOS were tested, along with the expression of MYPT-1(the protein substrates of ROCK)、mRNA of RhoA and ROCK. Results Compared with normal control group, the levels of VBG、ET-1、MYPT-1、mRNA of ROCK and Rho A were increased in model group[(6.18±1.14)mmol/L vs (26.36± 3.25)mmol/L,(72.13±6.13 )pg/mL vs (88.22±4.61) pg/mL, (0.54±0.08) v s (1.28±0.17),(0.16±0.20) vs (0.83± 0.12),(0.40±0.18) vs (0.78±0.13),P<0.05].TG、TC、LDL were also higher in model group(P<0.05), while eNOS level was lower in model group than in control group[(24.42±2.13)U/L sv (17.36±1.58)U/L,P<0.05].Compared with model group, the levels of VBG[(17.70±2.69)mmol/L]、TG、TC、LDL, ET-1 [(75.03±2.50)pg/mL]、MYPT-1(0.74±0.01)、mRNA of ROCK(04.0±0.08) and Rho A (0.25±0.07)in drug intervention group were lower(P<0.05), while eNOS level (0.74±0.11) was higher(P<0.05) in drug intervention group than in model group . Conclusion Endothelial dysfunction and over-expression of Rho kinase can be found in hyperglycaemia and hyperlipid rat models .Fa-sudil could improve endothelial function .

BACKGROUND: Exercise has been proved to accelerate the proliferation of intervertebral disc cells and extracellular matrix production in healthy rats. For the degenerative intervertebral disc, whether exercise also has positive effects on its cell proliferation, extracellular matrix production or pain relief remains unclear. OBJECTIVE: To investigate the effect of exercise on the extracellular matrix production in a rat model of intervertebral disc degeneration.METHODS: A rat model of intervertebral disc degeneration was prepared by Freund's complete adjuvant injection into the intervertebral disc at L5-6 levels. Then, the model rats were allowed to have a rest for 2 weeks. All rats were then randomly divided into exercise and control groups. Rats in the exercise group were forced to run every day, while the controls allowed free activities in the cage. The behavioral tests were performed at 7, 14, 28, 42, 56 and 70 days after modeling; meanwhile, the intervertebral disc samples were collected used for alcian blue staining and immunohistochemical staining to detect the levels of proteoglycan, aggrecan and collagen type Ⅱ in the intervertebral disc cells, respectively.RESULTS AND CONCLUSION: Vocalization threshold on the rat back of punctured disc was significantly decreased, while grooming and wet-dog shaking were significantly increased at 7 days after modeling compared with the baseline (P < 0.05), suggesting that Freund's complete adjuvant injection successfully induces disc degeneration, hyperalgesia and abnormal behaviors. Further, the vocalization threshold and wet-dog shaking in the exercise group showed significant improvement compared with the control group after 14 days of exercise (P < 0.05), while the grooming was significantly reduced until the 28th day (P < 0.01), indicating that exercise can alleviate pain caused by disc degeneration in model rats. At 21 days after modeling, the levels of proteoglycan, aggrecan and collagen type Ⅱ in the nucleus pulposus and annulus fibrosus were significantly decreased compared with the baseline (P < 0.01), indicating the occurrence of disc degeneration. After 14 days of training, the levels of proteoglycan, aggrecan, and collagen type Ⅱ in the nucleus pulposus and annulus fibrosus in the exercise group were significantly increased compared with the control group (P < 0.01). Moreover, after 8-week exercise, the level of proteoglycan in the nucleus pulposus and annulus fibrosus in the exercise group was increased by 4-5 times compared with the control group, and levels of aggrecan and collagen type Ⅱ in the nucleus pulposus in the exercise group also was increased by 3-4 times compared with the control group. To conclude, exercise can promote extracellular matrix increased by production by increasing the levels of proteoglycan, aggrecan, and collagen type II in the degenerative intervertebral disc.

