In this study, high-throughput promoter screening was employed to optimize the heterologous expression of Mesorhizobium loti carbonic anhydrase (MlCA) in order to reduce the costs associated with carbon capture and storage (CCS). To simplify the complexity of traditional vectors, a fusion protein expression system was constructed using superfolder green fluorescent protein (sfGFP) and MlCA. The synthetic promoter library in Escherichia coli was utilized for efficient one-step screening. Based on fluorescence intensity on agar plates, a total of 143 monoclonal colonies were identified, forming a library with varying expression levels. The top four recombinants with the highest fluorescence intensity were selected, among which MlCA driven by the promoter 342042/+ exhibited the highest enzymatic activity, with a specific activity of the 34.6 Wilbur-Anderson units (WAU)/mg. Optimization experiments revealed that MlCA exhibited the best performance when cultured for 4 days under pH 7.0 and 40 ℃ conditions. The Michaelis constant (K_m·hdy) and maximum reaction rate (V_max·hdy) for CO₂ hydration were determined to be 62.46 mmol/L and 0.164 mmol/(s·L), respectively. For esterase hydrolysis, MlCA showed the K_m and V_max of 639.8 mmol/L and 0.035 mmol/(s·L), respectively. MlCA accelerated the CO₂ hydration process, promoting CO₂ mineralized into CaCO₃ within 9 min at low pH and room temperature conditions. Scanning electron microscopy (SEM) and X-ray diffraction (XRD) analyses confirmed that the precipitated product was calcite. This study provides a low-cost and environmentally friendly alternative for future CCS applications.
Carbonic Anhydrases/biosynthesis* ; Promoter Regions, Genetic/genetics* ; Escherichia coli/metabolism* ; Carbon Sequestration ; Carbon Dioxide/metabolism* ; Green Fluorescent Proteins/metabolism*

In the management of obesity, aside from lifestyle interventions and surgery, pharmacotherapy is the most important choice. In recent years, research on the mechanisms of obesity and weight-loss drugs has been advancing rapidly. Exploring drugs that increase energy expenditure from the perspective of energy balance is beneficial for making clinical decisions based on the mechanisms of drugs and clinical needs in order to treat obesity effectively and improve the quality of life.

Objective:To identify a novel bombesin bioactive peptide from the skin secretion of Hylarana Latouchii, and to explore its effect on insulin secretion in islet cells.Methods:The skin secretion from Hylarana Latouchii was extracted by electrical stimulation, and the single chain of bombesin peptide was cloned and sequenced. The peptide QUB2995 was synthesized via solid-phase synthesis, then purified using reversed-phase high performance liquid chromatography (HPLC). Matrix assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF) was applied to validate. QPCR and ELISA were used to probe the effect of QUB2995 on insulin secretion in MIN6 and INS-1 cells.Results:A novel bombesin peptide named QUB2995 (GAFGDFLKGAAKA GALKILSIAQCKLSGTC) was found in the skin secretion of Hylarana Latouchii through molecular cloning. The bioactive peptide could significantly promote the proliferation and insulin secretion from mouse islet MIN6 cells and rat islet INS-1 cells. The effect reached a climax at the concentration of 10 -5 mol/L. Conclusion:A novel bombesin bioactive peptide named QUB2995 was found from Hylarana Latouchii. It could significantly promote insulin secretion in MIN6 cells of mouse islets and INS-1 cells of rat islets, indicating its potential in the treatment of diabetes.

