[Spectrometric analyses of larotaxel and larotaxel liposomes quantification by high performance liquid chromatography].

OBJECTIVE: Larotaxel is a new chemical structure drug, which has not been marketed worldwide. Accordingly, the standard identification and quantification methods for larotaxel remain unclear. The spectrometric analyses were performed for verifying weight molecular formula, molecular weight and chemical structure of larotaxel. Besides, a quantification method was developed for measuring larotaxel in the liposomes. METHODS: The molecular formula, molecular weight and chemical structure of larotaxel were studied by using mass spectrometry (MS), infra-red (IR), nuclear magnetic resonance (NMR) and ultraviolet-visible (UV-vis) spectrometric techniques. The absorption wavelength of larotaxel was investigated by UV-vis spectrophotometry full-wavelength scanning. Besides, a quantification method was developed by high performance liquid chromatography (HPLC), and then validated by measuring the encapsulation efficacy of larotaxel liposomes. RESULTS: The four spectral characteristics of larotaxel were revealed and the corresponding standard spectra were defined. It was confirmed that larotaxel had the structure of tricyclic diterpenoids, with the molecular formula of C45H53NO14, the molecular weight of 831.900 1, and the maximum absorption wavelength of 230 nm. The quantitative method of larotaxel was established by using HPLC with a reversed phase C18 column (5 µm, 250 mm×4.6 mm), a mobile phase of acetonitrile-water (75:25, volume/volume), and a detection wavelength of 230 nm. The validation study exhibited that the established HPLC method was stable, and had a high recovery and precision in the quantitative measurement of larotaxel in liposomes. In addition, a new kind of larotaxel liposomes was also successfully prepared. The particle size of the liposomes was about 105 nm, with an even size distribution. And the encapsulation efficiency of larotaxel in the liposomes was above 80%. CONCLUSION: The present study offers reference standard spectra of larotaxel, including MS, IR, NMR, and UV-vis, and confirms the molecular formula, molecular weight and chemical structure of larotaxel. Besides, the study develops a rapid HPLC method for quality control of larotaxel liposomes.

[Etiology and clinical analysis of central nervous system infection caused by Coxsackievirus B5 in severe hand, foot and mouth disease in Qingdao City, 2013-2014].

Objective: To illuminate the gene characteristics and clinical characterization of Coxsackievirus B5 (CV-B5) strains isolated from patients with sevre hand, foot and mouth disease (HFMD) in Qingdao city. Methods: A total of 1 844 patients of HFMD were consecutively admitted to Qingdao Women and Children's Hospital from 2013 to 2014. Information of the study population described above was collected retrospectively. The samples were collected from at least 1 site (throat swab, cerebrospinal fluid), which viral nucleic acid extracted and the entire VP1 gene sequences of CV-B5 isolates were amplified and sequenced, then the homology and phylogeny analysis were conducted by MEGA7.0. The prototype Faulkner strain and other VP1 amino acid sequences were derived from the GenBank database. Results: A total of 8 CV-B5 positive cases were obtained, including 4 males and 4 females; 6 severe hospitalized cases and 2 outpatients. The age of 6 hospitalized patients ranged from 3 to 48 months, with a median of 26 months. For the six inpatients, fever, convulsions vomiting, diarrhea and rash were the main clinical manifestation, and all combined with viral encephalitis. Compared with the prototype strain Faulkner, in the VP1 region,the nucleotide and the amino acid homologies was 77.3%-78.8% and 95.5%-97.0% respectively. Five out of the six severe cases with substitution of serine (S) to asparagine (N) at amino acid site 95 in the VP1 region. The sequences of 8 CV-B5 strains were classified into genogroup D. Conclusion: Hand, foot and mouth disease associated with CV-B5 virus infection can result in nervous system involvement and the main complication was viral encephalitis. The CV-B5 strains associated with severe hand, foot and mouth disease had high nucleotide homology and present a certain regional aggregation.

[Effects of different techniques on removal of vapor lock in the apical region of curved canals: a cone-beam computed tomography study].

