Objective:To investigate the impact of limonin(LIM)on the necrotic apoptosis of myocardial cells in rats with myocardial infarction(MI)by regulating the receptor-interacting protein 1(RIP1)/receptor-interacting protein 3(RIP3)/mixed-lineage kinase domain-like protein(MLKL)signaling pathway.Methods:A total of 60 Sprague-Dawley rats were randomly divided into sham-operation group(Sham group),MI model group(Model group),low-dose LIM group(LIM-L group,25 mg/kg LIM),high-dose LIM group(LIM-H group,50 mg/kg LIM),and high-dose LIM+RIP1 inhibitor Nec-1 group(LIM-H+Nec-1 group,50 mg/kg LIM+0.6 mg/kg Nec-1),with 12 rats in each group.A rat model of MI was established by ligation of the coronary artery.The changes of cardiac func-tion were examined for each group;HE staining was used to observe the pathological changes of myocardial tissue;the 2,3,5-triphen-yltetrazolium chloride-Evans blue method was used to measure myocardial infarct area;the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling method was used to observe the necrotic apoptosis of myocardial cells;quantitative real-time PCR and Western blot were used to measure the expression levels of mRNAs and proteins associated with the RIP1/RIP3/MLKL signaling pathway and the expression of apoptosis-related proteins.Results:Compared with the Sham group,the Model group had significant re-ductions in left ventricular ejection fraction,left ventricular systolic pressure,and the expression level of B-cell lymphoma-2(Bcl-2)in myocardial tissue(all P<0.001)and significant increases in left ventricular end-systolic volume,left ventricular end-diastolic pres-sure,left ventricular end-diastolic volume,myocardial infarct area,cell apoptosis rate,the mRNA and protein expression levels of RIP1,RIP3,and MLKL in myocardial tissue,and the protein expression levels of Bcl-2 associated X protein and cysteinyl aspartate-specific proteinase 3 in myocardial tissue(all P<0.001),as well as swelling and disordered arrangement of myocardial cells with ne-crosis and massive inflammatory cell infiltration on HE pathological sections.Compared with the Model group,the LIM-H group showed reverse changes in the above indicators(RIP1 mRNA:P=0.002,RIP3 mRNA:P=0.008,and the other indexes P were all<0.001),with alleviations of myocardial histopathological injury and inflammatory cell infiltration.Nec-1 promoted the effect of LIM in alleviating the necrotic apoptosis of myocardial cells in MI rats.Conclusion:LIM may alleviate the necrotic apoptosis of myocardial cells in MI rats by downregulating the RIP1/RIP3/MLKL signaling pathway.

Hepatic stellate cells (HSCs) represent a significant component of hepatocellular carcinoma (HCC) microenvironments which play a critical role in tumor progression and drug resistance. Tumor-on-a-chip technology has provided a powerful in vitro platform to investigate the crosstalk between activated HSCs and HCC cells by mimicking physiological architecture with precise spatiotemporal control. Here we developed a tri-cell culture microfluidic chip to evaluate the impact of HSCs on HCC progression. On-chip analysis revealed activated HSCs contributed to endothelial invasion, HCC drug resistance and natural killer (NK) cell exhaustion. Cytokine array and RNA sequencing analysis were combined to indicate the iron-binding protein LIPOCALIN-2 (LCN-2) as a key factor in remodeling tumor microenvironments in the HCC-on-a-chip. LCN-2 targeted therapy demonstrated robust anti-tumor effects both in vitro 3D biomimetic chip and in vivo mouse model, including angiogenesis inhibition, sorafenib sensitivity promotion and NK-cell cytotoxicity enhancement. Taken together, the microfluidic platform exhibited obvious advantages in mimicking functional characteristics of tumor microenvironments and developing targeted therapies.

