Objective: To investigate the protective effect and mechanism of rhubarb on intestinal mucosa epithelial cells apoptosis of piglet with sepsis.Methods: The changes of apoptosis of intestinal mucosa epithelial cells were tested by flow cytometry and TUNLE.Molecular biology methods were used to observe the mRNA expression of zonula occludens(ZO-1),Occludin,TNF-?/IL-10 and the content of MDA,activity of SOD.Results: Compared with model group,the number of apoptosis cells in treatment group decreased apparently(P

Objective To establish an HPLC method for the determination of ceftazidime for injection. Methods HPLC column was TSK-GEL G2000SWXL gel column. Mobile phase of phosphate buffer ( pH 7. 0) was 0. 005 mol · L-1 disodium hydrogen phosphate solution-0. 005 mol · L-1 sodium dihydrogen phosphate solution ( 61 : 39 ). The detection wavelength was 231 nm; column temperature was 30 ℃ ; the injection volume was 20 μL. Results Ceftazidime reference linear range was 0. 51-25. 64 μg·mL-1(r=0. 999), ceftazidime for injection polymer linear range was 0. 20-3. 92 mg·mL-1 (r=0. 99), and the limit of quantification polymer was 0. 71 μg. Conclusion The method is rapid and the separation was good. It can be used for the detection of ceftazidime for injection.

OBJECTIVE:To establish a method for the simultaneous determination of tetracycline hydrochloride and cortisone acetate in Cortisone tetracycline eye ointment. METHODS:HPLC was performed on the column of Phenomenex C18 and shimaduz GL C18 with mobile phase of 0.01 mol/L Sodium dodecyl sulfate solution(adjusted to pH 2.5 with phosphoric acid)-acetonitrile(60∶40,V/V)at flow rate of 1.0 ml/min,detection wavelength was 254 nm,column temperature was 30 ℃,and the injection volume was 20 μl. RESULTS:The linear range was 11.36-227.18 μg/ml for tetracycline hydrochloride(r=0.999 9)and 11.11-222.21 μg/ml for cortisone acetate(r=0.999 9);RSDs of precision,stability and reproducibility tests were no more than 1.2%;recoveries were 96.89%-100.67%(RSD=1.1%,n=9)and 100.04%-101.02%(RSD=0.3%,n=9),respectively. CONCLUSIONS:The method is simple,accurate and specific,and can accurately determine the contents of tetracycline hydrochloride and cortisone acetate in Corti-sone tetracycline eye ointment.

Aim To synthesize and identify artificial antigen of podophyllotoxin for the production of podophyllotoxin polyclonal antibody. Methods The hapten was synthesized by two different chemical approaches and characterized by TLC, IR, NMR, and MS. Mixed anhydride reaction (MAR) and active ester method (AEM) were used to couple the podophyllotoxin to carrier proteins (BSA and OVA). Characterization of artificial antigens was done by using spectroscopy and electrophoresis. The anti-podophyllotoxin polyclonal antibodies were obtained through immunizing rabbits. Results The results from IR, NMR and MS showed that 4-O-succinoyl podophyllotoxin (hapten) was successfully synthesized. The coupling molar ratios of the hapten and carrier proteins were 88.6 for Hapten-BSA1, 40.3 for Hapten-BSA2, 17.8 for Hapten-OVA1, and 54.2 for Hapten-OVA2. Hapten conjugates coupled with BSA yielded two sets of the specific and affinitive polyclonal antibodies. One set of antibodies showed an IC50 value of 2.21 μg·mL -1 with a detection limit of 0.12 μg·mL -1. Conclusion Antigenic conjugates were artificially synthesized, and based on these artificial antigens, polyclonal antibodies against podophyllotoxin were raised from rabbits immunized with two different immunogens and characterized with an indirect ELISA format.

