Objective To investigate the release kinetics of Zn2+ from nZCP-loaded polylactic acid/hydroxyapatite(PLA/HA)composite scaffold(PHZ)and determine the optimal nZCP content in the scaffold.Methods The particle size of nZCP was measured by DLS measurement,and PXRD,FTIR,and SEM were used to characterize the scaffolds and nZCP distribution;EDS was used to analyze element composition of the scaffold.Compression strength of the scaffold was determined,and ion release profile was investigated using ICP-MS.The biocompatibility of the materials was evaluated by CCK-8 assay and dead/alive staining of rat bone marrow stem cells(BMSCs)incubated with their aqueous extracts.ALP staining,alizarin red staining,RT-qPCR,and Western blotting were used to assess the osteogenic potential of the treated cells.In a rat model of bilateral ovariectomy(OVX)with femoral condylar bone defect,PHZ-1,PHZ-2,PHZ-3 or PLA/HA scaffold was implanted into the bone defect,and bone repair was observed using a microCT scanner and histological staining at 6 and 12 weeks.Results DLS,PXRD,SEM,FTIR,and EDS confirmed successful synthesis of 10-nm ZCP and efficient nZCP loading in the scaffold.PHZ-2 and PHZ-3 had significantly greater compression strength than PLA/HA.ICP-MS showed that Zn2+ release from PHZ-1,PHZ-2 and PHZ-3 were all optimal for promoting osteogenesis.In rat BMSCs,all the 4 scaffolds showed good biocompatibility,and their extracts enhanced ALP activity and extracellular matrix mineralization and promoted expressions of ALP,RUNX2,and OCN in the cells.In the rat models,nZCP in the implants improved bone graft integration at 6 weeks,and PHZ-2 and PHZ-3 more effectively induced new bone formation at 12 weeks(P<0.05).Conclusion PHZ scaffold is capable of stable Zn2+ release to promote osteoporotic bone defect healing,and PHZ-2 and PHZ-3 scaffolds with nZCP mass fraction of 4.5%-7.5%have better osteogenic activity.

Objective To investigate the release kinetics of Zn2+ from nZCP-loaded polylactic acid/hydroxyapatite(PLA/HA)composite scaffold(PHZ)and determine the optimal nZCP content in the scaffold.Methods The particle size of nZCP was measured by DLS measurement,and PXRD,FTIR,and SEM were used to characterize the scaffolds and nZCP distribution;EDS was used to analyze element composition of the scaffold.Compression strength of the scaffold was determined,and ion release profile was investigated using ICP-MS.The biocompatibility of the materials was evaluated by CCK-8 assay and dead/alive staining of rat bone marrow stem cells(BMSCs)incubated with their aqueous extracts.ALP staining,alizarin red staining,RT-qPCR,and Western blotting were used to assess the osteogenic potential of the treated cells.In a rat model of bilateral ovariectomy(OVX)with femoral condylar bone defect,PHZ-1,PHZ-2,PHZ-3 or PLA/HA scaffold was implanted into the bone defect,and bone repair was observed using a microCT scanner and histological staining at 6 and 12 weeks.Results DLS,PXRD,SEM,FTIR,and EDS confirmed successful synthesis of 10-nm ZCP and efficient nZCP loading in the scaffold.PHZ-2 and PHZ-3 had significantly greater compression strength than PLA/HA.ICP-MS showed that Zn2+ release from PHZ-1,PHZ-2 and PHZ-3 were all optimal for promoting osteogenesis.In rat BMSCs,all the 4 scaffolds showed good biocompatibility,and their extracts enhanced ALP activity and extracellular matrix mineralization and promoted expressions of ALP,RUNX2,and OCN in the cells.In the rat models,nZCP in the implants improved bone graft integration at 6 weeks,and PHZ-2 and PHZ-3 more effectively induced new bone formation at 12 weeks(P<0.05).Conclusion PHZ scaffold is capable of stable Zn2+ release to promote osteoporotic bone defect healing,and PHZ-2 and PHZ-3 scaffolds with nZCP mass fraction of 4.5%-7.5%have better osteogenic activity.

