Metabolic engineering provide powerful tools for the systematic manipulation of cellular metabolic activities. The ptsG gene for glucose-specific transporter Enzyme II CBGlc of the phosphotransferase system was knock-out so as to reduce the accumulation of acetic acid in the high cell-density culture of Escherichia coli on excess glucose. The chloramphenicol-resistant cassette with short shared sequences on both ends generated by PCR was electroporated into Escherichia coli DH5alpha and JM109. Recombination between linear DNA cassettes and Escherichia coli chromosomes took place by Red recombinase functions. Therefore, the ptsG gene was disrupted to construct the mutants called DH5alphaP and JM109P. There was no difference between the mutants and parent strains in LB media.However, in LB media supplemented with glucose, the mutants of Escherichia coli deficient in ptsG showed greater biomass, together with exploiting more glucose. The maximal cell density obtained with DH5alphaP was approximately 3 times more than that of DH5alpha, then the result of JM109P increased fourfold. The products of recombinant protein TNF respectively accounted for 24.3% of total cellular protein in DH5alphaP with A600 8.28 and 20.8% of total cellular protein in JM109P with A600 7.62. The specific volume expression amount of TNF was greater in the ptsG mutant than in its parent strain. These results demonstrate that the ptsG-mutant strains will be available for high cell-density culture.
Culture Media ; Escherichia coli ; genetics ; growth & development ; Fermentation ; Mutation ; Phosphoenolpyruvate Sugar Phosphotransferase System ; genetics ; Polymerase Chain Reaction ; Recombinant Proteins ; biosynthesis ; Tumor Necrosis Factor-alpha ; biosynthesis

Cardiovascular diseases are characterized by cardiac and vascular dysfunction. Prokineticin 2 ( PK2 ) is a newly found secretory peptide which plays a key role in the physiology homeostasis via prokineticin receptor 1 and 2 ( PKR1 and 2). Furthermore, PK2/PKR1 signaling pathway plays an important role in protecting cardiovascular diseases. Here we discuss the effect of PK2/PKR1 signaling in myocardial infarction, conges-tive heart failure and vascular endothelial dysfunction.

BACKGROUNDProteasome subunits (PSMB) and transporter associated with antigen processing (TAP) loci are located in the human leukocyte antigen (HLA) Class II region play important roles in immune response and protein degradation in neurodegenerative diseases. This study aimed to explore the association between single nucleotide polymorphisms (SNPs) of PSMB and TAP and Parkinson's disease (PD).

METHODSA case-control study was conducted by genotyping SNPs in PSMB8, PSMB9, TAP1, and TAP2 genes in the Chinese population. Subjects included 542 sporadic patients with PD and 674 healthy controls. Nine identified SNPs in PSMB8, PSMB9, TAP1, and TAP2 were genotyped through SNaPshot testing.

RESULTSThe stratified analysis of rs17587 was specially performed on gender. Data revealed that female patients carry a higher frequency of rs17587-G/G versus (A/A + G/A) compared with controls. But there was no significant difference with respect to the genotypic frequencies of the SNPs in PSMB8, TAP1, and TAP2 loci in PD patients.

CONCLUSIONChinese females carrying the rs17587-G/G genotype in PSMB9 may increase a higher risk for PD, but no linkage was found between other SNPs in HLA Class II region and PD.

ATP-Binding Cassette Sub-Family B Member 2 ; genetics ; ATP-Binding Cassette, Sub-Family B, Member 3 ; genetics ; Adult ; Aged ; Antigen Presentation ; Case-Control Studies ; Cysteine Endopeptidases ; genetics ; Female ; Humans ; Male ; Middle Aged ; Parkinson Disease ; genetics ; immunology ; Polymorphism, Single Nucleotide ; Proteasome Endopeptidase Complex ; genetics

OBJECTIVETo observe the effect of Shenluotong Decoction (SD) on serum levels of aldosterone, monocyte chemoattractant protein-1 (MCP-1), α-smooth muscle protein (α-SMA), and nuclear factor-KB (NF-κB) in obstructive nephropathy rats, and to explore the initial mechanism of SD for inhibiting renal interstitial fibrosis.

METHODSTotally 48 healthy Wistar rats were randomly divided into the sham-operation group (n =12) and the model group (n =36). Renal interstitial fibrosis rat model was established by unilateral ureteral obstruction (UUO). After successful modeling, 36 rats were randomly divided into the model group, the Chinese medicine group, and the Western medicine group, 12 in each group. Eplerenone was added in the forage at the daily dose of 100 mg/kg for rats in the Western medicine group. Chinese medicine was added in the forage at the daily dose of 26 g/kg for rats in the Chinese medicine group. Equal volume of normal saline was administered to rats in the sham-operation group and the model group. All medication was performed once daily. The obstructive kidneys were extracted ten days after medication. The pathomorphological changes were observed. The contents of serum aldosterone and MCP-1, and the protein or mRNA expression of MCP-1, α-SMA, and NF-KB were detected.