Objective To explore the difference of the contrast-enhanced ultrasound (CEUS) image and quantitative parameters between two different contrast agents Fluorocarbon and SonoVue. Methods The tumor model of colorectal carcinoma in nude mice was established by injecting CT26 cells into the subcutaneous space in hepatic area of 15 nude mice. CEUS was performed with Fluorocarbon and SonoVue on the 14th day after establishment. SonoLiver software was used to analyze the dynamic image quantitatively. The difference of the CEUS image and quantitative parameters between the two contrast agents was analyzed. Results Compared with normal liver parenchyma around the tumor, the tumor ultrasound contrast performance was fast forward and rewind with low enhancement. There was no signiifcant difference between the two kinds of microbubbles not only for CEUS image but also for quantitative parameters [maximum intensity (Imax):49.53%±24.38%vs 45.04%±17.03%, rise time (RT):11.68 s±3.07 s vs 13.76 s±2.92 s, time to peak (TTP):12.76 s±4.12 s vs 15.26 s±3.74 s, T1/2:50.57 s±28.32 s vs 48.75 s±9.85 s, Imax/TTP(V1):4.48±2.82 vs 3.18±1.49, (Imax-INT60)/(60-TTP)(V2):0.67±0.34 vs 0.60±0.20, AUC1:3032.78%±1343.12%vs 3258.77%±1369.84%, AUC2:11647.38%±6183.10%vs 10439.04%±4604.65%, AUC:14680.17%±7469.85%vs 13697.81%±5831.99%, Rate of AUC1:264.25±146.93 vs 222.24±92.16, Rate of AUC2:241.67±119.97 vs 231.97±100.34, all P>0.05). No mouse was dead during the CEUS examination. Conclusion Fluorocarbon had similar CEUS imaging effect and quantitative information compared with SonoVue.

Objective To analyze the genomic sequences of Cryptococcus neoformans var grubii strains of two genotypes with different virulence and to screen out the virulence-associated genes. Methods A clinical strain (IFM56800) with the strongest virulence and an environmental strain (IFM56731) with the weakest virulence were screened out for whole genome sequencing analysis. The results of sequencing analy-sis were comprehensively analyzed by using the method of comparative genomics. Genetic variations were ex-tensively screened by using the strategies of non-synonymous single nucleotide polymorphisms ( nsSNPs), nonsense SNPs and the insertions or deletions ( InDels) causing frameshift mutations. The filtered genes were sequenced in 20 experimental strains. The whole RNAs were extracted and then the full-length cDNAs were sequenced by using the rapid amplification of 5′ and 3′ cDNA ends (RACE) method. Results By whole genome sequencing, valid data with high coverage (127 times and 111 times) was obtained in both the environmental strain IFM56731 and the clinical strain IFM56800. The data of InDels and SNPs were statisti-cally analyzed, respectively. Six genes were chosen for further analysis based on the strategies of nonsense SNPs and the InDels causing frameshift mutations. The six genes were amplified and sequenced in all of the experimental strains, three of which were further analyzed with cDNA sequencing. Ultimately, the location and structure of CNAG_01032 gene were determined. The predicted nonsense mutation locus was verified to present in the actual mRNA. Conclusion The strategies of nonsense SNPs and the InDels causing frame-shift mutations showed high-efficiency in screening potential virulence-associated genes. The CNAG_01032 gene was screened out as a novel virulence-associated gene.

Molecular beacon probe was designed based on a specific DNA sequence of Nocardia to PCR detection of thisbacterium.The strains of Nocardia、Gordina and Rhodococcus were inoculated in Brain Heart Infusion Agar medium separately,then the growth condition was observed,DNA was extracted as a template;the molecular beacon probe was designed based on the partial secA 1 gene sequences of Nocardia strains,and the probe was added into the reaction system of real time fluorescence quantitative PCR (RT-PCR),and the fluorescence signal was tested at the end of PCR.Showed that the amplified secA1 gene of Nocardia could produce positive fluorescence signal in RT-PCR,but those of Gordonia and Rhodococcus with control groups showed negative results because of no fluorescence signal.In conclusion as a housekeeping gene,secA1 is an ideal target molecule to identify the actinomycetes strains on the species level in the systematic evolution research,and the technique of fluorescence molecular beacon probe is accurate,rapid and sensitive for detecting the Nocardia strains with secA1 gene.

