Objective:To investigate alterations in the glymphatic system of spinocerebellar ataxia type 3 (SCA3) patients based on quantitative MRI, and its association with genetic information and motor dysfunction.Methods:The study was a cross-sectional study. This prospective study recruited 39 confirmed SCA3 patients (SCA3 group) and 40 matched healthy controls (HC group) who were seen at the Southwest Hospital of Army Medical University from May 2017 to June 2023. All subjects underwent cranial MRI scanning. Clinical assessments were conducted on all participants using the scale for the assessment and rating of ataxia (SARA) and the international cooperative ataxia rating scale (ICARS). The automatic segmentation and volume measurement of the choroid plexus based on Freesurfer 6.0; the perivascular interstitial space (PVS) was automatically segmented based on the deep-learning model VB-Net, and the volume of the PVS in each brain region was quantified after manual correction. Independent samples t-test and Mann-Whitney U-test were used to analyze the changes in the class lymphatic system in the SCA3 group and the HC group. Pearson partial correlation analysis was used to explore the relationship between CAG repeats, the glymphatic system, and motor dysfunction. Results:The standardized choroid plexus volume in the SCA3 group was (1.24±0.36)×10 3 mm 3, and that in the HC group was (0.96±0.34)×10 3 mm 3, with a statistically significant difference ( t=4.01, P<0.001). PVS volumes in the frontal lobe, temporal lobe, parietal lobe, basal ganglia, cerebellum, thalamus, and brainstem regions in the SCA3 group were significantly higher than those of HC group ( P<0.05). Partial correlation analysis revealed that CAG repeats in SCA3 group were positively correlated with SARA, ICARS, and basal ganglia PVS volumes ( r=0.65, 0.58, 0.29; P=0.001, 0.001, 0.042). Cerebellar and temporal lobe PVS volumes were positively correlated with SARA ( r=0.59, 0.47; P=0.001, 0.003), and positively correlated with ICARS scores ( r=0.61, 0.40; P=0.001, 0.011). Choroid plexus volume was positively correlated with cerebellar and basal ganglia PVS volumes ( r=0.41, 0.31; P=0.009, 0.043). Conclusions:The glymphatic system of SCA3 patients have significant alteration and have association with CAG repeats and motor dysfunction.

Objective To apply PDCA(plan,do,check and action)cycle to optimize management and shorten the turnaround time(TAT)of five sex hormone tests for improving clinical efficiency and patient satisfaction.Methods A PDCA management team was established to retrospectively analyze the current status of sex hormone TAT in 2021.In 2022,PDCA tools were utilized,including fishbone diagrams for cause analysis,Pareto charts for root cause identification,formulation and implementation of corresponding im-provement measures and monitoring of TAT changes before and after PDCA.Results Through the corrective measures,such as staff personnel training,increasing testing equipment,optimization process and other improvement measures,the median TAT of sex hor-mone samples in 2022(post-PDCA improvement)was 54 minutes which was significantly lower than 61 minutes in 2021(pre-PDCA)(P＜0.05).Conclusion PDCA cycle management effectively shortened the TAT of sex hormone samples,enhanced clinical diagnostic efficiency and patient experience,improved doctor-patient relationships,while also elevated the quality management standards of labo-ratories.

ObjectiveTo perform a predictive analysis of the quality marker（Q-Marker） for the anticoagulant activity of Kunning granules. MethodThe chemical components of Kunning granules were analyzed by ultra high performance liquid chromatography-quadrupole-time of flight tandem mass spectrometry（UHPLC-Q-TOF-MS/MS） on a Waters ACQUITY UPLC HSS T3 column（2.1 mm×100 mm， 1.8 μm） with the mobile phase of acetonitrile（A）-25 mmol∙L^-1 ammonium acetate aqueous solution（B） for gradient elution （0-5 min， 5%-22%A； 5-10 min， 22%-30%A； 10-15 min， 30%-95%A； 15-20 min， 95%-5%A； 20-30 min， 5%A）， flow rate of 0.2 mL∙min^-1， column temperature at 30 ℃， injection volume of 1 μL， electrospray ionization（ESI）， positive and negative ion detection modes. Interaction analysis between the targets of chemical components and the targets of abnormal uterine bleeding（AUB） was performed by network pharmacology， and the key components were screened through network topology analysis. The fingerprints of 10 batches of Kunning granules were established by high performance liquid chromatography（HPLC）， the anticoagulant activity of the granules was determined by blood coagulation method and fibrinogen plate method， and the spectrum-effective relationship was established. The components co-occurring in the topological analysis and spectrum-effective relationship were selected as Q-Markers， and their anticoagulant activities were verified and confirmed. ResultA total of 475 chemical components were identified from Kunning Granule， of which 22 key components such as salvianolic acid B， paeoniflorin， naringin and neohesperidin， were the potential material basis for the treatment of AUB. The spectrum-effective analysis showed that peaks 7（paeoniflorin）， 9（naringin）， 10（neohesperidin） and 11（salvianolic acid B） were the optimal principal components， and in vitro activity test showed that these four components could better characterize their anticoagulant activity. ConclusionSalvianolic acid B， paeoniflorin， neohesperidin and naringin may be Q-Markers for the anticoagulant activity of Kunning granules.