OBJECTIVE: To compare the effect of four different techniques on removal of vapor lock in the apical region of curved root canals. METHODS: Forty simulated resin root canals with 45° curvature were prepared using WaveOne Primary, then the apical foramen were sealed with soft wax. The teeth were divided randomly into 4 groups thereafter (n=10). Contract solution was injected into the canals using a 30 G side-vented needle and scanned with cone-beam CT (CBCT) to identify the volume of the vapor lock. Four different techniques including photon-induced photoacoustic streaming (PIPS) laser-activated irrigation, gutta-percha cone technique, ultrasonic irrigation, and sonic irrigation were used to remove the vapor locks in the root canals. The residual volume of the vapor lock was identified again using CBCT scanning data. Accordingly, the reduction rates of the vapor lock were calculated. Furthermore, the initial and residual vapor lock length was calculated. The data were analyzed by using the One-way ANOVA analysis and Kruskal-Wallis H test at a significance level of P<0.05. RESULTS: There was no significant difference in the initial vapor lock volume (P>0.05). Residual volume of the vapor lock for PIPS laser-activated irrigation was 0 mm(3), and that for gutta-percha cone technique was (0.02±0.07) mm3, significantly lower than those of ultrasonic and sonic irrigation, the values being (0.20±0.09) mm(3) and (0.23±0.06) mm(3) (P<0.001), respectively. The reduction rates of the vapor lock of PIPS laser-activated irrigation and gutta-percha cone technique were 100.00% (100.00%, 100.00%) and 100.00% (77.66%, 100.00%), respectively, significantly higher than those of ultrasonic irrigation [70.37% (56.41%, 91.43%)] and sonic irrigation [63.54% (51.47%, 74.00%), P<0.001]. The length of the residual vapor lock for PIPS laser-activated irrigation was 0 mm, and that for gutta-percha cone technique was (0.15±0.47) mm, significantly lower than those of ultrasonic and sonic irrigation, values being (2.21±0.09) mm and (2.34±0.08) mm (P<0.001), respectively. The length of the residual vapor locks in the ultrasonic and sonic group remained approximately the same as the distance between the working tip and the apical foramen. CONCLUSION: PIPS laser activated irrigation and gutta-percha cone technique could remove the vapor lock from the apical region of curved canals effectively.

Porphyromonas gingivalis B cell Antigen Epitope Vaccine, pIRES-ragB'-mGITRL, Promoted RagB-Specific Antibody Production and Tfh Cells Expansion.

The outer membrane protein RagB is one of the major virulence factors of Porphyromonas gingivalis (P. gingivalis). To prevent periodontitis and associated systemic diseases induced by P. gingivalis, we built B cell antigen epitope vaccine characterized by pIRES-ragB'-mGITRL to induce a protective immune responses. The B cell antigen epitope and scrambled peptide of ragB were predicted, cloned into pIRES and constructed pIRES-ragB', pIRES-scrambled epitopes and pIRES-ragB'-mGITRL. pIRES-ragB'-mGITRL was transfected into COS-7 cells. Subsequently, the 6-week-old female BALB/c mice were challenged by P. gingivalis following three time immunization by pIRES, pIRES-ragB', pIRES-scrambled epitopes and pIRES-ragB'-mGITRL. The levels of RagB-specific antibody in the serum and Tfh cells in the spleen were measured by ELISA and flow cytometry, respectively. And higher levels of RagB-specific IgG were produced in the immunized mice with pIRES-ragB'-mGITRL. Additionally, the number of Tfh cells was also expanded and lesions were diminished in pIRES-ragB'-mGITRL mice comparing with control groups. Our results clearly demonstrated that P. gingivalis B cell antigen epitope vaccine, pIRES-ragB'-mGITRL, could induce protective immune responses. Furthermore, our data also indicated that pIRES-ragB'-mGITRL was a potential therapeutic vaccine against P. gingivalis.

Directional atherectomy combined with drug-coated balloon angioplasty for superficial femoral arteriosclerosis obliterans.

INTRODUCTION: This study is an analysis of the therapeutic effects of directional atherectomy combined with drug-coated balloon angioplasty (DA+DCB) in treating superficial femoral arteriosclerosis obliterans. METHODS: Patients in our hospital with superficial femoral arteriosclerosis obliterans who received DA+DCB during the period June 2016 to February 2019 were identified retrospectively. Preoperative demographics, operative details and postoperative follow-up outcomes were analysed statistically. RESULTS: Between June 2016 and February 2019, 48 patients were enrolled in this retrospective study. The average age of the patients was 66.85 ± 11.28 years; 83.3% of the patients were male. During the procedure, flow-limiting dissection occurred frequently (9/48 patients) and there were six bailout stent implantations owing to flow-limiting dissections. The incidence rate of target artery thrombosis was 4.2% (2/48). There was no vessel perforation, embolism or operation-related death. The technical success rate was estimated at 100%. The mean ankle-brachial index of the patients was 0.54 ± 0.28 before the operation and 0.93 ± 0.13 before discharge (p < 0.0001). The mean follow-up time was 19.6 ± 9.0 months. The primary patency rate was 89.4%, 82.4% and 76.5% at 12, 24 and 36 months. The freedom from target lesion revascularisation (TLR) was 97.9%, 93.8% and 84.4% at 12, 24 and 36 months. CONCLUSION: The use of DA+DCB showed good clinical benefit for superficial femoral arteriosclerosis obliterans, which had good primary patency and freedom from TLR. Multicentre randomised controlled trials with long-term follow-up are needed.