Objective To explore the effect and mechanism of microRNA (miR)-17-5p on the invasion and metastasis of hepatocellular carcinoma (HCC) cells. Methods Expression of miR-17-5p in the normal human L-02 hepatocyte and QGY-7703 HCC cells was detected by RT-PCR. QGY-7703 HCC cells were transfected by miR-17-5p mimic and mimic control respectively. Influence of miR-17-5p on the invasion and metastasis ability of HCC cells was detected using Transwell assay and scratch test. Target gene of miR-17-5p was confirmed by bioinformatic analysis, and its expression in HCC cells was detected by Western blot. After siRNA silenced by target gene, the invasion and metastasis ability of HCC cells were observed. Comparison of microRNA in the two kinds of cells was conducted by t test. Results Expression level of miR-17-5p in HCC cells was 0.16±0.04, significantly lower than 1.01±0.19 in normal L-02 hepatocytes (t=-9.67, P<0.05). Number of trans-membrane cells and metastasis rate of HCC cells transfected by miR-17-5p mimic were respectively 36±4 and (5.37±0.15) mm/d, significantly lower than 62±7 and (7.50±0.01) mm/d of control group (t=-15.40, -32.00; P<0.05). Bioinformatic analysis showed that AKT3 was the key target gene of miR-17-5p, and the expression of AKT3 in HCC cells was obviously higher than that of normal hepatocyte. Number of trans-membrane cells and metastasis rate of HCC cells transfected by siRNA-AKT3 were respectively 13±3 and (4.13±0.15) mm/d, significantly lower than 58±3 and (7.23±0.25) mm/d of control group (t=-17.88, -53.69; P<0.05). Conclusion miR-17-5p inhibits the invasion and metastasis ability of HCC cells through targeting effect on AKT3.

OBJECTIVE: To compare the accuracy of five warfarin-dosing algorithms and warfarin stable dose model (2.5 mg/day) for Shandong population. METHODS: One hundred and twenty five patients who achieved stable warfarin dose were enrolled. Clinical and genetic data were used to evaluate the value of each algorithm by calculating the percentage of patients whose predicted warfarin dose was within 20% of the actual stable therapeutic dose and mean absolute error (MAE). RESULTS: The frequency of patients with CYP2C9*1/*1, CYP2C9*1/*3 and CYP2C9*1/*2 genotype was 92.00%, 7.20%, 0.80%, respectively. That of VKORC1-1639 AA, AG and GG genotype was 82.40%, 15.20%, 2.40%, respectively. CYP4F2*1/*1, *1/*3, *3/*3 genotype was 50.40%, 39.20%, 10.40%, respectively. With the same genotypes for other loci, patients who carried at least one VKORC1-16398G mutant allele had increased warfarin stable daily dose compared with VKORC1-1639AA. Compared with CYP4F2*1/*1, those carrying at least one CYP4F2*3 mutant allele had warfarin stable daily dose increased by 5.9%-13.00%. The percentage of ideal prediction calculated from IWPC model (59.20%), Huang model (57.60%) and Ohno model (52.80%) were higher than others. The MAE were 0.35 (95%CI: 0.11-0.49), 0.15 (95%CI: 0.10-0.32), 0.39 (95%CI: 0.12-0.51), respectively. CONCLUSION The polymorphisms of CYP2C9, VKORC1 and CYP4F2 genes can influence the stable dose of warfarin in Shandong population. IWPC algorithm is suitable for guiding the use of warfarin in this population.
Anticoagulants ; administration & dosage ; Aryl Hydrocarbon Hydroxylases ; Cytochrome P-450 CYP2C9 ; genetics ; Cytochrome P450 Family 4 ; genetics ; Dose-Response Relationship, Drug ; Genotype ; Humans ; Models, Theoretical ; Polymorphism, Genetic ; Vitamin K Epoxide Reductases ; genetics ; Warfarin ; administration & dosage

ObjectiveTo investigate the effects of Yanghetang （YHT） on breast cancer 4T1 cells and their mitogen-activated protein kinase （MEK）/extracellular signal-regulated kinase （ERK） signaling pathway. MethodThe YHT-containing serum was prepared from SD rats. The rats were randomly assigned into a blank group （normal saline） and low-， medium-， and high-dose （5.8， 11.6， 23.2 g·kg^-1， respectively） YHT groups. The serum containing 10% YHT in each group was mixed with 90% RMPI 1640 complete medium， and the mixture was used to interfere with the cells. Cell counting kit-8 （CCK-8） method was used to detect the proliferation of the 4T1 cells treated with YHT for 24， 48， 72 h. The apoptosis， migration， and invasion of 4T1 cells were detected by flow cytometry， scratch test， and Transwell assay， respectively. Western blot was employed to determine the expression levels of MEK1/2， phosphorylation （p）-MEK1/2， ERK1/2， p-ERK1/2， and rat sarcoma virus （RAS） protein. ResultCompared with the blank group， the intervention with YHT-containing serum for 24， 48， and 72 h had significant inhibitory effect on 4T1 cell proliferation （P<0.05， P<0.01）. After intervention with YHT-containing serum for 48 h， the apoptosis rate of cells increased （P<0.01）. Compared with the blank group， the intervention with YHT for 24 h and 48 h decreased the healing ability of cells in the scratch test （P<0.01）. The invasive ability of cells treated with the low， medium， and high-dose YHT containing serum showed a decreasing trend （P<0.01）. Compared with the blank group， YHT-containing serum did not change the expression of MEK1/2 and ERK1/2 while down-regulating the expression of p-MEK1/2， p-ERK1/2， and RAS protein （P<0.01）. ConclusionYHT can inhibit the proliferation， migration， and invasion and promote the apoptosis of breast cancer 4T1 cells. In may promote the apoptosis by inhibiting the MEK/ERK signaling pathway and down-regulating the expression of p-MEK1/2， p-ERK1/2， and RAS protein.

