In this study, 34 deep eutectic solvents (DESs) were successfully prepared for the extraction of proanthocyanidin from Rhodiolae Crenulatae Radix et Rhizomes. The extraction process was optimized using single factor exploration and Box-Behnken design-response surface analysis. The extraction rate was significantly improved when the molar ratio of choline chloride to 1,3-propanediol was 1:3.5 and the water content was 30% (V/V) in DESs. AB-8 macroporous resin and ethyl acetate were used for separation and refining, and the oligomer-rich proanthocyanidin components were eventually obtained. The ultraviolet (UV) and infrared (IR) spectra showed that the proanthocyanidins were mainly composed of catechin and epicatechin. To further clarify the chemical composition of proanthocyanidin, an ion scan list containing 156 proanthocyanidins precursors was obtained by constructing a proanthocyanidins structural library and mass defect filtering (MDF) algorithm, combined with the full mass spectrometry (MS)/dd-MS² scan mode that turns on the "if idle pick others" function. By using ultra-high performance liquid chromatography and high-resolution MS (UHPLC/HRMS), the analysis used both targeted and non-targeted methods to detect proanthocyanidins. Finally, 50 oligomeric proanthocyanidin (OPC) compounds were identified, including 7 monomers, 22 dimers, 20 trimers, and 1 tetramer, most of which were procyanidins of proanthocyanidins (84%), and a small amount of prodelphinidin (14%) and other types of proanthocyanidins (2%), which enabled the systematic characterization of proanthocyanidin components from Rhodiolae Crenulatae Radix et Rhizomes. Meanwhile, the comparison with the grape seeds OPCs standard (United States Pharmacopeia) revealed that the proanthocyanidins in Rhodiolae Crenulatae Radix et Rhizomes were more abundant, suggesting that the proanthocyanidins in Rhodiolae Crenulatae Radix et Rhizomes has promising applications.

For wild natural medicine, unanticipated biodiversity as species or varieties with similar morphological characteristics and sympatric distribution may co-exist in a single batch of medical materials, which affects the efficacy and safety of clinical medication. DNA barcoding as an effective species identification tool is limited by its low sample throughput nature. In this study, combining DNA mini-barcode, DNA metabarcoding and species delimitation method, a novel biological sources consistency evaluation strategy was proposed, and high level of interspecific and intraspecific variations were observed and validated among 5376 Amynthas samples from 19 sampling points regarded as "Guang Dilong" and 25 batches of proprietary Chinese medicines. Besides Amynthas aspergillum as the authentic source, 8 other Molecular Operational Taxonomic Units (MOTUs) were elucidated. Significantly, even the subgroups within A. aspergillum revealed here differ significantly on chemical compositions and biological activity. Fortunately, this biodiversity could be controlled when the collection was limited to designated areas, as proved by 2796 "decoction pieces" samples. This batch biological identification method should be introduced as a novel concept regarding natural medicine quality control, and to offer guidelines for in-situ conservation and breeding bases construction of wild natural medicine.

Long noncoding RNAs (lncRNAs) are expressed in different species and different tissues, and perform different functions, but little is known about their involvement in the synthesis or secretion of follicle-stimulating hormone (FSH). In general, we have revealed lncRNA‍‒‍microRNA (miRNA)‍‒‍‍messenger RNA (mRNA) interactions that may play important roles in rat primary pituitary cells. In this study, a new lncRNA was identified for the first time. First, we analyzed the gene expression of lncRNA-m18as1 in different tissues and different stages by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and observed the localization of lncRNA-m18as1 with fluorescence in situ hybridization, which indicated that this lncRNA was distributed mainly in the cytoplasm. Next, we used RT-qPCR and enzyme-linked immunosorbent assay (ELISA) to analyze the regulation of FSH synthesis and secretion after overexpression or knockdown of lncRNA-m18as1 and found that lncRNA-m18as1 was positively correlated with FSH synthesis and secretion. In addition, mothers against decapentaplegic homolog 2 (Smad2) was highly expressed in our sequencing results. We also screened miR-18a-5p from our sequencing results as a miRNA that may bind to lncRNA-m18as1 and Smad2. We used RNA immunoprecipitation-qPCR (RIP-qPCR) and/or dual luciferase assays to confirm that lncRNA-m18as1 interacted with miR-18a-5p and miR-18a-5p interacted with Smad2. Fluorescence in situ hybridization (FISH) showed that lncRNA-m18as1 and miR-18a-5p were localized mainly in the cytoplasm. Finally, we determined the relationship among lncRNA-m18as1, miR-18a-5p, and the Smad2/3 pathway. Overall, we found that lncRNA-m18as1 acts as a molecular sponge of miR-18a-5p to regulate the synthesis and secretion of FSH through the Smad2/3 pathway.
Animals ; Cell Line, Tumor ; Cell Proliferation ; Follicle Stimulating Hormone/metabolism* ; Gene Expression Regulation, Neoplastic ; In Situ Hybridization, Fluorescence ; MicroRNAs/metabolism* ; RNA, Long Noncoding/metabolism* ; Rats