RESULTSCompared with the sham-operation group, infiltration of a large amount of inflammatory cells and collagen deposition significantly increased, serum contents of aldosterone and MCP-1 obviously increased (P < 0.01), the expression of MCP-1 mRNA and protein were significantly up-regulated (P <0.01), the protein expression of α-SMA and NF-KB were significantly enhanced in the model group (P <0.01). Com- pared with the model group, infiltration of inflammatory cells and renal collagen deposition were attenua- ted in the Chinese medicine group and the Western medicine group, the serum MCP-1 level were reduced, and the mRNA and protein expression of MCP-1 were significantly down-regulated (P <0.01), the protein expression of α-SMA and NF-KB were obviously inhibited (P <0. 01). At the same time, serum aldosterone level was reduced in the Chinese medicine group (P <0.01).

CONCLUSIONSinflammatory lesions of the renal tissue could promote the progress of interstitial fibrosis in rats with obstructive nephropathy. SD could attenuate interstitial fibrosis through reducing serum contents of aldosterone and MCP-1, down-regulating MCP-1/ NF-KB, and inhibiting the expression of α-SMA.

Animals ; Chemokine CCL2 ; drug effects ; metabolism ; Down-Regulation ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Fibrosis ; Kidney ; drug effects ; Kidney Diseases ; drug therapy ; genetics ; NF-kappa B ; drug effects ; metabolism ; RNA, Messenger ; biosynthesis ; Rats, Sprague-Dawley ; Ureteral Obstruction ; drug therapy ; genetics

Objective:To explore a better indicator that can predict septic shock induced acute kidney injury (AKI) by combining renal resistive index (RRI) and central venous pressure (CVP).Methods:A prospective observational study was conducted. Patients with septic shock admitted to department of critical care medicine of Hebei General Hospital from November 2017 to October 2018 were enrolled. Baseline characteristics such as age, gender, underlying diseases, infection sites, acute physiology and chronic health evaluation Ⅱ(APACHEⅡ) in the first 24-hour, sequential organ failure assessment (SOFA) were recorded; Doppler-based RRI was obtained on the first day when hemodynamics was relatively stable, meanwhile the dose of norepinephrine and hemodynamic parameters were assessed. Urine output per hour, the total duration of mechanical ventilation, the length of intensive care unit (ICU) stay and 28-day mortality were also collected. Observational end point was death at discharge or the 28th day after ICU admission, whenever which came first. The patients were divided into AKI and non-AKI groups according to the 2012 Kidney Disease: Improving Global Organization (KDIGO) clinical practice guideline. The baseline and prognostic indicators, variables potentially associated with AKI were compared between the two groups. The variables independently associated with septic shock induced AKI were identified using multivariable Logistic regression. The predictive value of RRI and RRI combining CVP for AKI were analyzed by the receiver operating characteristic (ROC) curve.Results:A total of 107 patients were enrolled, with 59 patients in AKI group and 48 patients in non-AKI group. There was significant difference in RRI, CVP, percentage of norepinephrine dosage ≥0.5 μg·kg -1·min -1, procalcitonin (PCT), lactate (Lac), and serum creatinine (SCr) between the two groups. Logistic regression analysis showed that high CVP, RRI, Lac and PCT were independent risk factors for septic shock induced AKI [CVP: odds ratio ( OR) = 1.20, 95% confidence interval (95% CI) was 1.03-1.40, P = 0.022; RRI: OR = 3.02, 95% CI was 2.64-3.48, P = 0.006; Lac: OR = 2.43, 95% CI was 1.32-4.50, P = 0.005; PCT: OR = 1.20, 95% CI was 1.05-1.38, P = 0.009]. ROC curve analysis showed that the area under ROC curve (AUC) values of CVP≥9.5 mmHg (1 mmHg = 0.133 kPa) and RRI≥0.695 for predicting septic shock induced AKI were 0.656 and 0.662 respectively. The AUC value of the combination of RRI and CVP was greater compared with either RRI or CVP alone in predicting septic shock induced AKI, which AUC value was 0.712, 95% CI was 0.615-0.809, the sensitivity was 59% and the specificity was 75%. Conclusions:High CVP and RRI were independent risk factors for septic shock induced AKI. The combination of RRI and CVP performs poorly in predicting septic shock induced AKI. Further studies are needed to describe factors influencing Doppler-based assessment of RRI, which may help clinicians to prevent AKI early.