Objective To investigate the pharmacological mechanism of Chaihudaxiong mixture in the treatment of coronavirus disease 2019 (COVID-19) based on a network pharmacology approach. Methods The effective ingredients and targets of Chaihudaxiong mixture were collected from Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP). The targets’ names were standardized by Uniprot database. Genes associated with coronavirus were obtained from the GeneCards and OMIM, which were intersected with effective therapeutic targets. A "herbs-ingredients-targets" network was compiled and analyzed by Cytoscape 3.7.2. The protein-protein interaction of the targets was analyzed by String. The GO gene annotation and KEGG signaling pathway analysis were performed using related packages of the R software. Results A total of 165 active ingredients and 51 targets were collected. Further analysis revealed that the main active ingredients were β-sitosterol and 11 flavonoids. The core targets were CASP3, MAPK3, IL-6, MAPK8, IL-10, CXCL8, MAPK1 and IL-1B. A total of 1722 GO entries were obtained from the GO gene annotation (P<0.05), including 1612 entries for biological processes, 30 entries for cell composition, and 80 entries for molecular functions. 156 signaling pathways (P<0.05) were obtained with KEGG signaling pathway screen. The important signaling pathways were AGE-RAGE signaling pathway in diabetic complication, Influenza A, IL-17 signaling pathway, TNF signaling pathway and hepatitis B. Conclusion This study revealed the synergistic features of multi-component, multi-target, and multi-pathway of Chaihudaxiong mixture in the treatment of COVID-19, which provided an important scientific basis for further understanding the mechanism of Chaihudaxiong mixture in the treatment of COVID-19.

Objective:To analyze the clinical characteristics and diagnosis and treatment experiences of Langerhans cell histocytosis (LCH) in skull.Methods:Sixteen patients with cranial LCH admitted to our hospital from January 2015 to December 2019 were chosen in our study. Their clinical data, diagnosis and treatment procedures and prognoses were retrospectively analyzed.Results:Among the 16 patients, there were 13 males and 3 females, aged from 1 to 31 years. The clinical manifestations included space-occupying lesions of the skull; and imaging showed bone destruction of the skull, with or without involvement of other bones or organs. All patients were pathologically confirmed to have LCH after surgical total resection of the lesions. Routine whole-body bone scanning was performed after surgery: one was found to have local abnormal metabolic activity and received local radiotherapy; 8 were combined with other bone or organ involvement, and received chemotherapy. All the patients were followed up for 1-5 years, and no recurrence was found, and no one died.Conclusion:Good prognosis can be achieved in cranial LCH patients accepted resection by giving additional treatment according to the results of postoperative reexamination and combination use of standardized radiotherapy and chemotherapy.

