Objective To compare magnetic beads kit,agrose gel recovery kit and heparinase I three methods to purify the micro DNA from crude heparin, then use q-PCR to identify the species origins and select the best method.Methods Using magnetic beads kit,agrose gel recovery kit and heparinase I to purify micro DNA from crude heparin and combined the porcine,bovine and ovine identification kits to identify the species origins and conformed the minimum detection limit of different percentage of ovine crude heparin in porcine crude heparin.Results Three pretreatment methods all can solve the pretreatment difficulties and we found that the haparinase was the best method; the minimum detection limit was 0.01%of ovine crude heparin in porcine crude heparin.Conclusion The heparinase method is the best pretreatment method and can successfully solve the pretreatment difficulties.Heparinase combine the porcine, bovine and ovine identification kits can identify the species origins from crude heparin.

Objective A high performance liquid chromatographic (HPLC) method was established for the determination of glycerol in propofol medium and long chain fat emulsion injection.MethodsThe chromatographic conditions were as follows: Kromasil 100-5-NH2 column(4.6×250mm,5μm) with the column temperature was 40℃,acetonitrile-water(8515)as mobile phase with flow rate of 1.0mL/min.Glycerol was detected by refractive index (RI) detector at 40℃.ResultsThe linear range of glycerol was 455.3916-2276.9580μg/mL(r=0.9999,n=7),the average recovery rate was 99.5%,RSD was 0.6%(n=9),the limit of detection(LOD) was 121ng and the limit of quantification(LOQ)was 364ng.ConclusionThe method was simple, rapid, strong specifity and accurate with good reproducibility, which is suitable for the content determination of glycerol in propofol medium and long chain fat emulsion injection.

Objective To evaluate the role of spinal c?Jun N?terminal kinase ( JNK ) signaling pathway in incisional pain in rats. Methods Sixty?three adult male Sprague?Dawley rats, weighing 200-250 g, were randomly divided into 3 groups ( n=21 each) using a random number table: incisional pain group ( IP group) , dimethyl sulfoxide ( DMSO) group, and JNK inhibitor SP600125 group ( SP group) . A 1?cm longitudinal incision was made through skin, fascia and muscle of the plantar aspect of the hindpaw in anesthetized rats. In group DMSO, 10% DMSO 10 μl was injected intrathecally at 30 min before surgery. In group SP, SP600125 25 μg (in 10 μl of 10% DMSO) was injected intrathecally at 30 min before sur?gery. Six rats in each group were sacrificed, and the mechanical paw withdrawal threshold ( MWT) and thermal paw withdrawal latency ( TWL) were measured at 24 h before establishment of the model and 2, 6, 24, 48 and 72 h after establishment of the model. After measurement of the pain threshold at 24 h before establishment of the model and 6, 24, 48 and 72 h after establishment of the model, the lumbar segment of the spinal cord was removed for determination of the expression of phosphorylated JNK ( p?JNK) by im?munofluorescence. Results The MWT was significantly lower, the TWL was shorter, and the expression of p?JNK was lower at each time point after establishment of the model than at 24 h before establishment of the model in group IP (P<0?05). Compared with group IP, the MWT was significantly increased, the TWL was prolonged, and the expression of p?JNK was down?regulated at each time point after establishment of the model in group SP ( P<0?05) , and no significant change was found in the parameters mentioned a?bove in group DMSO ( P>0?05) . Conclusion Spinal JNK signaling pathway is involved in the develop?ment and maintenance of incisional pain in rats.

Objective To establish acute hepatotoxic model induced by Amanita exitialis and to study the characteristics of acute toxic liver failure induced by mushrooms containing peptide toxins,in hope for providing some help to experimental research on poisoning induced by mushrooms containing peptide toxins.Methods UPLC-MS/MS (Ultra performance liquid chromatography-tandem mass spectrometry) method was used to detect peptide toxins in Amanita exitialis.To establish acute toxic liver hepatic failure model,the beagles were fed with 60 mg/kg of lyophilized powder of Amanita exitialis fungus which encapsulated in starch capsules.Toxic sighs were observed,coagulation function,hepatic and renal function,liver histopathological morphology,peptide toxin concentration in plasma and urine were detected during the experiment.Results Total peptide toxins in Amanita exitialis was (3 482.6 ± 124.94) mg/ kg.All the beagles had toxic signs including vomiting and diarrhea in 12-48 h after ingestion.On 24 h after ingestion,the beagles' ALT,AST,TBIL,ALP,PT and APTT levels increased obviously.On 36 h after ingestion,the beagles' ALT,AST,PT and APTT values reached their peaks (ALT:283.2 ± 112.9 Kallmann unit;AST:223.9 ±93.8 Kallmann units;PT:132.9 ± 152.6 s;APTT:131.4 ± 153.9 s).On 48 h after ingestion,the beagles' TBIL and ALP levels reached their peaks (TBIL:23.3 ± 14.6 mol/L;ALP:274.5 ± 115.5 U/L).The beagles' TBIL,TP and APTT returned to normal 1 week after ingestion,their ALT,AST and ALP levels returned to normal 3 weeks after ingestion.Three dogs died during 24-72 h after ingestion.Liver histopathological morphology study showed hemorrhagic necrosis of hepatocytes.Peptide toxins can be detected in plasma within 24 h after ingestion.Peptide toxins can be detected in urine within 96 h after ingestion.Conclusion Amanita peptide toxins can cause hemorrhagic necrosis of liver cells and lead to acute liver failure.This model is consistent with clinical pathophysiological process of acute toxic liver failure induced by mushrooms containing peptide toxins,and it can be applied to the study of diagnosis and treatment of poisoning induced by mushrooms containing peptide toxins.

