Healthy granulation tissue played an important role in the wound healing process. However, some factors which interfered the process would result in unhealthy granulation tissue. Unhealthy granulation tissue may affect wound repairing. This article would focus on the concept, mechanisms, interventions of unhealthy granulation tissue.

Objective To establish a flow-cytometric method to detect microvesicles (MVs) in rat peripheral blood, and to detect platelets-derived MVs (PMVs) and endothelial cell-derived MVs (EMVs) in blood from ischemic precondition-ing (IPC) treated rats. Methods Blood was withdrawn from rat abdominal aorta and anticoagulated with sodium citrate. Platelets-free plasma (PFP) was isolated through two centrifugations at room temperature. PFP was incubated with FITC-conjugated mouse anti-rat CD61 or PE-conjugated mouse anti-rat CD144. Standard beads in diameter of 1 and 2μm were used for calibration and absolute counting, respectively. Analysis was performed on flow cytometer. Results When 3.5%so-dium citrate was mixed with blood at volume ratio of 1∶4, clear supernatant was collected after centrifugation. Signals of parti-cles smaller than 1μm accounted for more than 99%of overall signals. PMVs and EMVs were CD61 positive and CD144 positive, respectively. Their diameters were both smaller than 1 μm. The concentration of PMVs and EMVs in peripheral blood from IPC treated rats was (4 053±1 987)/μL and (4 870±825)/μL, respectively. Conclusion The method for MVs de-tection by flow cytometry was successfully established and optimized, and verified through detecting PMVs and EMVs in pe-ripheral blood from IPC treated rats.

Objective To observe the effects of olprinone on ischemia/reperfusion (I/R) induced myocardial injury in male (Sprague-Dawley, SD rats) and explore its mechanisms. Methods Rats were subjected to a 30-min coronary arterial occlusion followed by 24-hour reperfusion. The survival rats were randomly divided into sham group (n=6), ischemia reperfusion group (I/R group, n=9), ischemia reperfusion+low dose of olprinone group(IR+olprinone-L group, n=6), ischemia reperfusion+medium dose of olprinone group (IR+olprinone-M group, n=6),ischemia reperfusion +high dose of olprinone group (IR+olprinone-H group, n=6). A MAP heart function analysis system was used to measure hemodynamic parameters; TTC staining method was used to detect the myocardial infarct size;24-hour mortality of SD rats was recorded; western blot was used to detect the levels of Caspase-3, Bax,Bcl-2, LC3B/LC3A,Beclin-1. Results Cardiac function in I/R group was lower than that in sham group, which was significantly improved by pretreatment with olprinone (P＜0.01),but systolic arterial pressure (SAP) diastolic arterial pressure (DAP) mean arterial pressure (MAP) mean pressure developed in left ventricle (Pmean) had no significant difference (P＞0.05). The percentage of myocardial infarct size in olprinone-M and olprinone-H group was lower than that in I/R group (P＜0.05).There was no significant difference in mortality among groups within 24 hours. Compared with sham group, the expressions of Caspase-3 and Bax were obviously up-regulated in I/R group (P＜0.01), whereas caspase-3 was down-regulated in olprinone-M group (P＜0.05) and Bax was inhibited by different doses of olprinone (P＜0.05), but the expression of Bcl-2 increased (P＜0.05); furthermore, the ratio of Bcl-2/Bax decreased in I/R group (P＜0.01) and increased with different degrees in different doses of olprinone (P＜0.05). Meanwhile, compared with sham group, the expression of Beclin-1 was up-regulated in I/R group(P＜0.05),and also increased in olprinone-L and olprinone-M groups(P＜0.05), but the ratio of Bcl-2 /Beclin-1 decreased in different doses of olprinone making statistically significant difference only in olprinone-M group (P＜0.05). Moreover, different doses of olprinone elevated the different ratios of LC3B/LC3A (P＜0.05), and this elevated ratio in olprinone-M group at median among groups. Conclusions Olprinone can strengthen the cardiac function after myocardial ischemia/reperfusion injury, without leading to disorders in hemodynamics; by regulating autophagy with anti-apoptotic protein, olprinone can make autophagy to an appropriate level using the mechanism of autophagy to preventing the myocardium from injury.

OBJECTIVETo investigate the anti-cancer effect of low-molecular-weight heparin (LMWH) combined with doxorubicin and explore the mechanism.