Excimer laser atherectomy combined with drug-coated balloon angioplasty for the treatment of femoropopliteal arteriosclerosis obliterans.

INTRODUCTION: It has been reported that excimer laser atherectomy combined with a drug-coated balloon (ELA+DCB) can achieve better results than simple balloon angioplasty, especially for the treatment of femoropopliteal in-stent restenosis. However, reports on the application of ELA+DCB in China for femoropopliteal arteriosclerosis obliterans are lacking. This study focuses on analysing the effectiveness and safety of ELA+DCB. METHODS: This was a single-centre retrospective study that enrolled patients from November 2016 to January 2019 who had femoropopliteal arteriosclerosis obliterans treated by ELA+DCB. Preoperative demographics, operative details and postoperative follow-up outcomes were analysed statistically. RESULTS: There were 43 patients with an average patient age of 68.0±8.6 years; 79.1% were male. In 30 cases, the lesions were de novo and the others were in-stent restenosis (ISR). During the procedure, flow-limiting dissection (48.8%) was the main adverse event and there were 17 bailout stent implantations due to dissection. Mean (±sd) ankle-brachial index (ABI) in the patients was 0.42±0.31 before the operation and 0.83±0.13 before discharge. The mean (±sd) follow-up time was 29.35±9.71 months. The primary patency rate was 66.8%, 64.3% and 60.9% at 12, 24 and 36 months. Freedom from target lesion revascularisation (TLR) was 85.7%, 80.7% and 75.3% at 12, 24 and 36 months. Rutherford categories also greatly improved during follow-up. Overall mortality was 6.9% (3/48), and no deaths were related to the intervention. CONCLUSION: The use of ELA+DCB had good clinical benefit for femoropopliteal arteriosclerosis obliterans, which had good primary patency and freedom from TLR, although intraoperative complications still required attention. Multicentre randomised controlled trials with long-term follow-up are needed.

The rag locus of Porphyromonas gingivalis might arise from Bacteroides via horizontal gene transfer.

Porphyromonas gingivalis is regarded as one of the risk factors of periodontitis. P. gingivalis exhibits a wide variety of genotypes. Many insertion sequences (ISs), located in their chromosomes, made P. gingivalis differentiate into virulent and avirulent strains. In this research, we investigated the prevalence of P. gingivalis in the gingival crevicular fluid (GCF) among periodontitis patients from Zhenjiang, China, detected the P. gingivalis rag locus distributions by multiplex polymerase chain reaction (PCR), and analyzed the origin of the P. gingivalis rag locus based on evolution. There were three rag locus variants co-existing in Zhenjiang. The results showed that the rag locus may be associated with severe periodontitis. This work also firstly ascertained that the rag locus might arise, in theory, from Bacteroides sp. via horizontal gene transfer.

Chronic osteomyelitis with proliferative periostitis of the mandibular body: report of a case and review of the literature.

Chronic osteomyelitis with proliferative periostitis is a rare form of osteomyelitis that is characterised by new bone formation with periosteal reaction. It is also traditionally known as Garre's osteomyelitis. The common sources of infection of the jaw include dental caries associated with periapical periodontitis, periodontitis, fractures and nonodontogenic infections. Chronic osteomyelitis with proliferative periostitis mainly presents in younger patients. Here, we present a case of a 12-year-old patient with chronic osteomyelitis with proliferative periostitis with no definitive infection source such as pericoronitis, caries and periodontitis. Therapeutic measures involved surgical debridement and antibiotics. Disease remission and a normal facial symmetrical morphology were observed at the six-month follow-up.

Layer-by-layer assembly of viral capsid for cell adhesion.

Cowpea mosaic virus (CPMV)-based thin films are biologically active for cell culture. Using layer-by-layer assembly of CPMV and poly(diallyldimethylammonium chloride), quantitatively scalable biomolecular surfaces were constructed, which were well characterized using quartz crystal microbalance, UV-vis and atomic force microscopy. The surface coverage of CPMV nanoparticles depended on the adsorption time and pH of the virus solution, with a greater amount of CPMV adsorption occurring near its isoelectric point. It was found that the adhesion and proliferation of NIH-3T3 fibroblasts can be controlled by the coverage of viral particles using this multilayer technique.