Objective: Heavy metal and harmful element contamination are frequently reported in Chinese herbal medicines (CHMs), and roots and rhizomes parts showed a higher content than other parts. To investigate the residue level and assess the potential human health risk of heavy metals and harmful elements in roots and rhizomes, 720 batches of the sample representing 20 species of herbs from different sources were collected. Methods: The content of Pb, Cd, As, Hg, and Cu in the digests was determined using ICP-MS. The chronic hazard index estimate based on non-cancer hazard quotient (HQ) was applied for potential health risk assessment of Pb, Cd, As, Hg, and Cu via consumption of CHMs. Results: Compared with the Chinese limit standard (Chinese Pharmacopoeia Commission, 2020 edition) of Pb, Cd, As, Hg, and Cu in Ginseng Radix et Rhizoma, the exceedance percentage of Pb in total samples was 14.1%, which were generally far higher than Cd, As, Hg, and Cu. Health risk assessment results based on hazard quotient calculating showed that total HQ of Cu, Pb, As, Cd, and Hg in Pulsatillae Radix and Clematidis Radix et Rhizoma exceeded 1, with the value of 1.543 and 1.235. Besides, Arsenic had the highest HQ value (0.957) in Pulsatillae Radix. Conclusion: Consuming raw materials of Pulsatillae Radix and Clematidis Radix et Rhizoma may pose a potential risk and Arsenic residues in Pulsatillae Radix deserved special attention.

Objective To investigate the clinicopathological characteristics and independent risk factors of hepatocellular carcinoma (HCC) differentiation.  Methods A total of 108 HCC patients who underwent operation and treatment were reviewed and classified into low differentiation group (n= 29, 26.85%), medium differentiation group (n=53, 49.07%) and high differentiation group (n=26, 24.07%) according to pathological diagnosis. The clinicopathological characteristics and the expression levels of Ki67 and P53 in each group were compared and analyzed. Logistic regression model was used to analyze the risk factors for low differentiation of HCC.  Results The proportion of cirrhosis, the positive rate of P53 and Ki67 expression level in different degrees of HCC differentiation were statistically significant (P<0.05). However, there were no significant differences in age, sex, tumor diameter, and hepatitis virus infection or tumor involvement of liver capsule between groups (P>0.05). Multivariate logistic analysis showed that cirrhosis (OR=3.408), high expression of Ki67 (OR=11.113) and positive P53 (OR=9.711) were the main risk factors for poorly differentiated HCC.  Conclusion There are differences in clinical characteristics and expressions of Ki67 and P53 in HCC patients with different degrees of differentiation. Logistic regression analysis can identify clinicopathological risk factors affecting the degree of differentiation of HCC, which can provide criterion support for accurate diagnosis and prognostic treatment.

