A diagnostic kit for the enteroviruses Coxsackie A6 and A10.

Recently, there has been an upward trend in the occurrence of hand-foot-mouth disease, which is correlated with Coxsackie A6 and A10 infections. Although two separate diagnostic reagents are available for these two viral strains, the protocol and diagnosis efficiency still need to be improved. More importantly, as co-infection with these viruses is common, the development of a single test kit that can diagnose both viruses would be most beneficial for clinical practice. In our study, specific primers targeting viral nucleic acids were designed and modified. Viral nucleic acids were extracted from fecal or throat swab samples by ultrasonic rupture and silicon membrane purification. The consistency, specificity, and sensitivity of the tests were further optimized by adjusting the polymerase chain reaction (PCR) conditions. The efficiency of viral nucleic acid extraction was significantly enhanced by the ultrasonic rupture and silicon membrane elution approach. Specific amplifications of both viral nucleic acids were achieved using modified primers. The optimal conditions for PCR were also determined (60°C for 30 min and 95°C for 2 min, followed by 40 cycles of denaturation for 30 s at 95°C, annealing for 30 s at 60°C, and elongation for 50 s at 72°C). Amplified products were confirmed as viral specific nucleotides by agarose gel electrophoresis and sequencing. The minimal nucleic acid concentration required for detection was 0.2 ng/L, which was adequate to yield satisfactory specificity and consistency. This novel diagnostic method has many advantages, including rapid protocols and accurate results, and can be promoted for large-scale clinical trials.

[In vitro study of non-thermal atmospheric plasma influencing the surface properties of dentin].

Objective: To study the influence of non-thermal atmospheric pressure plasma (NTAPP) on dentine surface temperature, wettability and morphology of collagen fibrils under different treatment condition. Methods: Helium was used as the operating gas at the flow rate of 3, 4, 5 L/min respectively. The plasma jet was operated at various input power of 8, 9, 10, 11 W. Thermal accumulation on human dentine surface (6 specimens per group, acquired from Department of Stomatology, The First Affiliated Hospital of Xi'an Medical University and Department of Stomatology, The Second Affiliated Hospital of Xi'an Medical University) of each group was measured continuously at 5 s intervals for 60 s by infrared thermography. Mean values were calculated and temperature curves were drawn. Dentine surface contact angles were measured after NTAPP treatment for 5, 10, 15, 20 s with gas flow rate and input power described above. The micro structure of the collagen fibrils of the negative control group (without NTAPP treatment) and NTAPP treatment groups (5 L/min gas flow rate, input power of 8, 9, 10, 11 W and treating time for 5, 10, 15, 20 s) was observed by field emission scanning electron microscopy (FE-SEM). All data were analyzed by SPSS 18.0. Results: Input power, gas flow and treatment time all showed significant influences on dentine surface temperature and wettability (P<0.01). Dentine temperature rose along with time. The greater input power was, the higher dentine temperature was. The greater gas flow rate was, the faster the temperature rose. Dentine surface temperature reached the highest point of (35.10±0.24) ℃ with NTAPP treatment for 60 s, at input power of 11 W and flow rate of 5 L/min. The contact angles of each experimental group decreased with time, and significant differences were found in the contact angles between the experimental groups and the negative control group (75.57°±1.45°). The contact angles showed a decreasing trend as the input power and the gas flow rate increased. The contact angles reached the lowest point of 13.19°±2.01° with NTAPP treatment for 20 s, at input power of 10 W and flow rate of 5 L/min. The FE-SEM results showed that, along with the increase of input power and extension of time, the demineralized collagen fibrils were destroyed in varying degrees. The collagen fibrils were curled, fractured, fused, and even disappeared. Conclusions: NTAPP could significantly increase the surface temperature, modify dentine wettability and alter the micro structure, which was significantly influenced by input power, gas flow rate and treating time.

Controlled-release mitomycin C-polylactic acid film prevents epidural scar hyperplasia after laminectomy by inducing fibroblast autophagy and regulating the expression of miRNAs.