Grafting materials for alveolar cleft reconstruction: a systematic review and best-evidence synthesis.

The purpose of this study was to compare the efficacy of alveolar bone reconstruction for alveolar cleft patients performed with the traditional iliac graft or alternative/supplementary bone grafting materials. Electronic databases, relevant journals, and reference lists of the included studies were searched to the end of June 2016. A best-evidence synthesis was performed to draw conclusions. A total of 38 studies were included, which provided 25 pieces of evidence: seven of moderate evidence and 18 of insufficient evidence. The seven pieces of moderate evidence indicated that (1) bone morphogenetic protein 2 bound to absorbable collagen sponge shares similar cleft repair efficacy to the iliac graft; (2) covering the iliac graft with an acellular dermis matrix membrane may increase bone retention for unilateral cleft patients; (3) mixing iliac graft with platelet-rich plasma may increase bone retention for skeletally mature patients, but (4) does not achieve the same result for younger patients; and compared with the iliac graft, (5) the mandible graft is more effective, whereas (6) the cranium graft and (7) rib graft are less effective for alveolar cleft reconstruction. The efficacy of the remaining grafting materials was supported by insufficient evidence. More well-designed controlled studies are needed to ascertain the long-term clinical results of alveolar cleft reconstruction.

Effects of bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) release from polylactide-poly (ethylene glycol)-polylactide (PELA) microcapsule-based scaffolds on bone.

Multiple growth factors can be administered to mimic the natural process of bone healing in bone tissue engineering. We investigated the effects of sequential release of bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) from polylactide-poly (ethylene glycol)-polylactide (PELA) microcapsule-based scaffolds on bone regeneration. To improve the double emulsion/solvent evaporation technique, VEGF was encapsulated in PELA microcapsules, to which BMP-2 was attached. The scaffold (BMP-2/PELA/VEGF) was then fused to these microcapsules using the dichloromethane vapor method. The bioactivity of the released BMP-2 and VEGF was then quantified in rat mesenchymal stem cells (rMSCs). Immunoblotting analysis showed that BMP-2/PELA/VEG promoted the differentiation of rMSCs into osteoblasts via the MAPK and Wnt pathways. Osteoblast differentiation was assessed through alkaline phosphatase expression. When compared with simple BMP-2 plus VEGF group and pure PELA group, osteoblast differentiation in BMP-2/PELA/VEGF group significantly increased. An MTT assay indicated that BMP-2-loaded PELA scaffolds had no adverse effects on cell activity. BMP-2/PELA/VEG promoted the differentiation of rMSCs into osteoblast via the ERK1/2 and Wnt pathways. Our findings indicate that the sequential release of BMP-2 and VEGF from PELA microcapsule-based scaffolds is a promising approach for the treatment of bone defects.

One-stage repair of absence of the aortopulmonary septum and interrupted aortic arch.

Absence of the aortopulmonary septum, interrupted aortic arch, aortic origin of the right pulmonary artery, intact ventricular septum, and patent ductus arteriosus is a rarely reported association. A 3-year-old boy underwent successful one-stage repair of this constellation of anomalies. A Dacron baffle was used both to close the huge aortopulmonary window and to direct blood to the right pulmonary artery. Type A interrupted aortic arch was repaired by direct anastomosis. Postoperatively, pulmonary artery pressure was less than half systemic pressure.

Characterization of polyethylene glycol-modified proteins by semi-aqueous capillary electrophoresis.

A new program to characterize polyethylene glycol-modified (PEGylated) proteins is outlined using capillary zone electrophoresis (CZE). PEGylated ribonuclease A and lysozyme were selected as examples. Five separation procedures were compared to select out the mixed buffer of acetonitrile-water (1:1, v/v) at pH 2.5 as the best to characterize the PEGylated proteins without sample pretreatment. Polyethylene oxide (PEO) with a high molecular mass of 8 x 10(6) was applied to rinse the capillary to form a dynamic coating which would decrease the undesirable proteins adsorbed to the inner wall of the silica. The electroosmotic flow (EOF) mobility of the five procedures was determined, respectively. It is found that acetonitrile is mainly responsible for the good resolution of PEGylated proteins with the help of PEO coating in the semi-aqueous system. The low EOF mobility and current in the semi-aqueous system might also have some responsibility for the high resolution. The semi-aqueous procedure described in this paper also demonstrates higher resolution of natural proteins than aqueous ones.

The design and application of a pediatric centrifugal pump.