METHODSHepatocellular cancer HepG2 cells exposed to LMWH, doxorubicin, or both were evaluated for cell viability with MTT assay and for changes in their migration ability using wound healing assay and Transwell migration assay. The changes in cellular expressions of matrix metalloproteinase-9 (MMP-9) and MMP-2 mRNA and proteins were analyzed with quantitative real-time PCR (qRT-PCR) and Western blotting, and ELISA was used to determine heparanase (HPA) concentration in the cell culture medium.

RESULTSHepG2 cells exhibited suppressed proliferation in response to LMWH and doxorubicin treatments. The combined treatment caused a significantly higher inhibition rate of cell migration than LMWH and doxorubicin alone. LMWH enhanced doxorubicin-induced down-regulation of MMP-9, MMP-2 and HPA in the cells.

CONCLUSIONSLMWH can enhance the inhibitory effect of doxorubicin on the migration of HepG2 cells, the mechanism of which may involve the down-regulation of MMP-9, MMP-2 and HPA expressions.

Cell Movement ; drug effects ; Cell Survival ; Down-Regulation ; Doxorubicin ; pharmacology ; Glucuronidase ; chemistry ; Hep G2 Cells ; Heparin, Low-Molecular-Weight ; pharmacology ; Humans ; Liver Neoplasms ; pathology ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Neoplasm Invasiveness ; RNA, Messenger ; Real-Time Polymerase Chain Reaction

Objective:To assess the application of extracorporeal membrane oxygenation (ECMO) regional treatment pattern in patients with severe cardiopulmonary diseases.Methods:A retrospective analysis was conducted. Patients with severe cardiopulmonary disease who were transferred to Henan Provincial People's Hospital after ECMO treatment in cooperative hospitals were selected. The patients who received regular ECMO treatment from June 2017 to May 2018 were enrolled as the control group, and the patients who received ECMO regional treatment from June 2018 to May 2019 were selected as the observation group. The ECMO regional treatment pattern referred to implement a referral program for critical patients in primary hospitals, which mainly included the establishment of ECMO regional cooperative treatment network and ECMO referral team, the formulation of ECMO referral management standards, and the promotion of the merging of high-quality medical resources. Time of establishment of ECMO, ECMO regional treatment satisfaction, and the incidence of adverse events were also compared.Results:There were 27 patients enrolled in the control group and 64 patients in the observation group. There were no significant differences in gender, age, body mass index (BMI), ECMO mode, hypertension or coronary heart disease history between the two groups. Compared with the control group, the time for establishment of ECMO in the observation group was significantly shorter (minutes: 38.10±17.19 vs. 54.67±41.30, t = 2.715, P = 0.008), the ECMO treatment satisfaction of the observation group was also significantly higher than that of the control group (98.4% vs. 88.9%, χ 2 = 4.120, P = 0.042), and the incidence of ECMO referral adverse events was significantly lower than that of the control group (6.25% vs. 25.93%, χ 2 = 6.918, P = 0.009). Conclusion:The ECMO regional collaborative pattern in patients with severe cardiopulmonary diseases can shorten the time for establishment of ECMO, improve the satisfaction of ECMO treatment, and reduce the incidence of adverse events in ECMO referral.