OBJECTIVE To establish the fingerprints of Ligusticum sinense from different habitats ，screen differential components and determine their contents. METHODS Using Z-ligustilide as reference ，HPLC fingerprints of 12 batches of L. sinense were established by using Similarity Evaluation System of Chromatographic Fingerprints of TCM （2012 edition）；common peaks were identified and their similarities were evaluated. Cluster analysis （CA），principal component analysis （PCA）and orthogonal partial least squares-discriminant analysis （OPLS-DA）were performed to screen differential components with variable importance in the projection （VIP）＞1 as standard ；meanwhile，the contents of above differential components were determined by the same HPLC method. RESULTS There were 17 common peaks in the fingerprints of 12 batches of L. sinense ，and their similarities ranged 0.989-1.000. A total of 9 common peaks were identified ，i.e. chlorogenic acid （peak 1），ferulic acid （peak 2）， senkyunolide Ⅰ（peak 7），coniferyl ferulate （peak 9），E-ligustilide（peak 13），senkyunolide A （peak 14），Z-ligustilide（peak 17）. CA results showed that 12 batches of L. sinense were divided into 3 categories，S1-S5（Wuning）were clustered into one category，S6-S8（Ruichang）were clustered into one category ，S9-S12（De’an）were clustered into one category ；the VIP values of peaks 2，13，14 and 17（corresponding to ferulic acid ，E-ligustilide，senkyunolide A ，and Z-ligustilide respectively ）were all greater than 1，respectively. In S 1-S5，S6-S8 and S 9-S12 samples，the contents of ferulic acid were 0.488-0.533，0.603-0.658 and 0.415-0.433 mg/g，respectively；senkyunolide A were 1.184-1.295，1.450-1.588 and 1.307-1.377 mg/g，respectively；E-ligustilide were 0.118-0.125，0.130-0.135 and 0.223-0.229 mg/g，respectively；Z-ligustilide were 7.200-7.681，8.076-8.643 and 4.508-4.996 mg/g， respectively；the differences between two groups were statisti-cally significant （P＜0.05）. CONCLUSIONS Established ARS-11）；fingerprint is simple and accurate ，and can be used for overall quality evaluation of L. sinense from different habitats by combining with multivariate statistical analysis. Ferulic acid ， senkyunolide A ，Z-ligustilide and E-ligustilide may be the differential components that affect the quality of L. sinense from different habitats ，the contents of the first 3 components in L. sinense from Ruichang are the highest ，and the content of E-ligustilide in samples from De’an is the highest.

OBJECTIVETo investigate the effect of modified bone morphogenetic protein-2 polylactic acid nanospheres (BMP-2-PLA-Ns) sustained-release system on rabbit mandibular defect repair.

METHODSThe polylactic acid s nanospheres (PLA-Ns) and BMP-2-PLA-Ns were prepared by ultrasonic emulsification after graft polymerization. Forty-five rabbits were randomly divided into 3 groups: Blank group, PLA-Ns gel group(control group), and BMP-2-PLA-Ns gel group (experimental group). The rabbit mandibular defect models were established. The defect area of control group was implanted with PLA-Ns gel, meanwhile, the experimental group was implanted with BMP-2-PLA-Ns gel, the blank group experienced no special handling. Rabbits were killed in 1, 2, 4 weeks after operation and the iconography, hematine eosin(HE) staining and PCNA immunohistochemistry were used to detect the reparative effect on rabbit mandible defects.

RESULTSImage observation showed that bone defect repair in the experimental group was well and the shadow was not obvious. Better repair effect was seen compared with the control group and blank group. HE staining showed that the experimental group and the control group had a large number of neovascularization and secondary callus formation, callus in experimental group was obviously higher than that of control group and blank group. Immunohistochemical observation showed that the experimental group's PCNA positive chondrocytes were more than those in the control group and the blank group in the first 2 weeks; all groups of PCNA positive cells were rare in the fourth week, PCNA positive expression rate of the fourth week was lower than that of the first 2 weeks.

CONCLUSIONThe modified BMP-2-PLA-Ns sustained-release system promotes mandibular defect repair obviously.

Animals ; Bone Morphogenetic Protein 2 ; Delayed-Action Preparations ; Lactic Acid ; Mandible ; Nanospheres ; Polyesters ; Polymers ; Rabbits ; Reconstructive Surgical Procedures