A method for measuring 13C isotopic abundance of intracellular metabolites of Saccharopolysporaerythraea by ultra-high performance liquid chromatography (UPLC)-triple quadrupole mass spectrometry was established.First, the chromatographic conditions of UPLC were optimized, and then the MS conditions such as unique tube lens voltage, collision energy, and ion pair were optimized.On the bases of length of the parent and daughter ions carbon chains and whether the daughter ions contain 13C atoms, the one-to-one method, one-to-many method and SIM method were established for measuring 13C isotopic abundance.Then these methods were used to measure naturally labeled intracellular metabolite standards and 13C labeled samples, and according to the gap between the experimental value and the theoretical value, the best method was established for each metabolite of different characteristics.The results showed that one-to-one method was most effective for measuring the metabolites of daughter ions not containing 13C atoms represented by sugar phosphates, one-to-many method was the best for measuring the metabolites of both parent and daughter ions containing 13C short carbon chains represented by carboxylic acids, SIM method could play a role in measuring the metabolites of both parent and daughter ions containing 13C long carbon chains represented by coenzyme A.This method had a good measurement precision and could be applied to the measurement of Saccharopolysporaerythraea intracellular metabolites, which contributed to the consequent study of metabolic mechanism and the efficient expression of erythromycin.

Objective: To explore the clinical outcome of advanced testicular nonseminomatous germ cell cancer patients undergoing post chemotherapy retroperitoneal lymph node dissection (PC-RPLND), and to analyze the relevant prognostic factors of lymph node pathological. Methods: A total of 43 consecutive testicular nonseminomatous germ cell cancer patients underwent PC-RPLND between March 2001 and December 2014 in Department of Urology at Sun Yat-sen University Cancer Center were retrospectively reviewed. The average age of the patients was (29.0±11.5) years (ranging from 12 to 58 years). Before PC-RPLND, 22 patients were classified as phase Ⅱ, while 21 were phase Ⅲ. Primary tumor histology revealed seminomatous elements in 19 cases, embryonal cell carcinoma in 22 cases, yolk sac tumor in 13 cases, chorionic carcinoma in 3 cases, mature teratomatous elements in 11 and immature teratomatous elements in 2 cases. Patients were treated with cisplatin-based chemotherapy after orchectomy and then underwent surgical resection of retroperitoneal lymph nodes.After PC-RPLND, all patients underwent a periodic review including the blood routine, biochemistry routine and computed tomography or ultrasonograph of the chest, the abdomen and the pelvis. The association of pathological data with patient′s clinic features and the correlations between molecular features detected with each other were assessed by the t test, χ² and Fisher′s exact test. Multivariate logistic regression were used to assess prognostic factors. Results: The median operative time was 278 minutes (ranging from 50 to 715 minutes). Median blood loss was 425 ml (ranging from 50 to 5 000 ml). Eight patients received blood transfusion intra-operatively, 2 patients underwent adjunctive surgical procedures, 4 patients developed ileus and 4 had an ascites chylosus following PC-RPLND, 1 patient had a postoperative hyperthermia and retrograde ejaculation was present in 10 patients. The transverse diameter of the residual tumor in patients ranged from 0.8 to 18.2 cm. Necrosis, teratoma and viable germ cell tumors were found in 15, 17 and 11 of all patients. The median follow-up time was 46 months (ranging from 6 to 169 months). There were 39 patients had no tumor recurrence, 7 patients were found recurrence after PC-RPLND, 5 died of malignant germ cell tumor. The normal serum lactate dehydrogenase (LDH) level before chemotherapy (HR=25.811, 95%CI: 0.678 to 982.624, P=0.017) and relative changes more than 50% in retroperitoneal lymph node size (HR=0.016, 95%CI: 0 to 0.698, P=0.032) were statistically significant prognostic factors of the presence of necrosis. Conclusions Since most residual masses are not sensitive to chemotherapy, PC-RPLND is still an essential part of the treatment of metastatic testicular nonseminomatous germ cell cancer. Patients with the normal serum LDH level before chemotherapy and a shrinkage of 50% or more in retroperitoneal mass have a considerably chance of having necrosis in the retroperitoneum resection. This may help to refine the selection of candidates for PC-RPLND.