Objective:To explore the microbiological characteristics of donor blood culture and donor liver perfusion culture and summarize the clinical experiences to provide basic rationales for preventing donor-derived infections.Methods:From August 1, 2018 to November 26, 2018 and November 27, 2018 to December 31, 2020 at First Affiliated Hospital, Zhengzhou University, culture results of donor blood and donor liver perfusate were retrospectively reviewed.According to whether or not donor liver was obtained without breaking diaphragm, removing gallbladder intraoperatively and flushing bile through cystic duct, two stages were assigned: before and after improvement measures of liver donor, i.e.August 1, 2018 to November 26, 2018 and November 27, 2018 to December 31, 2020.The culture results of donor blood samples and donor liver perfusion fluid samples in two stages of liver transplantation were statistically analyzed and infection preventing measures during donor liver maintenance and obtaining donor liver examined.Results:A total of 486 cases of blood culture from potential donors and 478 cases of liver perfusion culture were analyzed.The results showed that the incidence of blood culture infection was 4.5% and 4.3% before and after improvement measures( χ2=0.008; P=0.927)while the incidence of perfusion fluid infection was 56.8% and 46.2%( χ2=4.569; P=0.031); Klebsiella pneumoniae was a major pathogen cultured in perfusion solution before improvement measures and Staphylococcus epidermidis after improvement measures. Conclusions:Before organ donation, infection screening and prevention of potential donors and corresponding measures during donor liver acquisition can reduce donor source infection and effectively lower the mortality of recipients.

Objective:To evaluate the effects of different doses of ulinastatin on lung function in patients undergoing total aortic arch replacement.Methods:One hundred and thirty five patients with acute Stanford type A aortic dissection of both sexes, aged 20-70 yr, with body mass index of 16.2-33.3 kg/m 2, of American Society of Anesthesiologist physical status Ⅳ, were divided into 3 groups ( n=45 each) using a random number table method: high-dose ulinastatin group (group H with total dose of 30 000 U/kg), low-dose ulinastatin group (group L with total dose of 20 000 U/kg) and control group (group C). In group H and group L, half of the total dose of ulinastatin was given after induction of anesthesia, the rest of the total dose was primed after being added to cardiopulmonary bypass (CPB) circuit, while normal saline 100 ml was given at the same time point in group C. After induction of anesthesia (T 0), and at 3, 6, 12, 24 and 48 h after the beginning of CPB (T 1-5), blood samples from the central vein were collected for determination of plasma concentrations of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6). The oxygenation index (OI) and alveolar-arterial partial pressure of oxygen difference (P A-aO 2) at T 0 and T 2-T 5, intraoperative blood loss and blood transfusion, postoperative mechanical ventilation time, length of intensive care unit (ICU) stay and the incidence of complications within 30 days after surgery were collected. Results:Compared with group C, the plasma concentrations of TNF-α and IL-6 were significantly at T 1-T 5, OI was increased, and P A-aO 2 was decreased at T 2, 3 in H and L groups ( P<0.05). There was no significant difference in the mechanical ventilation time, length of ICU stay and incidence of complications within 30 days after surgery among the 3 groups ( P>0.05). Conclusion:Ulinastatin can inhibit inflammatory responses and improve lung function in patients undergoing total aortic arch replacement, but it has no value for clinical outcomes.