Objective: To investigate the effect of overexpression of Notch intracellular domain (NICD) on proliferation and osteogenic differentiation of human periodontal ligament stem cells (hPDLSC). Methods: The third generation hPDLSC with stable overexpressing of NICD were assigned as experimental group, normal hPDLSC were as negative control group and hPDLSC transfected with empty vector were as blank control group. The effect of overexpressing NICD on proliferation ability of hPDLSC was detected by using cell counting kit-8 (CCK-8). Alizarin Red staining and real-time quantitative PCR (qPCR) were used to detect the effects of NICD on cementum attachment proteins (CAP), osteocalcin (OCN), Runt-related transcription factor 2 (RUNX2) and Notch signal pathway receptor Notch1. The effect of overexpressing NICD on hPDLSC osteogenic protein RUNX2 and flag marker protein (used to label NICD) were detected by using Western blotting. Results: CCK-8 results showed that there were no significant differences in A values amongst the three groups for 1-2 days (P>0.05). The number of cells in the experimental group was significantly increase than that of the two control groups from the third to seventh days (A values were 0.203±0.016, 0.364±0.014, 0.449±0.020, 0.549±0.020 and 0.570±0.020, respectively) (P<0.05). Alizarin red staining showed that compared with the blank control group and negative control group, the mineralized nodules in the experimental group had smaller formation range and lighter color, and the differences were statistically significant (P<0.05). The expressions of CAP gene (0.751±0.058, 0.887±0.025), osteocalcin gene (0.592±0.051, 0.670±0.045) and RUNX2 gene (0.319±0.038, 0.684±0.055) at 14 and 21 days in the experimental group were significantly lower than those in the negative control group respectively (P<0.05). However, the expression levels of Notch1 gene at 14 and 21 days (2.507±0.047, 4.041±0.219) were significantly higher than those of negative and blank control groups (P<0.05). The results of Western blotting showed that the expressions of flag marker protein (0.167±0.007, 0.204±0.010) at 14 and 21 days in the experimental group were significantly higher than those in the negative and blank control groups (P<0.05). However, the expressions of RUNX2 protein (0.075±0.006, 0.074±0.013) at 14 and 21 days were significantly lower than that in the negative control group (0.092±0.003, 0.118±0.008) and blank control group (0.174±0.006, 0.212±0.008) (P<0.05). Conclusions Overexpression of NICD can promote the proliferation capacity of hPDLSC and inhibit its osteogenic differentiation.

Objective To evaluate the biological effect of tumor necrosis factor-α(TNF-α) on the proliferation and multi-directional differentiation of stem cells from rat apical papilla(SCAP).Methods SCAP was extracted by combining enzyme digestion method with tissue block method.The cells were divided into control group(TNF-α 0 ng/mL) and experimental group(TNF-α 5,10,20,50 ng/mL).The ability of proliferation of SCAP was measured by MTT method.The ability of osteogenic/dentinogenic differentiation of SCAP was measured by alizarin red staining and quantitative real-time PCR.The ability of adipogenic of SCAP was measured by oil red O staining.The expression of vascular related genes of SCAP was measured by quantitative real-time PCR.Results SCAP was consistent with the characteristics of mesenchymal stem cells and possessed the ability of multi-directional differentiation.The MTT results showed that experimental group promoted the proliferation of SCAP in comparison with the control group.The difference was statistically significant(P<0.05),and 10 ng/mL was the optimum concentration.The results of alizarin red staining showed that with the increase of the concentration of TNF-α,the mineralized nodules in the experimental group gradually became smaller,and the number of the formation decreased gradually.The results of quantitative real-time PCR showed that the expression of OC,DMP-1 and DSPP in the experimental group was significantly lower than that of the control group at 3 and 7 days,in which the expression of OC was statistically significant different(P<0.05);at 14 days,the expression of OC,DMP-1 in the experimental group was significantly lower than that of the control group(P<0.05).The result of Oil red O staining showed that with the increase of the concentration of TNF-α,the lipid droplets formation in the experimental group gradually decreased.The result of quantitative real-time PCR showed that the expression of ANGPT1,VEGFA,PECAM-1 in the experimental group was significantly lower than that of the control group(P<0.05).Conclusion TNF-α might promote the proliferation and inhibit the multi-directional differentiation of SCAP.

Objective: This in vitro study aimed to investigate the remineralization effect of the natural medicine epigallocatechin gallate on artificial enamel caries in primary human teeth. Methods : We divided 30 sound primary upper anterior teeth into 3 groups according to a random number table, including experimental group (epigallocatechin gallate group), positive control group (NaF group) and blank control group (artificial saliva group), with 10 teeth in each group. After test in vitro, Micro Hardness Tester was applied to measure hardness of samples before and after demineralization. Scanning electron microscopy (SEM) was used to observe the result of primary enamel surface remineralization. Results: A significant increase in enamel surface microhardness between the three groups after remineralization (F=1 199.975, P < 0.05). The difference between 2 groups was compared with each other among 3 groups. Statistical significance was found (vs experimental group q=41.986, P < 0.05; vs positive control group q=68.174, P < 0.05), suggesting that both positive control group and experimental group could promote the remineralization of primary enamel, and the effect of epigallocatechin gallate was weaker than NaF (q=26.188, P < 0.05 ). The results from SEM indicated that there was large amount of sediment on the surface of primary enamel surface of incisors in the experimental group and positive control group, while primary enamel surface of incisors in the blank control group was honeycomb and uneven, with less sediment. Conclusion Based on this in vitro study, epigallocatechin gallate can promote the remineralization of demineralized enamel of primary teeth, indicating its potential use as a natural remineralization medicine.