The PCR-amplified severe fever with thrombocytopenia syndrome virus(SFTSV)Gn-DⅢ-Ⅲ gene was inserted into the pET-30a(+)prokaryotic expression vector to generate the re-combinant plasmid pET-SFTSV-Gn-D Ⅲ-Ⅲ.The plasmid was transformed into E.coli BL21(DE3)for Gn-DⅢ-m protein expression and the expression conditions were optimized.The Gn-DⅢ-Ⅲ protein purified with Ni-NTA column affinity chromatography was applied as the captured antigen to establish an indirect ELISA method for the detection of SFTSV antibody.The results demonstrated that the recombinant plasmid pET-SFTSV-Gn-D Ⅲ-Ⅲ was successfully constructed as identified by PCR and sequencing.The recombinant protein SFTSV Gn-D m-Ⅲ was soluble ex-pression in E.coli under the optimal induction conditions of 0.4 mmol/L IPTG at 25 ℃ for 4 h,and the protein purity was 91.77％after purification by Ni-NTA column.The optimal reaction con-ditions for the indirect ELISA of SFTSV antibody were as follows:coating antigen concentration(5 μg/mL),primary antibody(incubation at 37 ℃ for 1.5 h),and secondary antibody(diluted 1:10 000 and incubated at 37 ℃ for 1 h).The established method had no cross-reactivity with Rift Valley fever virus(RVFV),Ebola virus(EBOV),and tick-borne encephalitis virus(TBEV)posi-tive sera.The method had a high sensitivity,with P/N＞2.1 for SFTSV-positive sera diluted to 81920.Coefficients of variation for intra-and inter-batch reactions were less than 10％.Detection of four SFTSV-infected human clinical serum samples showed the serum samples from patients in re-mission were tested as positive(P/N＞2.1),while serum samples from patients with multiple or-gan failure were detected as negative(P/N＜2.1).The results indicated that the SFTSV Gn-D Ⅲ-Ⅲ protein was successfully expressed and purified,and it was used as the coating protein to estab-lish an indirect ELISA assay for SFTSV antibody,which possesses good specificity,sensitivity and reproducibility.This method might be applied to detect human SFTSV clinical serum samples.

OBJECTIVE To analyze the clinical characteristics of nephrotic syndrome induced by sunitinib， and to provide reference for clinical rational drug use. METHODS Retrieved from CNKI， VIP， Wanfang data， PubMed， Web of Science and Medline， case report about sunitinib-induced nephrotic syndrome were collected from the inception to Oct. 30th， 2022. Those case reports were analyzed statistically in terms of gender， age， primary disease， drug use， clinical manifestations， treatment and outcome. RESULTS A total of 15 pieces of literature were collected and 17 patients were involved， including 10 males and 7 females. The average age of patients was （59.35±15.72） years. Among 17 patients， there were 10 patients with renal cell carcinoma and 7 patients with gastrointestinal stromal tumor， all of whom received evidence-based medication； the dosage of sunitinib in 15 cases was recorded， and all of them were within the recommended range of the instructions； 9 patients received combined therapy； the time from sunitinib application to the occurrence of nephrotic syndrome was 21 days-52 months， of which 11 cases were ≤2 years. The clinical manifestations in 13 patients were described， including edema， oliguria， foamy urine， weight gain， fatigue， dyspnea on exertion， etc. Eight patients had other adverse reactions induced by sunitinib before suffering from nephrotic syndrome， including new hypertension or worsening of original hypertension， and hand-foot syndrome. Renal biopsy mainly manifested as thrombotic microangiopathy， focal segmental glomerular sclerosis and immune complex glomerulonephritis. Sunitinib withdrawal or dosage reduction was adopted in all patients， and they were given symptomatic treatment such as glucocorticoids and antihypertensive agents. Symptoms of 16 patients were improved， and renal function of one patient deteriorated and hemodialysis was started. Sunitinib was re-challenged in 6 patients， elevated creatinine and substantial proteinuria recurred in 5 patients. CONCLUSIONS In clinical use of sunitinib， it is advisable to periodically monitor renal function. In case of deterioration of renal function， albuminuria， edema， etc.， relevant examinations should be implemented in time， and symptomatic intervention should be taken as soon as possible. Besides， we should be alert to the recurrence of nephrotic syndrome after sunitinib rechallenge.

Gene editing technology has been a hotspot in the field of biotechnology. CRISPR/Cas systems are efficient gene editing tools because of its specificity, simplicity and flexibility, these features enabled the rapid application of CRISPR/Cas systems in a variety of organisms. Moreover, the combination of transcriptional activator with dead Cas protein can achieve specific regulation of gene expression at the transcription level, which has made important contributions to the development of biotechnology in medical and agriculture. Overexpression of foreign genes is a common method to verify gene function and regulation. However, due to the limitation of vector capacity, it is difficult to achieve overexpression of multiple genes. CRISPR/Cas9 activation system can regulate the expression of multiple genes under the guidance of different guide RNAs to verify gene functions at the regulatory level. This review summarizes the composition of the CRISPR/Cas9 activation system and different activation strategies, and summarizes solutions for excessive activation. It may facilitate the application of CRISPR/Cas9 activation system in genetic improvement of cotton and herbicide resistance research.
Biotechnology ; CRISPR-Cas Systems/genetics* ; Gene Editing ; Phenotype ; RNA, Guide, Kinetoplastida/metabolism*