Objective:To observe whether endothelial cells undergo pyroptosis in the inflammatory periodontal environment by using a model in vivo and in vitro, providing an experimental basis for indepth understanding of the underlying pathogenesis of periodontitis. Methods:According to the classification of periodontal diseases of 2018, gingival tissues were collected from periodontally healthy subjects and patients with stage Ⅲ-Ⅳ, grade C periodontitis, who presented Department of Oral and Maxillofacial Surgery and Department of Periodontology, School of Stomatology, The Fourth Military Medical University from April to May 2022. Immunohistochemical staining was performed to detect the expression level and distribution of gasdermin D (GSDMD), a hallmark protein of cell pyroptosis, in gingival tissues. Periodontitis models were established in each group by ligating the maxillary second molar teeth of three mice for 2 weeks (ligation group). The alveolar bone resorption was determined by micro-CT (mice without ligation treatment were used as the control group), and the colocalization of GSDMD and CD31 were quantitatively analyzed by immunofluorescence staining in gingival tissues of healthy and inflammatory mice. Human umbilical vein endothelial cells (HUVECs) were cultured in vitro and treated with lipopolysaccharide (LPS) of Porphyromonas gingivalis (Pg) combined with adenosine triphosphate (ATP) at various concentrations of 0.5, 1.0, 2.5, 5.0, and 10.0 mg/L, respectively, and the 0 mg/L group was set as the control group at the same time. Scanning electron microscopy was used to observe the morphology of HUVECs. Western blotting was used to detect the expression of gasdermin D-N terminal domains (GSDMD-N) protein and immunofluorescence cell staining was used to detect the expression and distribution of GSDMD. Cell counting kit-8 (CCK-8) was used to detect the proliferative ability of HUVECs, and propidium iodide (PI) staining was used to detect the integrity of cell membrane of HUVECs. Results:Immunohistochemistry showed that GSDMD in gingival tissues of periodontitis was mainly distributed around blood vessels and its expression level was higher than that in healthy tissues. Micro-CT showed that alveolar bone resorption around the maxillary second molar significantly increased in ligation group mice compared with control subjects ( t=8.88, P<0.001). Immunofluorescence staining showed significant colocalization of GSDMD with CD31 in the gingival vascular endothelial cells in mice of ligation group. The results of scanning electron microscopy showed that there were pores of different sizes, the typical morphology of pyroptosis, on HUVECs cell membranes in the inflammatory environment simulated by ATP combined with different concentrations of LPS, and 2.5 mg/L group showed the most dilated and fused pores on cell membranes, with the cells tended to lyse and die. Western blotting showed that the expression of GSDMD-N, the hallmark protein of cell pyroptosis, was significantly higher in 2.5 and 5.0 mg/L groups than that in the control group ( F=3.86, P<0.01). Immunofluorescence cell staining showed that the average fluorescence intensity of GSDMD in 2.5 mg/L group elevated the most significantly in comparison with that in the control group ( F=35.25, P<0.001). The CCK-8 proliferation assay showed that compared to the control group (1.00±0.02), 0.5 mg/L (0.52±0.07), 1.0 mg/L (0.57±0.10), 2.5 mg/L (0.58±0.04), 5.0 mg/L (0.55±0.04), 10.0 mg/L (0.61±0.03) groups inhibited cell proliferation ( F=39.95, P<0.001). PI staining showed that the proportion of positive stained cells was highest [(56.07±3.22)%] in 2.5 mg/L group ( F=88.24, P<0.001). Conclusions:Endothelial cells undergo significant pyroptosis in both in vivo and in vitro periodontal inflammatory environments, suggesting that endothelial cell pyroptosis may be an important pathogenic factor contributing to the pathogenesis of periodontitis.