Objective:To investigate whether silence information regulator 1 (SIRT1) could regulate nuclear factor E2-related factor 2/heme oxygenase 1 (Nrf2/HO-1) signaling pathway and its role in acute lung injury (ALI) in sepsis rats.Methods:Twenty-four male Sprague-Dawley (SD) rats were randomly divided into sham operation group (Sham group), cecal ligation and puncture (CLP) induced sepsis group (CLP group), sepsis+SIRT1 specific agonist group (CLP+SRT1720 group,10 mg/kg SRT1720 was intraperitoneally injected 2 hours before CLP), sepsis+SIRT1 specific inhibitor group (CLP+EX527 group, 10 mg/kg EX527 was intraperitoneally injected 2 hours before CLP), with 6 rats in each group. The rats were killed 24 hours after modeling and their lung tissues were taken for pathological score (Smith score), superoxide dismutase (SOD), glutathione (GSH), malondialdehyde (MDA), 8-hydroxydeoxyguanosine (8-OHdG), tumor necrosis factor-α (TNF-α), interleukins (IL-6, IL-1β), and SIRT1, Nrf2 and HO-1 mRNA and protein expression were detected.Results:The lung tissue of the CLP group mice was severely damaged, the alveolar interval was widened and a large number of inflammatory cells infiltrated, and there was visible pulmonary capillary hyperemia. The Smith score, the levels of TNF-α, IL-6, IL-1β, MDA and 8-OHdG were significantly increased, the levels of SOD, GSH, SIRT1, Nrf2 and HO-1 were significantly decreased in CLP group. After using SIRT1 specific agonist, the lung injury in CLP+SRT1720 group was significantly alleviated compared with that in CLP group, Smith score and lung tissue TNF-α, IL-6, and IL-1β levels were significantly decreased [Smith score: 2.83±0.75 vs. 5.67±0.52, TNF-α (ng/L): 36.78±5.36 vs. 66.99±5.44, IL-6 (ng/L): 23.97±3.76 vs. 45.70±4.16, IL-1β (ng/L): 16.76±1.39 vs. 39.64±2.59, all P < 0.05], SOD activity and GSH content increased [SOD (kU/g): 115.88±3.31 vs. 101.65±1.09, GSH (μmol/g): 8.42±0.81 vs. 5.74±0.46, both P < 0.05], MDA and 8-OHdG contents decreased [MDA (μmol/g): 5.24±0.33 vs. 9.86±0.66, 8-OHdG (ng/L): 405.76±8.54 vs. 647.12±10.64, both P < 0.05], the mRNA and protein expressions of SIRT1, Nrf2 and HO-1 were increased [SIRT1 mRNA (2 -ΔΔCT): 1.49±0.15 vs. 0.64±0.03, Nrf2 mRNA (2 -ΔΔCT): 1.19±0.08 vs. 0.84±0.02, HO-1 mRNA (2 -ΔΔCT): 1.80±0.41 vs. 0.64±0.11, SIRT1 protein (SIRT1/β-actin): 1.03±0.06 vs. 0.52±0.05, Nrf2 protein (Nrf2/β-actin): 1.14±0.10 vs. 0.63±0.05, HO-1 protein (HO-1/β-actin): 1.01±0.11 vs. 0.73±0.03, all P < 0.05]. The lung injury in CLP+EX527 group was more severe than that in CLP group, Smith score and lung tissue TNF-α, IL-6, IL-1β levels were significantly increased [Smith score: 8.00±0.89 vs. 5.67±0.52, TNF-α (ng/L): 87.15±4.23 vs. 66.99±5.44, IL-6 (ng/L): 66.79±2.93 vs. 45.70±4.16, IL-1β (ng/L): 58.99±2.12 vs. 39.64±2.59, all P < 0.05], SOD activity and GSH content decreased [SOD (kU/g): 72.84±3.85 vs. 101.65±1.09, GSH (μmol/g): 3.30±0.67 vs. 5.74±0.46, both P < 0.05], the contents of MDA and 8-OHdG were increased [MDA (μmol/g): 14.14±0.70 vs. 9.86±0.66, 8-OHdG (ng/L): 927.66±11.47 vs. 647.12±10.64, both P < 0.05], the mRNA and protein expressions of SIRT1, Nrf2 and HO-1 were decreased [SIRT1 mRNA (2 -ΔΔCT): 0.40±0.07 vs. 0.64±0.03, Nrf2 mRNA (2 -ΔΔCT): 0.48±0.07 vs. 0.84±0.02, HO-1 mRNA (2 -ΔΔCT): 0.27±0.14 vs. 0.64±0.11, SIRT1 protein (SIRT1/β-actin): 0.20±0.05 vs. 0.52±0.05, Nrf2 protein (Nrf2/β-actin): 0.45±0.01 vs. 0.63±0.05, HO-1 protein (HO-1/β-actin): 0.36±0.08 vs. 0.73±0.03, all P < 0.05]. Conclusions:In the rat model of ALI induced by sepsis, SIRT1 can regulate the activation of Nrf2/HO-1 signaling pathway, upregulate the expression of downstream antioxidant enzymes, reduce oxidative stress injury, and then alleviate the ALI induced by sepsis in rats.