Objective To evaluate the effect of autologous platelet-rich plasma(aPRP)on postop-erative outcomes in the patients undergoing Sun's procedure.Methods One hundred and sixteen patients with acute Stanford type A aortic dissection of both sexes,aged 18-64 yr,with body mass index of 19-34 kg/m2,of American Society of Anesthesiologist physical status Ⅳ,were divided into aPRP group(group A)and control group(group C)by a random number table method,with 58 cases in each group.After an-esthesia induction and intubation,aPRP preparation was completed before surgery in group A,and surgery began in group C.After heparin neutralization,group A received retransfusion of platelet-rich plasma,while group C received conventional fluid management.Thromboelastography was performed on all patients,and the results were recorded.The intraoperative amount of allogeneic blood products and blood loss were recorded in two groups.Plasma concentrations of tumor necrosis factor-alpha(TNF-α)and interleukin-6(IL-6)were recorded after anesthesia induction(T0),immediately after completion of aPRP preparation(T1),3 h after heparin neutralization(T2),and at 24 and 48 h after surgery(T3,4).The volume of post-operative chest drainage and amount of allogeneic blood products,mechanical ventilation time,lengths of intensive care unit and hospital stay,incidence of important complications and fatality rate within 30 days after surgery were also collected.Results Compared with group C,the intraoperative amount of allogeneic blood products and blood loss were significantly reduced,α angle and MA values of thromboelastograph were larger,plasma concentrations of TNF-α and IL-6 were reduced at T2,3,postoperative chest drainage volume and allogeneic blood transfusion were reduced,the postoperative mechanical ventilation time and length of hospital stay were shortened,and the incidence of pulmonary complications was reduced in group A(P＜0.05).There was no significant difference in the length of intensive care unit stay,incidence of other complications or fatality rate within 30 days after surgery between the two groups(P>0.05).Conclu-sion aPRP can improve postoperative outcomes in the patients undergoing Sun's procedure.

Objective: To evaluate the effect of autologous platelet-rich plasma (aPRP) on postoperative outcomes in the patients undergoing Sun′s procedure. Methods: One hundred and sixteen patients with acute Stanford type A aortic dissection of both sexes, aged 18-64 yr, with body mass index of 19-34 kg/m², of American Society of Anesthesiologist physical status Ⅳ, were divided into aPRP group (group A) and control group (group C) by a random number table method, with 58 cases in each group.After anesthesia induction and intubation, aPRP preparation was completed before surgery in group A, and surgery began in group C. After heparin neutralization, group A received retransfusion of platelet-rich plasma, while group C received conventional fluid management.Thromboelastography was performed on all patients, and the results were recorded.The intraoperative amount of allogeneic blood products and blood loss were recorded in two groups.Plasma concentrations of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) were recorded after anesthesia induction (T₀), immediately after completion of aPRP preparation (T₁), 3 h after heparin neutralization (T₂), and at 24 and 48 h after surgery (T_{3, 4}). The volume of postoperative chest drainage and amount of allogeneic blood products, mechanical ventilation time, lengths of intensive care unit and hospital stay, incidence of important complications and fatality rate within 30 days after surgery were also collected. Results: Compared with group C, the intraoperative amount of allogeneic blood products and blood loss were significantly reduced, α angle and MA values of thromboelastograph were larger, plasma concentrations of TNF-α and IL-6 were reduced at T_{2, 3}, postoperative chest drainage volume and allogeneic blood transfusion were reduced, the postoperative mechanical ventilation time and length of hospital stay were shortened, and the incidence of pulmonary complications was reduced in group A(P<0.05). There was no significant difference in the length of intensive care unit stay, incidence of other complications or fatality rate within 30 days after surgery between the two groups (P>0.05). Conclusion aPRP can improve postoperative outcomes in the patients undergoing Sun′s procedure.