Objective To compare the clinic efficacy and safety of renal osteodystrophy between hemodiafiltration (HDF)com-bined low dose calcitriol and high flux hemodialysis combined high dose Calcitriol .Methods 25 patients ,who were in accordance with the inclusion criteria ,were randomly divided into HDF combined low dose Calcitriol group (group HDF) (n=13) and high flux hemodialysis combined high dose Calcitriol (group VitD)(n=12) .Patients in group HDF took Calcitriol for 0 .5μg after every dialysis session ,while those in group VitD took it for 1 .0 μg after dialysis at first ,then the dose would be adjusted every 4 weeks according to the level of blood Calcium .The intervention continued for 16 weeks in total .Then detecting the levels of intact parathy-roid hormone (iPTH) ,bone alkaline phosphatase (BAP) ,serum calcium (Ca) ,serum phosphorus (P) ,and calculating Ca × P sedi-mentation and clinical symptom score .Results he level of Ca in group HDF and group VitD increased significantly after interven-tion (P= 0 .000) .No difference was no statistically significant between the two groups (P= 0 .141) .The P ,iPTH ,BAP of two groups were significantly decreased (P<0 .05) .P in group HDF decreased more significantly(P=0 .003) .iPTH and BAP in group VitD decreased more significantly (P=0 .034 ,0 .046);Ca × P in group HDF decreased significantly (P=0 .031) ,but increased obvi-ously in group VitD(P=0 .037) ,which was much higher than group HDF(P=0 .001) .Clinical symptom in the two groups im-proved significantly after intervention(P<0 .05) ,There were no significant difference between the two groups(P=0 .632) .Conclu-sion The clinic efficacy of HDF combined low dose calcitriol is same as that of high flux hemodialysis combined high dose calcitri-ol .It could also reduce calcitriol side-effect ,which is an effective theraphy for high turnover osteodystrophy .

OBJECTIVETo establish a method of comprehensive chemical pattern recognition of plantain seed via HPLC fingerprint, principal component analysis (PCA) and cluster analysis.

METHODThe chromatographic separation was performed on a C18 column (4.6 mm x 250 mm, 5 microm). The mobile phase was a mixture of acetonitrile-0.5% acetic acid aqueous solution in gradient elution. The HPLC fingerprint of ethyl acetate fraction of 24 batches Plantaginis Semen from different habits and varieties was set up and 10 common peaks were obtained.

RESULTThe result of the principal component analysis (PCA) and cluster analysis is similar but there is disparity between them.

CONCLUSIONThe method could be used for the quality control and comprehensive evaluation of Plantaginis Semen.

Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; chemistry ; Plants, Medicinal ; chemistry ; Principal Component Analysis

Objective To investigate the mechanism of treatment of granulocyte colony-stimulating factor(rhG-CSF),recombinant human interleukin-11(rhIL-11)and recombinant human interleukin-2 (rhIL-2)on hematopoietic injuries induced by 4.5 Gy60 Coγ-ray irradiation in beagles,and to provide experimental evidence for the clinical treatment of extremely severe myeloid acute radiation sickness (ARS).Methods Sixteen beagle dogs were given 4.5 Gy60 Co γ-ray total body irradiation(TBI),then randomly assigned into irradiation control group,supportive care group or cytokines+supportive care (abbreviated as cytokines)group.In addition to supportive care,rhG-CSF,rhlL-11 and rhIL-2 were administered subcutaneously to treat dogs in cytokines group.The percentage of CD34+cells,cell cycle and apoptosis of nucleated cells in peripheral blood were examined by Flow cytometry.Results After 4.5 Gy 60 Co γ-ray irradiation,the CD34+cells in peripheral blood declined obviously(61.3%and 52.1% of baseline for irradiation control and supportive care group separately).The cell proportion of nucleated cells in Go/G1 phase was increased notably(99.27% and 99.49% respectively).The rate of apoptosis(26.93% and 21.29% separately)and necrosis(3.27% and 4.14%,respectively)of nucleated cells were elevated significantly when compared with values before irradiation(P<0.05) 1 d post irradiation.When beagles were treated with cytokines and supportive care,the CD34+cells in peripheral blood were markedly increased(135.6% of baseline).The effect of G0/G1 phase blockage of nucleated cells became more serious(99.71%).The rate of apoptosis(5.66%)and necrosis(1.60%)of nucleated cells were significantly lower than that of irradiation control and supportive care groups 1 d after exposure.Conclusions Cytokines maybe mobilize CD34+cells in bone marrow to peripheral blood,indce cell cycle block at G0/G1 phase and reduce apoptosis,and eventually cure hematopoieticinjuries induced by irradiation.

