In order to confirm the role that the 49th amino acid residue plays in enzymatic inactivity of Glutamine 49 phospholipase A2(Gln49-PLA2),site-directed mutagenesis of its 49th amino acid gene codon was conducted using PCR.Aspartic acid 49 phospholipase A2(Asp49-PLA2-Q49D-PLA2),the mutant of Gln49-PLA2 was expressed in E.coli with pET32a+ vector.The fusion protein,expressed as inclusion body,after being denatured,was on-column refolded and purified by immobilized metal affinity chromatography(IMAC),and then cleaved by Factor Xa.The mature Q49D-PLA2 mutant was obtained by Hitrap SP cation exchange and Superdex 75 gel filtration chromatography,with the recovery rate of 1.3%,and the specific activity of the mature Q49D-PLA2 mutant was 72 U/mg.It has been demonstrated that the 49th glutamine amino acid residue is the main reason in enzymatic inactivity of Gln49-PLA2 and the results are helpful for denatured protein refolding,especially in rich disulfide bonds conditions.

Animals ; Burns ; physiopathology ; Female ; Glycoproteins ; genetics ; metabolism ; Hydrogen Peroxide ; metabolism ; Male ; Mice ; RNA, Messenger ; genetics ; metabolism ; Skin ; injuries ; metabolism ; Wound Healing ; physiology

Hemagglutinin and neuraminidase, two important glycoproteins on the surface of influenza virus, play a considerable role in the entry and release stage of the viral life cycle, respectively. With in-depth investigation of influenza virus glycoproteins and the continuous innovation of drug discovery strategies, a new generation of glycoproteins inhibitors have been continuously discovered. From the point of view of medicinal chemistry, this review summarizes the current advances in seeking small-molecule inhibitors targeting influenza virus glycoproteins, hoping to provide valuable guidance for future development of novel antiviral drugs.

To study the effect of plant-derived smoke water on the accumulation of biomass and active substance of Salvia miltiorrhiza f. alba, seedlings of S. miltiorrhiza were treated with different concentrations of smoke water (1:500, 1: 1 000, 1: 2 000). The fresh weight and dry weight of underground part, the number of split-root, maximum root diameter, average root diameter, average root length, the content of lipophilic components and water-soluble components were measured. Results showed that fresh weight and dry weight of underground part were respectively improved by 98.01%, 44.32% and 85.71%, 28.57% with significant difference by smoke water treatment with concentration of 1: 500 and 1: 1 000. Maximum root diameter and dry weight of underground part were respectively enhanced by 58.44% and 85.71% by smoke water with concentration of 1:500. The content of tanshinone I and tanshinone II(A) were improved by smoke water treatment, however there were no significantly difference on the content of cryptotanshinone and dihydrotan shinone. This study indicates that smoke water treatment could be used to improve the accumulation of biomass and active substance content of S. miltiorrhiza f. alba, which could provide new ideas for its green cultivating.
Agriculture ; methods ; Biomass ; Drugs, Chinese Herbal ; analysis ; metabolism ; Salvia miltiorrhiza ; chemistry ; growth & development ; metabolism ; Smoke ; analysis ; Water ; chemistry ; metabolism

Trichosanthes kirilowii has been widely cultivated as its medicinal use, health care and food value. Drought resistance of seedlings is an important feature in breeding. Seeds of two T. kirilowii strains were used to research the difference of surface ultrastructure characteristic and drought resistance. Scanning electron microscope was used to identify the surface ultrastructure characteristic of seeds and PEG was used to simulate drought stress. The seeds germination rate, MDA content, chlorophyll content and the antioxidant enzymes activity were measured under the drought stress. The results showed that the seed surface colour of KXY-001 was lighter than that of KXY-005. The testa cobwebbing of KXY-001 was more intensive than that of KXY-005. The germination rate of KXY-001 was higher than that of KXY-005 under drought stress. The MDA content was increased and the chlorophyll content was decreased with the increasing of drought degree. The SOD activity of KXY-001 was higher than that of KXY-005, while the activity of POD and CAT was also increased firstly and decreased later. Surface reticulate of seeds and hilar traits can be used as identification points to identify the investigated strains. SOD and POD are activated to resist drought in T. kirilowii seedlings and the drought resistance of KXY-001 is superior than that of KXY-005.
Adaptation, Physiological ; drug effects ; Catalase ; metabolism ; Chlorophyll ; metabolism ; Droughts ; Germination ; Malondialdehyde ; metabolism ; Microscopy, Electron, Scanning ; Peroxidase ; metabolism ; Polyethylene Glycols ; pharmacology ; Seedlings ; metabolism ; Seeds ; growth & development ; metabolism ; ultrastructure ; Species Specificity ; Superoxide Dismutase ; metabolism ; Trichosanthes ; classification ; growth & development ; metabolism