OBJECTIVETo transfect a short hairpin RNA (shRNA) against survivin gene into human T lymphoblastic leukemia cell line Jurkat, and to explore the effects on apoptosis and proliferation of transfected cells.

METHODSThe survivin-shRNA expression vector were constructed and transfected into Jurkat cells. Expression of survivin mRNA and protein were assessed by RT-PCR and Western blot analysis respectively. Apoptosis index of transfected Jurkat cells was quantified by flow cytometry. The potential of cell proliferation was described by cell growth curves.

RESULTSIn survivin-shRNA transfected Jurkat cells, survivin mRNA levels were significantly reduced by 66.67% ( transient transfection) and 60.69% ( stable transfection) respectively, compared with that in control-shRNA treated group and PBS treated group (P < 0.05); and the levels of survivin protein were significantly reduced by 63.41% (transient transfection) and 60.18% (stable transfection), compared with that in the two control groups (P < 0.05). Apoptosis index was significantly increased during both transient and stable transfection, respectively [(22. 41 +/- 2.83)% and (20.73 +/- 2.56)% (P < 0.05)]. Survivin-shRNA also inhibited the proliferation of Jurkat cells.

CONCLUSIONSVector-based survivin-shRNA can effectively reduce the expression of survivin gene, induce apoptosis

Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; Gene Expression ; Gene Silencing ; Humans ; Inhibitor of Apoptosis Proteins ; Jurkat Cells ; Microtubule-Associated Proteins ; biosynthesis ; genetics ; Neoplasm Proteins ; biosynthesis ; genetics ; RNA Interference ; RNA, Messenger ; biosynthesis ; RNA, Small Interfering ; pharmacology

OBJECTIVETo summarize the Chinese experience in pathologic diagnosis of liver biopsies after orthotopic liver transplantation (OLTx).

METHODS1123 post-transplant liver biopsies from 665 OLTx patients from the Shanghai Eastern Hepatobiliary Surgery Hospital, Tianjin First Central Hospital, Guangzhou Sun Yat-sen University and Chongqing Southwest Hospital were retrospectively analyzed. All liver biopsies were stained with hematoxylin and eosin. Immunohistochemical studies for cytomegalovirus, HBsAg, CK19, CD4 and CD8 were also performed in selected examples.

RESULTSIn the involved hospitals, 4 to 12 types of complications were encountered after OLTx. The number of liver biopsies performed for each patient ranged from 1 to 9 (mean = 2.2). The timing of these biopsies varied from the second to the 2877 th post-transplant day. The 5 most common complications were acute cellular rejection (35.6%), ischemic-reperfusion injury (13.4%), biliary stricture (5.6%), drug complication (5.0%) and chronic rejection (4.7%). The 5 earliest complications after OLTx were primary non-function (occurring at day 4.7 +/- 2.1), ischemic-reperfusion injury (occurring at day 14.0 +/- 4.0), acute cellular rejection (occurring at day 32.1 +/- 62.9), hepatic artery thrombosis / stricture (occurring at day 62.9 +/- 74.2) and cytomegalovirus infection (occurring at day 107.7 +/- 93.0).

CONCLUSIONSThis study has evaluated the types, incidence and timing of major complications occurring after OLTx. The most important issue is the distinction between rejection and non-rejection pathology. Thorough understanding of atypical pathologic features of these complications is necessary. The Banff Schema (rejection activity index) for grading liver allograft rejection is useful for monitoring anti-rejection therapy and should be used routinely.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Biopsy, Needle ; Child ; Child, Preschool ; Cholestasis, Intrahepatic ; pathology ; Female ; Graft Rejection ; pathology ; Hepatic Artery ; pathology ; Humans ; Infant ; Liver Transplantation ; pathology ; Male ; Middle Aged ; Postoperative Complications ; pathology ; Reperfusion Injury ; pathology ; Retrospective Studies ; Thrombosis ; pathology

OBJECTIVETo observe the effect on intervention of sub-health with pestle needle (Chuzhen).

METHODRandomized controlled trail was adopted for this research. One hundred and fifty-three cases were randomly divided into two groups of a Chuzhen group (79 cases) and a massage group (74 cases). Acupoint of Bazhen (Baihui Bazhen, Shendao Bazhen, Zhiyang Bazhen, Mingmen Bazhen, Yaoyangguan Bazhen), Hechelu on the head, the neck and the lumbar area were adopted in Chuzhen group. While regular whole-body massage was applied in the massage group. The human sub-health score, the cornell medical index (CMI) and thermal texture maps system (TTM) technology of the two groups before and after the intervention were observed.