Objective: To analyze the safety and effectiveness of different types of stents with the treatment of vertebraebasilar artery dissection aneurysms(VBADA). Methods : The clinical data of 80 patients with VBADA treated by intravascular intervention in the First Hospital of University of Science and Technology of China(Anhui Provincial Hospital) from February 2018 to November 2023 were retrospectively analyzed. Patients were divided into laser engraved stent group(n=34) and braided stent group(n=46) based on the type of stent used. O′Kelly-Marotta(OKM) grade was used to evaluate the embolization effect of aneurysms in DSA images, and a modified Rankin Scale(mRS) score was used to evaluate the clinical prognosis of patients. The baseline data, aneurysm characteristics, intraoperative and perioperative treatment details and postoperative follow-up details between the two groups were compared. Results : There was no significant difference in baseline data, mRS score, ischemic and hemorrhage complications and mortality between the two groups(allP>0.05). Compared with the laser engraved stent group, the mean diameter of aneurysms was larger(P<0.000 1) and the proportion of ruptured aneurysm(P<0.01), parent artery stenosis and beaded vascular lesions(P<0.05) and branch artery dissecting aneurysm were higher(P<0.01) in the braided stent group. Conversely, the proportion of coil-assisted embolization(P<0.000 1) and the immediate aneurysm occlusion rate(OKM C and D)(P<0.000 1) were lower. Finally, 21 patients were obtained by controlling for maximum diameter of aneurysms and whether coil-assisted embolization, the aneurysm occlusion rate half a year later in the braided stent group was higher than that in the laser engraved stent group(P<0.05), but the recurrence rate of postoperative aneurysm was lower than that of laser engraved stent group(P<0.05). It was worth noting that the cure rate and vascular remodeling rate of middle-large size VBADA which the maximum diameter being over 15 mm in the braided stent group reached 72.2%, and the whole effect was ideal. Conclusion Braided stents are relatively safe and effective in the treatment of VBADA and are significantly better than laser engraved stents in reducing VBADA recurrence and remodeling lesion vessels without increasing postoperative complication.

Objective To analyze the differences in the expression of specific antibodies targeting different human immunodeficiency virus (HIV) antigens among patients at different clinical stages in Qiandongnan Prefecture of Guizhou Province, and to explore their association with the clinical staging of acquired immunodeficiency syndrome. Methods A total of 307 HIV-positive blood samples from Qiandongnan of Guizhou Province were selected for specific HIV antibody immunoblotting assays. CD3+CD4+T cell counts andHIV viral load nucleic acid testing were performed on the blood samples. Multivariate regression analysis was conducted on relevant indicators during HIV infection and AIDS stages. Results Among the 307 HIV-infected individuals, 218 were male and 89 were female, with a mean age of (48.53±16.03) years. The composition ratios of specific antibodies gp160, gp120, gp41, p66, p51, p31, p55, p24 and p17 were 98.7%, 90.9%, 92.2%, 74.3%, 66.4%, 86.6%, 3.9%, 97.4% and 73.9%, respectively. Multivariate binary Logistic regression model analysis showed that expression of p24-specific antibodies were more likely to be judged as influencing factors in the infection stage (OR=0.158, 95%CI, 0.032 to 0.768, P < 0.05). There was a certain correlation between p24-specific antibodies and the clinical staging of HIV infection (OR=0.217, 95%CI, 0.005 to 0.944, P < 0.05). Using the ratio of CD3⁺ CD4⁺ T cell count to viral load when p24 was specifically expressed as the test variable, and the clinical stage as the state variable (AIDS stage=1, infection stage=2), a receiver operating characteristic (ROC) curve was plotted. The area under the curve was 0.653 (95%CI, 0.529 to 0.774, P < 0.05). Conclusion Differential expression of p24-specific antibodies may indicate that patients are in the infection stage and could potentially serve as an early warning indicator of immune function for HIV-infected individuals.

Cell-free transcription and translation (TXTL) system is a cell extract-based system for rapid in vitro protein expression. The system bypasses routine laboratory processes such as bacterial transformation, clonal screening and cell lysis, which allows more precise and convenient control of reaction substrates, reduces the impact of bacteria on protein production, and provides a high degree of versatility and flexibility. In recent years, TXTL has been widely used as an emerging platform in clusterd regularly interspaced short palindromic repeat (CRISPR) technologies, enabling more rapid and convenient characterization of CRISPR/Cas systems, including screening highly specific gRNAs as well as anti-CRISPR proteins. Furthermore, TXTL-based CRISPR biosensors combined with biological materials and gene circuits are able to detect pathogens through validation of related antibiotics and nucleic acid-based markers, respectively. The reagents can be freeze-dried to improve portability and achieve point-of-care testing with high sensitivity. In addition, combinations of the sensor with programmable circuit elements and other technologies provide a non-biological alternative to whole-cell biosensors, which can improve biosafety and accelerate its application for approval. Here, this review discusses the TXTL-based characterization of CRISPR and their applications in biosensors, to facilitate the development of TXTL-based CRISPR/Cas systems in biosensors.
CRISPR-Cas Systems ; Bacteria