Objective To investigate effects of exogenous substance P (SP) on proliferation of humau skin fibroblast (HSF) and expression of its monocyte chemoattractant protein-1 (MCP-1) under high glucose condition.Methods HSF cultured in high glucose DMEM medium were treated with SP of various concentrations at different time points.Levels of MCP-1 in the supernatants were determined by ELISA and time-effect relationship of SP regulating expression of MCP-1 was analyzed.Expression of MCP-1 mRNA in HSF was detected by semi-quantitative RT-PCR and Real Time-PCR,Conclusion,while cxpression of MCP-1 by Western blot.Proliferation of HSF treated with SP of different concentrations was detected by cell counting Kit-8 (CCK-8).Results MCP-1 had a strong expression at 8 hours after SP disposal (P ＜0.05) and reached the peak at 24 hours (P ＜ 0.01).Meanwhile,MCP-1 expressed the most at 10 nmol/L and 100 nmol/L SP (P ＜0.01).SP significantly enhanced the expression of MCP-1 mRNA and its synthesis under high glucose condition,especially at the concentration of 10 nmol/L (P ＜0.01).Also,SP induced obvious proliferation of HSF at concentrations of 10 nmol/L and 1 000 nmol/L(P ＜ 0.05).Conclusion SP promotes proliferation of HSF and expression of MCP-1 in high glucose DMEM medium,which may be of significance in promoting diabetic wound healing.

Objective To establish a stable animal model for sequentially dynamic and direct monitoring of the skin wound infection. Methods The mice with full-thickness skin incisions were replicated. After immediate subcutaneous suture,the mice were randomly divided into four groups,ie,Group A was inoculated with 50 μl sterile PBS solution),Groups B,C and D were inoculated with 50 μl suspension containing 1 × 106,1 × 108 and 1 × 1010 colony forming unit (CFU)/ml bioluminescent methicillin-resistant staphylococcus aureus (MRSA) respectively.Then,the diet behavior of each group was observed and the mean weight and mortality of each group were also recorded at different time points.The bioluminescent intensity of fluoresce in the wounds was recorded at different time points by using the charge-coupled device (CCD) based imaging system.Local wound tissues were incised at 24 hours after inoculation for HE staining so as to observe wound inflammatory reaction in each group.Wound healing time of each group was also recorded. Results ( 1 ) Average weight:Groups A and B showed unobvious changes in weight; Group C lightened until day 3 after inoculation and then recovered gradually to the preinoculation level at day 14; Group D lightened gradually until death.(2)Mortality:Groups A and B had no death; Group C had 10％ deaths at day 14; Group D had 100％ deaths.(3) Bioluminescent intensity of wounds:Groups A and B showed a gradual weakened luminescence since the day of inoculation and had almost complete disappearance at days 5 and 7 respectively; there was no sign of obvious increase or decrease in Group C from the day of inoculation till day 14 ; Group D had a gradual increase since the day of inoculation and the luminous area expanded until the death.(4) HE staining at 24 hours after inoculation:all the four groups showed inflammatory cell infiltration,especially in Groups C and D.(5) Wound healing time:wound healed at days 5 and 7 after inoculation in Groups A and B; the wounds showed no healing even at day 14 in the Group C,but the wounds length and area did not show obvious enlargement or diminishment ; the wounds extended gradually until the death in the Group D,since the day of inoculation. Conclusions The inoculation of 50 μl suspension with 1 × 108 CFU/ml bioluminescent MRSA to full-thickness skin incision rats allows direct,real-time dynamic and continuous detection of the occurrence and development of the wound infections.The infection model is easy to make and has stability and high repeatability.