OBJECTIVEWe explored the expressions of the Notch and Wnt signaling pathways and their significance in the repair process of alveolar bone defects by establishing animal models with a composite of autologous bone marrow mesenchymal stem cells (BMSCs) and platelet-rich fibrin (PRF) to repair bone defects in the extraction sockets of rabbits.

METHODSA total of 36 two-month-old male New Zealand white rabbits were randomly divided into four groups, and the left mandibular incisors of all the rabbits were subjected to minimally invasive removalunder general anesthesia. BMSC-PRF compounds, single PRF, and single BMSC were implanted in Groups A, B, and C. No material was implanted in Group D (blank control). The animals were sacrificed at 4, 8 and 12 weeks after surgery, the bone defect was immediately drawn, and the bone specimens underwent surgery after four, eight, and twelve weeks, with three rabbits per time point. The expressions of Notch1 and Wnt3a in the repair process of the bone defect were measured via immunohistochemical and immunofluorescence detection.

RESULTSImmunohistochemistry showed that the expressions of Notch1 and Wnt3a in Groups A, B, and C were higher than that in Group D at the fourth and eighth week after operation (P<0.05). By contrast, the expressions of Notch1 and Wnt3a in Group D were higher than those in Groups A, B, and C at the twelfth week (P<0.05). Immunofluorescence showed that the expressions of both Notch1 and Wnt3a reached their peaks in the new bone cells of the bone defect after four weeks following surgery and gradually disappeared when the bone was repaired completely.

CONCLUSIONNotch1 and Wnt3a signaling molecules are expressed in the process of repairing bone defects using BMSC-PRF composites and can accelerate the healing by regulating the proliferation and differentiation of BMSCs. Moreover, the expressions of Notch and Wnt are similar, and a crosstalk between them may exist it.

Alveolar Bone Grafting ; methods ; Animals ; Blood Platelets ; Bone Marrow Cells ; cytology ; Bone Transplantation ; methods ; Bone and Bones ; abnormalities ; Cell Differentiation ; Fibrin ; administration & dosage ; Male ; Mesenchymal Stem Cell Transplantation ; methods ; Mesenchymal Stromal Cells ; Platelet-Rich Plasma ; Rabbits ; Random Allocation ; Receptor, Notch1 ; metabolism ; Tissue Engineering ; Wnt Signaling Pathway ; Wnt3A Protein ; metabolism ; Wound Healing

BACKGROUND:The periodontal ligament stem cels can promote periodontal tissue regeneration, providing a new way for the treatment of periodontitis. OBJECTIVE:To observe the inflammatory microenvironment effects on the biological properties of periodontal ligament stem cels. METHODS: Periodontal ligament stem cels from healthy controls and patients with periodontitis were primarily cultured by tissue digestion method, purified using limited dilution method, and identified through detection of CD146 and STRO-1. Then, passage 3 cels were taken and denoted as normal control and inflammation groups folowed by osteogenic induction. RESULTS AND CONCLUSION:Purified cels from two sources both expressed STRO-1 and CD146. Periodontal ligament stem cels in the inflammation group showed higher multiplication capacity, but the osteogenesis ability was lower compared with the normal control group. The expressions of Runx2 mRNA and Osterix mRNA were dropped significantly after the stimulus of tumor necrosis factor-α (P < 0.05), but the interleukin-1β and interleukin-6 did not have a significant impact. Tumor necrosis factor-α at 0.1 and 1 μg/L had no significant effects on the expression of Runx2 mRNA, but the expression of Runx2 mRNA was decreased significantly after treatment with 10 μg/L tumor necrosis factor-α (P< 0.05). It is confirmed that the molecular signaling mechanism inside the periodontal ligament stem cels is changed under inflammatory microenvironment, so that the differentiation capacity of cels from the inflammatory sources is lowered. Moreover, tumor necrosis factor-α is one of the key factors and its optimalconcentration is 10 μg/L.