OBJECTIVE: To design a new controlled-release MMC-PLA film and explore whether and how this film could prevent epidural scar hyperplasia and adhesion in a post-laminectomy rat model. MATERIALS AND METHODS: All procedures were performed under the approval and supervision of the Institutional Animal Care and Use Committee (IACUC) of Nanjing Medical University. A total of 120 Sprague-Dawley (SD) rats were randomly placed into four groups after laminectomy (each group=30 rats). In Group I, the laminectomy area was flushed with saline as a control; in Group II, 25 mg of PLA film was applied to the dura mater in the laminectomy area; in Group III, a cotton pad soaked with 0.01% MMC solution was kept on the laminectomy area; and in Group IV, 25 mg of PLA film containing 0.01% MMC was implanted on the laminectomy area. Magnetic resonance imaging (MRI), hematoxylin-eosin (HE) staining and Masson staining were used to evaluate scar adhesion and collagen deposition one month after the operation. Autophagy-related proteins, including autophagy-related gene 5 (ATG5), beclin 1, light chain-3B-2/1 (LC3B-2/1) and protein 53 (p53), were detected by Western blotting. A microRNA microarray analysis was performed to screen for scar tissue miRNAs, especially those associated with autophagy, and changes in expression were confirmed by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: A total of 112 rats recovered uneventfully from the surgery. MRI showed that the scar adhesion and scar area of the MMC-PLA group were significantly reduced compared with those of the PLA, MMC, and saline groups. Accordingly, scar adhesion and the deposition of collagen in the rats treated with MMC-PLA were also significantly reduced, as indicated by HE and Masson staining. In the scar tissue, the levels of autophagy-related proteins (ATG5, beclin 1, LC3B-2/1 and p53) were significantly elevated in the MMC-PLA group. Additionally, in the MMC-PLA group, the expression levels of miR-34a, miR-146a and miR-200 were significantly increased, while the levels of miR-16, miR-221 and miR-378a were significantly decreased. CONCLUSIONS: The controlled-release MMC-PLA film could alleviate epidural scar hyperplasia after laminectomy; this outcome might be associated with increased autophagy and altered expression of miRNAs in the scar tissue.

[Simultaneous determination of o-phthalaldehyde, p-chloro-m-xylenol and triclosan in compound chemical disinfectants and daily chemicals by micellar electrokinetic chromatography].

A novel method for the separation and determination of o-phthalaldehyde (OPA), p-chloro-m-xylenol (PCMX) and triclosan in daily chemicals and compound chemical disinfectants in a single run by micellar electrokinetic chromatography (MEKC) was established. The factors such as the buffer concentration and pH, the concentration of sodium dodecyl sulfate (SDS), and the sample buffer, were investigated in detail. The analysis was carried out using a 50 microm uncoated capillary of 40.2 cm in total length (effective length: 30 cm). The running buffer was 20 mmol/L Na2B4O7, and 80 mmol/L SDS. The sample buffer was 2 mmol/L Na2B4O7-8 mmol/L SDS (without pH adjustment) containing 10% (v/v) methanol. The detection wavelength was 214 nm. The relative standard deviations (RSDs) of the corrected peak areas of the three components were in the range of 1.1% - 3.8%, and the RSDs of migration times were less than 0.9%. The limits of detection (LODs, S/N = 3) were 4.0, 0.4 and 0.4 mg/L, and the limits of quantification (LOQs, S/N = 10) were 12, 1.2, and 1.2 mg/L for OPA, PCMX and triclosan, respectively. The corrected peak areas and the concentrations of the three components showed good linear relationship within the ranges of 12 - 2 000 mg/L, 1.2 - 200 mg/L and 1.2 - 200 mg/L with the correlation coefficients of 0.999 4, 0.999 3 and 0.999 5 for OPA, PCMX and triclosan, respectively. The method was used for the determination of the three components in compound chemical disinfectants, hand washing liquids, soaps and a toothpaste. The results showed that the three components could be assayed in a single run with simple sample pretreatment, rapidity, accuracy and low cost, and the method is convenient for routine analysis.