Objective:To evaluate the effects of ketogenic diet(KD) on pancreatic β-cell dedifferentiation in db/db mice.Methods:In animal study, 8-week-old db/db male mice with type 2 diabetes mellitus(T2DM) were randomly divided into 3 groups: T2DM model group(ND), KD group, 75% caloric restriction(CR) group, and male C57BL/6 mice of the same age as normal control group(C) fed with standard diet. Both C and ND groups were on ad lititum feeding of chow, the KD group was free to eat the ketogenic diet, and the CR group was the positive control group, consuming 75% of the calories of the ND group every day. Four weeks after different diet intervention, body weight, fasting blood glucose, fasting insulin, glucose tolerance and blood β-hydroxybutyric acid(BHB) were measured. Morphology and structure of pancreatic islet was observed by hematoxylin-eosin staining(HE). Immunofluorescence co-staining was used to observe the expression of mouse pancreatic β-cell specific transcription factors.Results:After 4 weeks diet intervention, the fasting blood glucose, insulin and the area under the curve of blood glucose in KD group was significantly decreased( P<0.05); When compared with ND group, the morphology and structure of the islets in the KD group were more regular, and the number of islet cells increased as revealed with HE staining. Pancreatic immunofluorescence co-assay showed that KD not only restored the number and arrangement of β-cells and the ratio of β/α-cell in the pancreatic islets, but also reversed the expression of specific β-cell transcription factors such as pancreatic duodenal homeobox factor-1(PDX1). Conclusion:KD can reduce fasting blood glucose, fasting insulin and improve glucose tolerance in db/db mice, which may be related to its ability to restore the expression of specific β-cell transcription factors and reverse the dedifferentiation of pancreatic β-cells.

The 21st century is regarded as the century of biotechnological drugs, among which monoclonal antibodies and their derived targeting drugs have established themselves as the leading modality of biopharmaceutical pharmaceutics for a wide range of indications covering malignant tumors and autoimmune disorders. Since the manufacturing of the first antibody drug from hybridoma cells, the technologies have been intensely studied and there emerged numerous breakthroughs in recombinant cell line establishment, antibody expression and purification, quality control and other related areas. This article summarizes the critical progresses of antibody drugs expression technologies, especially of mammalian cell expression system, Escherichia coli expression system, the transgenic animal reactor and the cell free protein synthesis system, to give a detailed illustration of the recent advances in antibody drugs development.
Animals ; Antibodies, Monoclonal ; Biotechnology ; Cell Line ; Escherichia coli ; Hybridomas

Metabolic syndrome(MS)is a kind of metabolic disorder, including abdominal obesity, hyperglycemia, dyslipidemia, hypertension, etc. It reflects susceptibility to diabetes, cardiovascular disease, and other pathological conditions. In recent years, ketogenic diet(KD), as one of the natural therapies, has been proved to be effective in reducing weight, lowering blood glucose, regulating lipid metabolism, and improving insulin resistance. Therefore, its application in metabolic diseases has attracted more attention. There is growing evidence demonstrating directly or indirectly that KD may be an effective treatment for MS. How to safely and effectively implement KD and explore the biochemical mechanism of KD in the treatment of MS will be the focus of clinical research in the future.

Objective:To accurately and effectively identify the most critical needs of extracorporeal membrane oxygenation (ECMO) treatment for patients with severe cardiopulmonary diseases, and to better carry out continuous improvement of medical service quality an patients' satisfaction.Methods:Patients who underwent ECMO and transferred from 56 medical institutions in the Henan Provincial People's Hospital Critical Care Medicine Specialist Alliance [the patients who were transported before applying quality function deployment (QFD) from June 2017 to May 2018 were enrolled as the control group, and patients who were transported after applying QFD from June 2018 to May 2019 were the observation group], medical staff in the alliance hospitals, ECMO transfer teams and transfer driver teams were enrolled as the subjects of the survey. QFD was applied to convert the collected requirements into quality improvement elements for targeted improvement measures.Results:A total of 125 questionnaires were distributed in this survey, and 116 valid questionnaires were collected, including 91 from patients (including 27 from the control group and 64 from the observation group), 10 from the medical staff of the alliance hospitals, 10 from the ECMO transport teams and 5 from the transport driver teams. The questionnaire recovery rate was 92.8%. The improvement elements of ECMO treatment for patients with critical cardiopulmonary diseases were ranked according to the importance, and the top five were as follows: the accuracy of the first diagnosis, the specialization of ECMO team, the guarantee of vehicle safety, the seamless responses, and the smooth coordinated rescue protocol.Conclusion:The top five improvement elements should be prioritized in ECMO treatment of patients with critical cardiopulmonary disease in all hospitals of the Alliance to ensure more accurate and timely treatment.