Objective : To investigate the effect of different decontamination methods, including photodynamic therapy, sandblasting and titanium curette, on titanium surface morphology and bacterial adhesion for the treatment of peri-implant disease. Methods: Porphyromonas gingivalis (Pg) and Fusobacterium nucleatum (Fn) were inoculated on the surface of polished titanium specimens, and titanium specimen surfaces were treated with different decontamination methods after incubation. The titanium specimens were divided into a no-treatment control group, photodynamic group, sandblasting group and titanium curette group according to different decontamination methods. The changes in titanium surface roughness were observed by atomic force microscopy (AFM), and the remaining bacteria on the titanium surface were observed by scanning electron microscopy (SEM) and live/dead bacteria staining tests. After reinoculation of Pg and Fn, bacterial readhesion was observed on the surface of decontaminated titanium specimens. Results : The AFM results showed that the surface roughness of the titanium curette group was significantly higher than that of the no-treatment control group, photodynamic group and sandblasting group (P<0.05), and there was no statistically significant difference between the no-treatment control group, photodynamic group and sandblasting group (P>0.05). The results of contact angle measurement showed that the surface contact angle of each treatment group was smaller than that of the no-treatment control group (P<0.05). The SEM results obtained after the titanium specimen surface was decontaminated showed that the number of bacteria on the no-treatment control group surface was higher and the bacteria were relatively concentrated. The bacteria on the surface of the photodynamic group, sandblasting group and titanium curette group were scattered and distributed in small numbers, and most bacteria on the surface of the photodynamic group were ruptured. The results of the live/dead bacteria staining experiment showed that the percentage of dead bacteria on the surface of the photodynamic group was significantly higher than that of the no-treatment control group, sandblasting group and titanium curette group (P<0.05). The remaining bacteria on the surface of the sandblasting group and titanium curette groups were mainly live bacteria. The remaining bacterial adhesion on the surface was significantly reduced for the sandblasting group compared to the no-treatment control group and the photodynamic and titanium curette groups (P<0.05). SEM and live/dead bacteria staining results of bacterial readhesion on the surface of titanium specimens showed that there was an aggregation of Pg on the surface of the titanium curette group, and its surface bacterial adhesion was significantly higher than that of the no-treatment control group, photodynamic group and sandblasting group. Conclusion In mechanical decontamination, sandblasting machines are a better option than photodynamic therapy and titanium curettes; however, sandblasting does not remove all bacterial contamination. For sterilization, photodynamic therapy is more effective than sandblasting and titanium curettes. A combination of sandblasting and photodynamic therapy methods for the treatment of peri-implant disease may be considered in clinical practice.

OBJECTIVE To establish the index system of quality evaluation standard for pharmacist training in cough and wheeze pharmaceutical care （CWPC） outpatient department， and to provide a basis for the selection of CWPC pharmacist training teachers and the improvement of training plans. METHODS Based on Kirkpatrick model， using the Delphi method， a total of 15 experts from 13 tertiary hospitals in 10 provinces in China were consulted to establish the standard index system of the quality evaluation for CWPC pharmacists training. Analytic hierarchy process （AHP） was adopted to determine the weights of each indicator and quantify the index system according to the weights of indicators at all levels. RESULTS The coefficient of expert authority was 0.810， the judgment coefficient was 0.727， and the familiarity coefficient was 0.893. The Kendall coordination coefficient of each index was 0.308-0.687. The P values of χ2 test were all less than 0.05， which indicated that the degree of coordination of the experts was high. After two rounds of correspondence， Kirkpatrick model-based index system of quality evaluation standard for CWPC pharmacist training was determined. The index system included 4 first-level indexes （participants’ reaction layer， learning gain layer， behavior improvement layer， training outcome layer）， 12 second-level indexes （such as training needs， teaching methods， theoretical knowledge， practical skills， job abilities， patient benefits， etc.） and 44 third-level indexes （such as clear training objectives， core system of CWPC， special device operation and evaluation， professional knowledge related to the treatment of cough and wheeze patients， promoting the construction of CWPC， improving patient compliance）. CONCLUSIONS The constructed index system of quality evaluation standard for CWPC pharmacist training has a certain level of authority and scientificity， and provides a scientific theoretical basis for quality evaluation of CWPC pharmacist training.