Objective:To explore clinical significance of sinus heart rate turbulence (HRT) phenomenon in aged pa‐tients with stable angina pectoris (SAP) .Methods :A total of 120 aged SAP patients ,who received 24h DCG in our hospital from Jan 2013 to Oct 2015 ,were selected as SAP group .Meanwhile ,another 144 aged patients ,who re‐ceived 24h DCG examination simultaneously and coronary angiography results were normal ,were regarded as nor‐mal control group .According to coronary lesion severity ,SAP group was further divided into single vessel coronary disease group (single vessel group ,n=35) ,double‐vessel coronary disease group (double‐vessel group ,n=48) and multi‐vessel coronary disease group (multi‐vessel group ,n=37) .The 24h DCG ,HRT indexes ,including turbulence onset (TO) and turbulence slope (TS) ,were measured and compared among all groups .Results:Compared with normal control group ,there was significant rise in TO [(0.77 ± 0.37)% vs .(1.26 ± 0.92)% ] and significant reduc‐tion in TS [(5.45 ± 4.02) ms/RR interval vs .(1.53 ± 0.70) ms/RR interval] ,P<0.01 both ;significant rise in ab‐normal rates of TO (19.44% vs .42.50% ) ,TS (15.97% vs .31.67% ) and TO + TS (11.11% vs .30.83% ) in SAP group ,P<0.01 all .Compared with single vessel group ,there was significant rise in TO [(0.66 ± 0.22)% vs .(1.28 ± 1.11)% vs .(1.46 ± 1.20)% ] and significant reduction in TS [ (2.04 ± 0.82) ms/RR interval vs .(1.66 ± 0.38) ms/RR interval vs .(1.29 ± 0.58) ms/RR interval] in double‐vessel group and multi‐vessel group ,and TO of multi‐vessel group was significantly higher than that of double‐vessel group ,TS of multi‐vessel group was significantly low‐er than that of double‐vessel group , P<0.01 all .Conclusion:Sinus heart rate turbulence can be used as risk predic‐tor for aged patients with stable angina pectoris ,which can provide basis for clinical effective treatment and progno‐sis assessment .

OBJECTIVEThis aimed to investigate the effect of specific sequence oligodeoxynucleotide MT01 on the biological properties of osteoblasts invaded by Porphyromonas gingivalis (P. gingivalis ) by evaluating proliferation, cell cycle, and apoptosis.

METHODSMG63 osteoblasts were recovered and incubated with MT01, CpG ODN, metronidazole (MNZ), and gentamicin (GEN) for 3 h. P. gingivalis (the multiplicity of infection was 100:1) was added subsequently and cocultured for another 24 and 48 h. Cells with PBS comprised the blank group, whereas cells with P. gingivalis comprised the negative controls. Six experimental groups were established: PBS group, P. gingivalis group, MT01+P. gingivalis group, CpG ODN+ P. gingivalis group, MNZ+P. gingivalis group, and GEN+P. gingivalis group. The proliferative ability was measured by methyl thiazolyl tetrazolium assay, and the percentages of apoptosis and cell cycle were examined by flow cytometry.

RESULTSCompared with the blank group, proliferation increased significantly in the MT01+P. gingivalis group (P < 0.05). The ratio of cells was lower at the G₁ phase and higher at the S phase in the MT01+P. gingivalis group compared with the results in the P. gingivalis group (P < 0.05). Early cell apoptosis in the MT01+P. gingivalis group was significantly lower than that in the P. gingivalis group (P < 0.05).