RESULTS1) After treatment, sub-health condition score, the CMI score, the M-R score and the TTM index were all increased in both groups (all P<0.01) 2) Comparison of D-value of the two groups before and after the intervention: the level of the sub-health score, the total score of CMI, and the index of sleep, pressure, Governor Vessel, Hukou (first web), blood lipid, viscosity of blood, microcirculation of TTM index of the Chuzhen group changed more obvious (all P<0.01), but there was no statistic significances in the M-R score and blood sugar of the TTM (both P>0.05). 3) The sub-health condition score in Chuzhen group was higher than that in the massage group (P<0.01).

CONCLUSIONChuzhen therapy has definite effect on intervention of sub-health, which is better than regular general massage.

Acupuncture Points ; Acupuncture Therapy ; Adolescent ; Adult ; Aged ; Cornell Medical Index ; Female ; Humans ; Male ; Massage ; Middle Aged ; Young Adult

BACKGROUNDSevoflurane is currently used as a volatile inhalation anesthetic with many clinical advantages. A representative degradation product, compound A, was quantitatively measured to investigate whether there are different reactions between two kinds of water content sevoflurane formulations with different carbon dioxide (CO2) absorbents.

METHODSA closed-circle breathe bag with the Dräger Fabius GS anesthesia apparatus was used as an artificial rubber lung. The experiments were grouped according to different sevoflurane formulations: group A: higher-water sevoflurane (Ultane); group B: lower-water sevoflurane (Sevoness). During the experiment, CO2 (200 ml/min) was continually perfused to keep the end-tidal pressure of CO2 (P(ET)CO2) at 35 - 45 mmHg. The artificial ventilation was set to 6 L/min, and the breathing rate at 12 breaths/min. The circuit was operated with constant fresh gas flow rate (1 L/min) and the sevoflurane concentration was kept at 1.0 minimum alveolar concentration (MAC) for 240 minutes. At 0, 10, 20, 30, 60, 90, 120, 180 and 240 minutes, gas was collected from the Y-piece. Gas chromatography/mass spectrometry (GC/MS) was used to quantify the major degradation product, compound A, with different water content sevoflurane. PETCO2 and sevoflurane concentration, and the temperature of the canister were continuously monitored during the experiment.

RESULTSThere were no significant differences in P(ET)CO2 and sevoflurane concentrations between the two groups. Drägersorb 800 plus produced the highest concentrations of compound A compared with other sodalimes, and Sevoness in Drägersorb 800 plus generated more compound A than Ultane (P < 0.05). There were significant differences in the peak and average compound A concentrations between Ultane and Sevoness with Drägersorb 800 plus (P < 0.05), while the compound A concentration produced by Sodasorb grase and sofonolime in the two groups showed no significant difference (P > 0.05). In the same group, the peak and average of compound A concentration produced by Sodasorb grase and sofonolime showed significant difference with Drägersorb 800 plus (P < 0.05).

CONCLUSIONThe water content of sevoflurane and potassium hydroxide in CO2 absorbent can influence compound A production.

Absorption ; Carbon Dioxide ; chemistry ; Ethers ; chemistry ; Gas Chromatography-Mass Spectrometry ; methods ; Hydrocarbons, Fluorinated ; chemistry ; Methyl Ethers ; chemistry

To observe the gene polymorphism of Rh blood group in unrelated random individuals and families for Chinese Han nationality, polymerase chain reaction-sequence specific primer (PCR-SSP) was used to amplify the Rh C/E gene, RhD gene, exons, intron 2 and 10, insert and Rh Box in 160 blood samples of RhD positive unrelated individuals and 71 samples of RhD negative unrelated individuals and 7 samples of families whose probands were RhD-negative. The results showed that RhD genes of RhD-negative individuals with C antigens were polymorphism, three forms were found for D exon including intact, partial deletion and complete deletion exons. Insert fragments and Rh Box were found in most cases of families whose probands were RhD-negative and its inheritance accorded with the Mendel's Law, and it did not affect the expression of RhD gene. "Normal" RhD exon 4 amplifying product was not found in all of the samples. It was concluded that gene structure of the RhD-negative in Chinese was polymorphism, intact, partial deletion and complete deletion exons were found in the individuals with C antigen and probably existed specific D (nf) Ce haplotype. The function of insert was uncertain. The Rh gene sequences of Chinese Han nationality are different from those of Caucasian and the Rh gene library based on Han nationality should be established.
Asian Continental Ancestry Group ; genetics ; China ; ethnology ; Humans ; Introns ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Rh-Hr Blood-Group System ; genetics