Objective:To explore the asymmetry of neonatal brain white matter using fractional anisotropy (FA) parameter generated from diffusion tensor imaging (DTI).Methods:From January 2011 to June 2013, 88 neonates with normal cranial MRI performance, aged 1-14 (7.7±0.3) days and gestational age 31-42 (37.5±0.3) weeks, were retrospectively collected from the First Affiliated Hospital of Xi′an Jiaotong University. According to the gestational age at birth, the neonates were divided into preterm group (37 cases) and term group (51 cases). DTI was processed to generate FA parametric maps. Tract-based spatial statistics (TBSS) was used to analyze the asymmetry in neonatal white matter. Regions of interest were placed in bilaterally symmetrical white matter tracts. The white matter tracts included the bilateral anterior thalamic radiation, corticospinal tract-posterior limb of the inner capsule (CST-IC), corticospinal tract-corona radiata, cingulum hippocampus part (CGH), inferior frontal occipital fasciculus (IFOF), inferior longitudinal fasciculus (ILF), superior longitudinal fasciculus (SLF), uncinate fasciculus, superior longitudinal fasciculus temporal part (SLF-temp). The FA value was measured and asymmetry index (AI) was calculated. The independent-sample t test or Mann-Whitney U test was used to compare the AI of each tract between the preterm and term groups. Partial correlation was used to analyze the effects of neonatal gestational age and birth anthropometric measures on the asymmetry of neonate at birth. Results:The results of TBSS showed that the left lateralized areas of brain white matter in the neonatal period were mainly located in the corticospinal tract, external capsule and genu of corpus callosum. The right lateralized areas of brain white matter in the neonatal period were mainly located in the optic radiation and splenium of corpus callosum. In both the preterm and term groups, the tracts related to motor (CST-IC) and language function (ILF, SLF, SLF-temp) were left asymmetry. The CGH of the preterm and the term neonates were right asymmetry, and AI was -0.086±0.114 and -0.140±0.108 respectively, with a statistically significant difference ( t=2.27, P=0.026), while the differences of AI in the remaining tracts were not statistically significant ( P>0.05). Only gestational age was correlated with the AI of ILF ( r=0.234, P=0.033), SLF ( r=0.259, P=0.018), SLF-temp ( r=0.252, P=0.022), CST-IC ( r=0.235, P=0.033). No significant correlation was found between birthweight, head circumference, body length and AI. Conclusions:The asymmetries of brain white matter already exist in neonates aged less than two weeks. The tracts associated with motor and language function are predominantly left asymmetry.

Due to the special structure of the nasal cavity, the addition of functional excipients in nasal drugs can improve the transmembrane absorption of the active pharmaceutical ingredient or reduce the clearance of nasal mucocilia, which plays an important role in maintaining the effective concentration of active pharmaceutical ingredient in the target area and promoting the absorption of nasal mucosa. In this paper, the excipients used in drug new approved by the United States Food and Drug Administration(FDA) in the past decade were summarized, and the current research advances and safety evaluation of excipients for nasal drug were reviewed, providing ideas and references for the selection of excipients for nasal drug development.