Objectivc To observe the therapeutic effects of combined cytokines on hematopoietic injuries induced by 4.5 Gy60 Co γ-rays irradiation in beagles,and to provide experimental evidences for the clinical treatment of extremely severe myeloid acute radiation sickness(ARS).Methods 16 beagles were given 4.5 Gy60 Co γ-rays total body irradiation,and then randomly assigned into irradiation control group,supportive care group and cytokines group.In addition to supportive care,recombinant human granulocyte colony-stimulating factor (rhG-CSF),recombinant human interleukin-11(rhIL-11)and recombinant human interleukin-2(rhIL-2)were administered subcutaneouly to dogs in cytokines group.Peripheral blood hemogram was examined once every two days.Bone marrow and peripheral blood were collected to proceed colony cultivation 4 d pre-irradiation and 1 and 45 d post-irradiation.Conventional histopathological sections of sternum were prepared to observe the histomorphology changes. Results After irradiation,the population of all kinds of cells in peripheral blood declined sharply.WBC nadir Was elevated(1.04×109/L,but 0.28×109/L and 0.68×109/L for the irradiation control group and the supportive care group separately),the duration of thrombocytopenia was shortened (24 days,but 33 days for the supportive care groug) and red blood cell counts were maintained in the range of normal values after cytokincs treatment in combination.The colony forming efficiency of haemopoietic stem cells(HSCs)in bone marrow and peripheral blood decreased obviously 1 d post irradiation,but recovered to the level of that before irradiation 45 d post irradiation after supportive care and cytokines treatment.Hematopoietic cells disappeared in bone marrow of animals in irradiation control group,but hematopoietic functions were recovered after cytokines were administrated.Conclusions RhG-CSF.rhIL-11 and rhIL-2 used in combination could elevate WBC nadir,accelerate the recovery of leukocytes,platelets and red blood cells and promote the proliferation,differentiation and maturity of HSPCs left in the body after 4.5 Gy γ-rays total body irradiation,eventually restore the hematopoietic function.Hence,combination of rhG-CSF,rhIL-11 and rhIL-2 could serve as better therapeutic strategy to treat extremely severe myeloid ARS.

Objective To explore the mechanisms of cytokines on acute radiation disease in irradiated beagles.Methods The sera of beagles irradiated with 4.5 Gy γ-rays with cytokines treatment was collected at different time points post irradiation.The two-dimensional gel electrophoresis(2-DE)was used to isolate and compare the differentially expressed proteins in sera.HD-MS was used to analyze the differentially expressed proteins with significance,and the amino acid sequences should be determined. Results High resolution 2-DE gel map was obtained.There were six differentially expressed proteins in sera of irradiated beagles at different time points.Four protein spots were successfully identified by MS.A significant spot was identified as serum amyloid A(SAA)by HD-MS,with relative molecular mass of 13 077 and isoelectfie point of 6.26.Expression of SAA was not found 1 d pre-irradiation and 36 d postirradiation,but increased slightly 1 d(0.2166)and significantly 14 d post-irradiation(0.4577). Conclusions The expression of serum amyloid A was consistent with the process of acute radiation injury,which might indicate the turnover of the disease.

Objective To evaluate the effects of combined administration of recombinant human interleukin-11(rhIL-11),recombinant human G-CSF(rhG-CSF)and recombinant human interleukin-2 (rhIL-2)on acute radiation sickness(ARS)beagles.Methods Sixteen beagle were irradiated with 4.5 Gy60 Co γ-rays to establish ARS models,and were divided into irradiation control group,supportive care group and combined cytokines treatment group.After irradiation irradiation control group was given no treatment,the dogs in supportive care group received purely symptomatic treatment,while combined cytokines treatment group received rhIL-11 50μg/(kg·d)and rbG-CSF 10μg/(kg·d)subcutaneously(0-14 d)and rhIL-2 1×1 06 U/d(29-43 d)besides symptomatic treatment.Manifestation and characteristics of ARS beagles were observed,and the survival time were recorded.At last,post-mortem examination and histological examination were performed.Results All animals underwent nausea,diarrhea and fever.After irradiation,all animals in irradiation control group died in two weeks,and the mean survival time was 12.7 d,while only one died at 33 d in supportive care group.All dogs in combined cytokine group survived at 45 day after exposure,and their haematopoiesis and gastrointestinal tract were recovered.Conclusions Combination of rhIL-11 + rhG-CSF + rhIL-2 treatment could be significantly effective on ARS beagles irradiated by 4.5 Gy60 Co γ-rays,which could accelerate injured haemotopoiesis and intestinal tract recovery,increase the survival rate and improve the life quality of animals.