Objective: To observe the effect of acupuncture on the intestinal flora in Parkinson disease (PD) model mice and explore the mechanism of acupuncture in improving the locomotor function in PD. Methods: Thirty-two C57BL/6 mice were randomly divided into a control group, a 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP) group, a MPTP + acupuncture group (MPTP+A), and a MPTP + madopar group (MPTP+M), with 8 mice in each group. Except for the control group, the other groups were intraperitoneally injected [25 mg/(kg·bw)] with MPTP to establish PD mouse models. After successful modeling, the MPTP group received no intervention, the MPTP+A received acupuncture at Tianshu (ST25), Guanyuan (CV4), and Zusanli (ST36), and the MPTP+M was given madopar [125 mg/(kg·bw)] by intragastric gavage. After consecutive 10-day interventions, the intestinal function and behaviors of the mice were detected. The 16S rRNA gene sequence was used to analyze the composition of fecal intestinal flora in each group. Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay were used to detect the expression levels of inflammatory cytokines in the brain and serum. The expression levels of tyrosine hydroxylase (TH) and α-synuclein in the substantia nigra (SN) were detected by immunohistochemical staining. Toll-like receptor (TLR) 2 and lipopolysaccharide receptor CD14 (CD14) in the SN were determined by RT-qPCR. Myeloid differentiation factor (MyD) 88, nuclear factor kappa-B (NF-κB) and Akt1 in the SN were detected by Western blotting. Results: After the intervention, compared with the control group, the intestinal motility, fecal water content, and the expression of TH in the SN were significantly decreased in the MPTP group (P<0.05), along with an increased α-synuclein expression (P<0.05). Additionally, the results of the fecal microflora test showed that the alpha diversity of the MPTP decreased, and the levels of inflammatory cytokines [tumor necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS), interleukin (IL)-1β, and IL-6] in the serum and SN, and the expression of NF-κB in the SN were significantly increased (P<0.05). Compared with the MPTP group, acupuncture intervention significantly enhanced the autonomous horizontal movement and coordination ability of PD mice (P<0.05); acupuncture and madopar interventions significantly reduced the levels of α-synuclein, inflammatory cytokines (TNF-α, iNOS, IL-1β, and IL-6) in the serum and SN, and the NF-κB expression in the SN, along with significantly increased alpha diversity richness index (P<0.05). In addition, the relative abundance of Bacteroides increased significantly in the MPTP+A (P<0.05), while the relative abundance of Firmicutes and Cyanobacteria decreased significantly (P<0.05). Conclusion: Acupuncture intervention can improve locomotor function, reduce α-synuclein aggregation and inflammatory factors expression, and increase the Akt signaling pathway in PD mice. In addition, acupuncture intervention can benignly regulate the intestinal flora of PD mice. Therefore, it suggests that acupuncture intervention can protect PD model mice probably by regulating intestinal flora and activating Akt signaling pathway.

OBJECTIVETo assess the effect of CatWalk automated gait analysis system for evaluation of motor function of rats with traumatic brain injury (TBI) after umbilical cord mesenchymal stromal cell (UC-MSC) treatment.

METHODSEighteen Wistar rats were randomized equally into normal control group, TBI ∓ saline group, and TBI ∓ UC-MSCs group. The rats in the latter two groups were subjected to weight-drop impact to induce TBI followed by injection UC-MSCs or saline into the lesion 7 days after TBI. The neurological function was assessed using CatWalk system and modified neurological severity scores (mNSS) before and 3 days after TBI and 7 days after UC-MSC transplantation. The rats were sacrificed 14 days after the cell transplantation and the brain sections were stained for immunohistochemical analyses.

RESULTSThree days after TBI, mNSS test showed moderate injury of the rats. Seven days after the cell transplantation, the rats showed significant motor function improvement and CatWalk analysis indicated partial recovery of the gait parameters of the 4 limbs compared to the rats with saline treatment. Histological analyses showed that DiO-labeled UC-MSCs were present in the lesion boundary and expressed glial fibrillary acidic protein and β-tubulin III.

CONCLUSIONUC-MSC transplantation can promote functional improvement of the brain after TBI in rats. Compared with mNSS test, CatWalk analysis is more sensitive and objective for assessing neurological function and also provides more detailed information on specific gait parameters.

Animals ; Brain Injuries ; physiopathology ; surgery ; Disease Models, Animal ; Gait ; Male ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; cytology ; Rats ; Rats, Wistar ; Recovery of Function ; Umbilical Cord ; cytology

Forensic science is looking for clues at a crime scene in order to reconstruct the crime scene.Classic clues include DNA and fingerprints.Forensic microbiology is a branch of forensic medicine that uses microbes as clues,providing us information about lifestyle,circadian rhythms,geographic locations,postmortem intervals,cancers,and oral or systemic diseases.Oral cavity,as the place with the second largest number of microorganisms,can provide researchers with microbial information of each ecological niche,and assist in the prediction,diagnosis and monitoring of oral or systemic diseases.This paper reviews the composition of oral microbiome,the application in oral diseases,systemic diseases and forensic medicine,with the aim of providing some references for the development of forensic microbiology based on oral microbiome.