This centrifugal pump (CP) includes two parts: the blood pump and the driving apparatus. They are connected by six twin magnetic disc plates and driven by a magnetic DC motor (120W). The blood pump had six leaves deadlocked between two plastic discs. Six leaves were set at 30 degrees angles, separately. In the lower chamber of the CP, there was an inlay magnetic disc, which is connected with the disc leaves by an axis. This axis was sealed by silicon rubber and a ceramic ring. The priming volume of the blood chamber was 34 ml. In vitro testing showed that the free hemoglobin caused by the CP was much less than that caused by a roller pump after 180 min. The effect of this CP on blood cell damage was also studied in an animal model. Six goats were placed on cardiopulmonary bypass for 180 min. Perfusion flow rates were maintained between 1.5 and 2.5 L/min. The plasma free hemoglobin was lower in the CP group (6.04 mg/dL) than in the roller pump group (32.25 mg/dL), p < 0.01. The CP has been used in ten pediatric patients undergoing cardiopulmonary bypass surgery. The patients' ages were from three to five years, and body weights were from 15 to 20 kg. Perfusion flow rates were maintained between 1.8 and 2.5 L/min, and bypass times were from 30 to 50 min. The rotation speeds were from 2000 to 2500 rpm. All the patients recovered smoothly, and no hemoglobinuria occurred.

Effects of bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) release from polylactide-poly (ethylene glycol)-polylactide (PELA) microcapsule-based scaffolds on bone

Multiple growth factors can be administered to mimic the natural process of bone healing in bone tissue engineering. We investigated the effects of sequential release of bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) from polylactide-poly (ethylene glycol)-polylactide (PELA) microcapsule-based scaffolds on bone regeneration. To improve the double emulsion/solvent evaporation technique, VEGF was encapsulated in PELA microcapsules, to which BMP-2 was attached. The scaffold (BMP-2/PELA/VEGF) was then fused to these microcapsules using the dichloromethane vapor method. The bioactivity of the released BMP-2 and VEGF was then quantified in rat mesenchymal stem cells (rMSCs). Immunoblotting analysis showed that BMP-2/PELA/VEG promoted the differentiation of rMSCs into osteoblasts via the MAPK and Wnt pathways. Osteoblast differentiation was assessed through alkaline phosphatase expression. When compared with simple BMP-2 plus VEGF group and pure PELA group, osteoblast differentiation in BMP-2/PELA/VEGF group significantly increased. An MTT assay indicated that BMP-2-loaded PELA scaffolds had no adverse effects on cell activity. BMP-2/PELA/VEG promoted the differentiation of rMSCs into osteoblast via the ERK1/2 and Wnt pathways. Our findings indicate that the sequential release of BMP-2 and VEGF from PELA microcapsule-based scaffolds is a promising approach for the treatment of bone defects.

Direct response to proton beam linear energy transfer (LET) in a novel polymer gel dosimeter formulation.

Linear energy transfer (LET) of clinical proton beams is an important parameter influencing the biological effects of radiation. This work demonstrates LET-induced response enhancement in novel formulations of polymer gel dosimeters, potentially useful for LET mapping of clinical proton beams. A series of four polymer gel dosimeters (labeled A through D), prepared based on the BANG3-Pro2 formulation, but with varying concentrations of polymerization modifiers, were irradiated by a clinical proton beam with a spread out Bragg peak modulation (SOBP) and read out using the OCTOPUS-IQ optical CT scanner. The evaluation of optical density profiles in the SOBP (constant physical dose) revealed response deviations at the distal end consistent with variations in gel composition. Maximum response deviations were as follows: -3% (under-response) for gel A, and over-response of 2%, 12%, and 17% for gels B, C, and D, respectively, relative to the mean dose in the center of the SOBP. This enhancement in optical response was correlated to LET by analytical calculations. Gels A and B showed no measurable dependence on LET. Gel C responded linearly in the limited range from 1.5 to 3.5 keV/µm. LET response of gel D was linear up to at least 5.5 keV/µm, with the threshold at about 1.3 keV/µm. These results suggest that it may be possible to develop a polymer gel system with direct optical response to LET for mapping of LET distributions for particle therapy beams.

[Removal of PEG and salt from recombinant human tumor necrosis factor-alpha(rhTNF-alpha) with serial operation of gel filtration chromatography].

A serial gel filtration operation with the combination of G25 and G100 in one column was developed to remove salt and PEG from upper phase rich in recombinant human tumor necrosis factor alpha after simultaneous cell disruption and two-phase aqueous extraction. Buffer exchanging and primary purification were also achieved at same time. The purification factor of 4.4 was obtained with an activity recovery of 97% by one single step of serial operation.