Objective: To observe the clinical efficacy of tuina manipulations in treating different types of tic disorders (TD). Methods: Eligible TD patients were classified into three types, transient tic disorders (TTD), chronic multiple tic disorders (CMTD) and Tourette syndrome (TS), according to their disease duration and severity. The three types of children were treated with the same tuina manipulations. Changes in the Yale global tic severity scale (YGTSS) score, effective rate for tic, and cervical spine imaging examination results (including asymmetry of the lateral atlanto-dental interval, broadened anterior atlanto-dental interval, C2 spinous process deviation, occipito-atlanto-axial flexion/ extension instability) were observed after 1-month and 3-month treatments respectively. Results: The YGTSS score changed significantly after 1-month and 3-month treatments compared with that before treatment (both P<0.01); the effective rate for TD was 46.6％ and 86.7％ respectively after 1-month and 3-month treatments; there were significant differences comparing the effective rate for tic between different types of TD after 1-month and 3-month treatments (all P<0.05); comparing the effective rate for tic after 1-month treatment with that after 3-month treatment for the same type, the intra-group differences were statistically significant [TTD group (P<0.01), CMTD group (P<0.01), TS group (P<0.05)]; the abnormal parameter rates in neck imaging examination after 3-month treatment were significantly different from those before treatment (all P<0.01). Conclusion: Tuina manipulation is effective for TTD, CMTD and TS. It can correct the abnormal alterations of patients' cervical vertebrae, and its efficacy for TTD is most significant.

OBJECTIVETo study the clinical value of expression of peripheral blood neutrophil CD64 in adults with measles complicating pneumonia.

METHODS106 patients were divided into two groups by clinical manifestation and bacteria study: measles complicating bacterial pneumonia group and measles complicating viral pneumonia, using flow cytometry determination of CD64, C-reactive protein (CRP) and white blood cell (WBC) count.

RESULTSThe expression of CD64 in the bacterial pneumonia group with eruptive stage was (32.15 +/- 11.07) MFI, which was significantly higher than that in the group of with recovery stage (10.6 +/- 3.23) MFI (P < 0.01) and viral pneumonia (9.55 +/- 3.48) MFI (P < 0.01). These markers were considered positive if CD64 > or = 8.5 MFI, CRP > or = 10 mg/L and WBC > or = 9.05 x 10(9)/L. Their sensitivity was 78.12%, 80.48% and 59.37%. Their specificity was 76.19% ,67.67% and 64.28%. Their accuracy rate was 77.35%, 74.52%, 61.32%; CD64 has a positive relationship with CRP.

CONCLUSIONCompared to CRP, expression of peripheral blood neutrophil CD64 can be a better marker in the early diagnosis of patients with measles complicating bacterial pneumonia and as one of the indicators of disease conditions.

Adult ; Biomarkers ; blood ; metabolism ; C-Reactive Protein ; metabolism ; Female ; Humans ; Leukocyte Count ; methods ; Male ; Measles ; blood ; immunology ; microbiology ; Pneumonia, Bacterial ; blood ; immunology ; virology ; Receptors, IgG ; biosynthesis ; blood ; immunology

The data collected from 34 type 2 diabetic patients receiving intensive insulin therapy for six years showed that the yearly mean HbA1C was less than 7.0％,and none of the patients showed severe hypoglycemia,occurrence or evident progression of retinopathy or nephropathy,and the islet β cell function gained improvement.The DQOL score,used to evaluate the quality of patients' life had no significant change during the observation ( P ＞0.05 ).It is satisfactory and safe to maintain long-term glycemic control with prolonged intensive insulin therapy in patients with type 2 diabetes,and that such therapy does not induce untoward influence on the quality of diabetic patients life.