Objective:To investigate the therapeutic value of modified multipoint drainage for biliary complications after liver transplantation.Methods:A total of 125 patients treated by endoscopic retrograde cholangiopancreatography (ERCP) for biliary complications after liver transplantation in Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine from May 2018 to May 2020 were enrolled. Patients were divided into endoscopic retrograde biliary drainage (ERBD) group ( n=67, treated with multiple drainage of bile duct stent) and modified multipoint drainage group [ n=58, treated with ERBD combined with endoscopic nasobiliary drainage (ENBD)] by random number table. Modified multipoint drainage group were further randomly divided into two groups, modification group 1, 31 cases, where nasobiliary ducts were cut proximal to duodenal papilla after one week under endoscopy and modification group 2, 27 cases, where they were cut proximal to duodenal papilla after two weeks under endoscopy. The changes of serological indexes in 2 weeks after the operation in three groups were compared, and the incidence of short-term and long-term complications were analyzed. Results:The serological indexes were improved in patients at 1 d, 7 d and 14 d after ERCP, especially in modified multipoint drainage groups. Two weeks after the operation, the improvement of serological indexes in modification group 2 was better than that in modification group 1. Incidence of recent complications including cholangitis, hyperamylasinemia, and pancreatitis in the ERBD group were higher than those in modification group 1 [32.84% (22/67) VS 12.90% (4/31), 46.27% (31/67) VS 19.35% (6/31), 20.90% (14/67) VS 3.23% (1/31), all P<0.05] and modification group 2 [32.84% (22/67) VS 11.11% (3/27), 46.27% (31/67) VS 22.22% (6/27), 20.90% (14/67) VS 3.70% (1/27), all P<0.05]. ERBD group had a higher incidence of long-term complications including recurrent biliary infection and jaundice than modification group 1 [ 58.21% (39/67) VS 35.48% (11/31), P=0.036; 49.25% (33/67) VS 25.81% (8/31), P=0.027] and modification group 2 [58.21% (39/67) VS 11.11% (3/27), P<0.001; 49.25% (33/67) VS 25.93% (7/27), P=0.038]. The incidence of recurrent biliary infection in modification group 1 was higher than that in modification group 2 [35.48% (11/31) VS 11.11% (3/27), P=0.030]. Conclusion:Multiple drainage with indwelling nasal bile duct by ERCP can effectively reduce the short-term and long-term complications and improve the recovery of serological indexes for patients with biliary complications after liver transplantation. It is suggested that the nasobiliary duct should be retained for 2 weeks and then transformed into a built-in tube to continue drainage.

Objective To investigate the effect of metformin on the growth of human anaplastic thyroid cancer cell HTh74Rdox which is doxorubicin resistant. Methods The HTh74Rdox was treated with different concentrations of metformin for 48 h. Cell morphology was observed by microscope, cell viability was tested by methylthiazoletetrazolium (MTT), cell apoptosis by annexin Ⅴ and propidium iodide double staining, the anti-oncogenic miRNA was assayed by realtime fluorescence quantitative PCR (RT-PCR), and the adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway tested by western blot. Furthermore, the anti-oncogenic miRNAs were knockdown by miRNA inhibitors (miR-34a, miR-101, miR-125b, and miR-138 inhibitors) and the cells were treated by metformin for 48 h, after that, cell apoptosis was detected by annexin Ⅴ and propidium iodide double staining, the expression of protein related to AMPK/mTOR signaling pathway was detected by western blot. Results Metformin inhibited the growth of human anaplastic thyroid cancer cell HTh74Rdox in a concentration-dependent manner, the cell apoptosis was induced by metformin, and there was a significantly lower expression of miR-34a, miR-101, miR-125b, and miR-138 in the HTh74Rdox. However, the four above miRNAs were upregulated by metformin, and AMPK/mTOR pathway was also activated by metformin. When these miRNAs were suppressed by miR-inhibitors (miR-34a, miR-101, miR-125b, miR-138 inhibitors), the stimulating effect of apoptosis and AMPK/mTOR pathway by metformin were reversed. Conclusion Metformin significantly suppresses cell viability of human anaplastic thyroid cancer cell HTh74Rdox, and stimulates AMPK/mTOR pathway and apoptosis by upregulating the expressions of miR-34a, miR-101, miR-125b, miR-138 in HTh74Rdox cell.

Objective:To study the causes that resulted in delayed surgery for bronchial ruptures and the results.Methods:The cases with the bronchial ruptures by the delayed surgery last decade were retrospectively reviewed.The causes and unsatisfactory results were analysed.Results:The severe complications usually occurred after the delayed surgery and the results were not as satisfactory as those by early surgery.Conclusion:The bronchial ruptures ought to be operated in the early stage after being wounded.