Staphylococcal nuclease A (SNA) may be used to produce bacterial ghosts for further inactivation of host bacteria and elimination of residual genetic materials. It is still controversial if SNA without signal peptide can be secreted to extracellular matrix and if fusion with other peptide is required for its function in the cytoplasm of host bacteria. To clarify this dispute, a series of temperature-inducible plasmids carrying SNA alone or SNA fused with partial sequences of λ phage cro gene (cSNA) or Mycobacterium tuberculosis urease gene (uSNA) were constructed and evaluated in Escherichia coli. Results show that the percentages of inactivated E. coli by SNA, cSNA and uSNA after 4 h of induction were 99.9%, 99.8% and 74.2%, respectively. Moreover, SNA and cSNA in the cytoplasm of host bacteria were initially detectable after 30 min of induction, whereas uSNA was after 1 h. In comparison, SNA and cSNA in culture supernatant were initially detectable 1 h later, whereas uSNA was 2 h later. The nuclease activity in the cytoplasm or supernatant was ranked as follows: SNA > cSNA > uSNA, and the activity in the supernatant was significantly lower than that in the cytoplasm. Furthermore, host genomic DNA was degraded by SNA or cSNA after 2 h of induction but not by uSNA even throughout the whole experiment. In conclusion, this study indicates that SNA, cSNA and uSNA expressed in host bacteria all have nuclease activity, the enzymes can be released to culture media, and fusion with exogenous peptide negatively reduces the nuclease activity of SNA.
Bacteriolysis ; Bacteriophage lambda ; DNA ; chemistry ; Escherichia coli ; Genetic Vectors ; Micrococcal Nuclease ; chemistry ; Plasmids ; Protein Sorting Signals

Objective To investigate the therapeutic effect of three-dimensional interactive reduction of femoral neck and shaft in treatment of irreducible femoral neck fracture.Methods The study enrolled 40 patients with irreducible femoral neck fractures treated by three-dimensional interactive reduction of femoral neck/shaft and internal fixation with three cannulated screws from June 2011 to July 2013 (study group).Frontal and lateral X-ray films were taken after operation.Garden index was used to evaluate the fracture reduction quality and Harris hip score was recorded.Meanwhile,a retrospective analysis was performed on 32 patients with irreducible femoral neck fractures treated by open reduction and internal fixation with cannulated screws between January 2008 and December 2010 (control group).Results Patients in study group obtained satisfactory reduction by minimally traumatic closed reduction.According to the Garden index,fracture reduction quality was level 1 in 24 patients and level 2 in 16 patients.Thirty-two patients were followed up for 12-28 months (mean,21 months),which showed fracture healing.At the final follow-up,Harris hip score was 87 points (range,61-100 points) and 4 patients had femoral head avascular necrosis.By contrast,6 patients in control group showed fracture nonunion.At the final follow-up,Harris hip score was 60 points (range,20-100 points) and 8 patients had femoral head avascular necrosis.Conclusion Three-dimensional interactive reduction of femoral neck and shaft provides good hip function recovery and decreased incidence of femoral head avascular necrosis and fracture disunion compared with the open reduction and internal fixation.

Objective To approach the effective treatment for simple bone cyst in adult. Methods Forty-eight cases were divided into two groups by visiting order, 20 cases (steroid group) were treated by prednisolone acetate and 28 cases (open resection and allograft group) were treated by open resection and allograft. The curative effect were contrasted between two groups. Results The follow-up time was 5-46(26.58 ± 10.81) months. Aecording to the Chigira's healing criteria for simple bone cyst, grade Ⅰ , Ⅱ , Ⅲ,V were 1, 2, 10 and 7 cases respectively in steroid group, contrasting to 0, 3, 5 and 20 cases respectively in open resection and allograft group. The recovery rate was 85.0%(17/20) in steroid group and 89.3%(25/28) in open resection and allograft group, which was no statistically significant difference between two groups (P＞0.05). Conclusions Although the two remedies are no obvious difference in curative effect,simple bone cyst at lower extremity of weight-bearing with obvious osteolysis in adult is recommended to open resection and allograft, otherwise or at upper extremity to steroid injection.