Objective:To explore the risk factors associated with tibia fractures in children with congenital anterolateral bowing of the tibia (ALBT).Methods:A retrospective analysis was conducted on data from 87 children diagnosed with ALBT at the Children's Hospital of Hunan Province from January 2012 to January 2020. The collected data included age at initial diagnosis, affected limb side, whether there was a concomitant type I neurofibromatosis, whether there was a concomitant fibular pseudoarthrosis, whether there was concomitant ankle joint deformity, whether there was bone cystic change in the region of tibial bowing deformity, location of the apex of the bowing deformity, diameter of the tibial bowing deformity on the affected side, diameter on the healthy side in the same plane as the tibial bowing deformity, angle of lateral bending deformity of the tibia, angle of anterior bending deformity of the tibia, occurrence of tibia fracture, history of trauma before fracture, location of fracture, and age at the time of fracture. The follow-up endpoint was January 2023. Receiver operating characteristic (ROC) curves were used to determine the optimal cutoff values for the angles of lateral and anterior bending deformity of the tibia and the ratio of cross-sectional areas. The correlation between the above factors and tibial fractures in children was analyzed by single factor survival analysis, and the indicators with statistical significance were included in multivariate Cox proportional risk regression analysis to determine the risk factors for tibial fractures in children with ALBT.Results:Of the 87 children diagnosed with ALBT, the median age at initial diagnosis was 14.0 months (range, 1-93 months), with 42 males and 45 females, 44 left-sided and 43 right-sided cases. The median follow-up time for non-fracture cases was 42.0 months (range, 1-124 months). At the last follow-up, 43 children had experienced fractures, while 44 had not. The average time to fracture-free survival was 70.3 months, the median fracture-free survival time was 55.0 months, and the median survival time without fractures was 42.0 months. The ROC curve results indicated a cutoff value of 25.55° for the lateral bending angle of the tibia and 32.63° for the anterior bending angle of the tibia, with no statistically significant significance for the cross-sectional area ratio [AUC=0.54, 95% CI (0.42, 0.66), P=0.530]. Single-factor analysis of fracture-free survival suggested that there were statistically significant differences in the intergroup fracture-free survival rates of four factors: lateral bending angle of the tibia (χ 2=7.06, P=0.008), anterior bending angle of the tibia (χ 2=8.96, P=0.003), history of trauma (χ 2=18.26, P<0.001), and tibial bone cystic change (χ 2=4.30, P=0.038). The results of the multivariate Cox proportional hazards regression analysis showed that a lateral bending angle of the tibia&ge;25.55° ( HR=2.73, P=0.007), tibial bone cystic change ( HR=2.35, P=0.018), and history of trauma ( HR=2.65, P=0.004) were all positively correlated with fractures. Conclusion:The main risk factors for tibia fractures in children with ALBT include trauma, tibial bowing deformity with concomitant bone cystic change, and lateral bending angle of the tibia&ge;25.55°.

China ranks the first in the morbidity and mortality of malignant tumors in the world. Traditional Chinese medicine （TCM） has played a crucial part in preventing recurrence of malignant tumors after surgery， enhancing efficacy and reducing toxicity of radiotherapy and chemotherapy， and promoting the survival of patients with advanced tumors. According to the whole process of tumor occurrence and development and via the long-term clinical practice， we proposed the "deficiency-cold-toxin-blocking-failure evolution theory" of malignant tumors， with depression in the whole process and cancer toxin as the core. The theory is a summary of pathophysiological status at different stages of tumor development based on the disease and syndrome. This paper analyzes the biological basis of the theory. To be specific， we believe that the ''deficiency'' is mainly caused by the tumor immunoediting abnormality and thus the escape of clearance. In addition， the ''cold'' is in line with the metabolic reprogramming of the tumor microenvironment. ''Toxin'' is manifested as the stemness of cancer cells induced by the tumor microenvironment. ''Blocking'' is closely related to the enhanced permeability and retention （EPR） effect of tumor neovasculature. ''Failure'' is exactly the cachexia caused by change of dominant metabolism. ''Depression'' is essentially stress-the accelerated progress of the neuroendocrine system. In combination with clinical practice， we conclude that method of reinforcing healthy Qi and adjusting the balance can help recover tumor immunoediting and restore the immunity to regulate the "deficiency". Moreover， replenishing Qi and warming Yang can correct the tumor metabolic programming to avoid "cold". Removing toxin and reinforcing healthy Qi can reverse tumor cell stemness and thus suppress the ''toxin''. Eliminating the ''blocking'' source can suppress tumor angiogenesis and help regulate the EPR. Regulating Yin and Yang will help relieve the change of dominant metabolism and futile cycle to avoid ''failure''. The method of relieving depression and regulating Qi can relieve neuroendocrine stress and prevent tumor progression， thus alleviating the "depression". Moreover， effect of these methods was tested with effective formulas. This study is expected to lay a foundation for the establishment of a new system for the clinical prevention and treatment of malignant tumors.