CONCLUSIONMT01 can promote the proliferation, reduce the ratio of the G₁phase, increase the ratio of the S phase, and inhibit the early apoptosis of osteoblasts invaded by P. gingivalis.

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Division ; Cell Proliferation ; drug effects ; Flow Cytometry ; Gentamicins ; pharmacology ; Humans ; Metronidazole ; pharmacology ; Oligodeoxyribonucleotides ; pharmacology ; Osteoblasts ; cytology ; drug effects ; Porphyromonas gingivalis ; pathogenicity

OBJECTIVEThis paper aimed to determine the mRNA expression of osteoclast-related factors interleukin-6 (IL-6), interleukin-12 (IL-12) p35, IL-12p40, matrix metalloproteinase-9 (MMP-9), nuclear factor of activated T-cells cytoplasmic 1 (NFATcl), receptor activator of nuclear factor-KB (RANK), and tumor necrosis factor-α (TNF-α) mRNA in murine macrophages infected by a periodontitis patient's own tissue nucleic acid. Another aim was to investigate the effects of a periodontitis patient's own tissue nucleic acid on the differentiation of macrophages into osteoclasts.

METHODSInflammatory periodontal tissue samples of chronic periodontitis patients were taken during periodontal flap surgery, and healthy gingival tissue samples were taken from orthodontic patients during tooth extractions. Total RNA from periodontal tissue was extracted and reversely transcribed into cDNA and then cryo-preserved until further use. First, specific sequence oligodeoxynucleotide MT0I at a concentration of 1 µg · mL⁻¹ was added in murine macrophage RAW264.7, and the cells were incubated for 3 hours. Cells with PBS (1 µg · mL⁻¹) were used as negative controls. The inflammatory periodontal tissue cDNA and healthy periodontal tissue cDNA (1 µg · mL⁻¹) was added subsequently. There were four experimental groups: healthy periodontal tissue cDNA+ RAW264.7, inflammatory periodontal tissue cDNA+RAW264.7, MT01+healthy periodontal tissue cDNA+RAW264.7, and MT01+inflammatory periodontal tissue cDNA+RAW264.7. Real-time quantitative polymerase chain reaction was used to detect the mRNA expression of osteoclast-related factors IL-6, IL-12p35, IL-12p4O, MMP-9, NFATcl, RANK, and TNF-α mRNA after 3, 6, 12, and 24-hours.

RESULTSThe mRNA levels of osteoclast-related factors NFATc1, MMP-9, TNF-a, IL-6, IL-12p40, IL-12p35, and RANK in RAW264.7 were markedly upregulated with the treatment of periodontitis patient's own tissue nucleic acid. However, the mRNA expression of osteoclast-related factors was inhibited by use of an immunosuppressant MT01.

CONCLUSIONThe periodontitis patient's own tissue nucleic acid could promote the differentiation of murine macrophage into osteoclasts.

Animals ; Cell Differentiation ; Cytokines ; metabolism ; Gene Expression ; Gingiva ; Humans ; Interleukin-12 Subunit p40 ; Interleukin-6 ; Macrophages ; Matrix Metalloproteinase 9 ; Mice ; Osteoclasts ; metabolism ; Periodontitis ; RNA, Messenger ; Tumor Necrosis Factor-alpha

The construction of featured specialties is the current development strategy of community health service institutions to improve the service scope and to meet the health needs of residents. The rehabilitation medicine has undergone 12 years of development and become a relatively mature featured specialty in Fenglin Community Health Service Center. Based on the Fenglin′s experience, this article discusses the development status and restriction bottlenecks of general practice, and the development status and trend of rehabilitation medicine in the community; and also explores the integrated development model of community-featured specialty with general practice.

Rehabilitation medicine is one of the most important specialties in community health institutions. This article introduces the 12 year′s development of rehabilitation medicine in Fenglin Community Health Service Center, focusing on the talent allocation, service capabilities, resource expansion, basic facilities, personnel recruiting, department operating, service scope, and its achievements and influence, to provide reference for planning and construction of featured specialty in community health service centers.

Community health institutions have entered a new development stage of featured specialty construction. After 12 years of development, rehabilitation medicine now is the featured specialty of Fenglin Community Health Service Center. This article presents the train of thought and key points of specialty construction in primary care institutions based on the Fenglin′s experience. The positioning of featured specialty should be based on the community. The construction process should include 7 elements, namely, the standard operation procedure(SOP)of service system construction, the detailed publicity and implementation of the collaboration of specialists, prevention and control knowledge promotion for general practitioners, prevention and control knowledge education for community residents, service list, clinical efficacy evaluation, and clinical database. In the later iterations, the head of the department should always focus on the service system construction SOP and clinical database construction, and the rest parts can be assigned to the relevant team members.