Objective To screen out specific microsatellite markers for use in Tupaia belangeri chinensis genetic testing. Methods Firstly to screen about 700 microsatellite loci from whole genome.Secondly to choose about 100 better loci without defect factors.Lastly 46 primers were designed by 33 tree shrew’ s microsatellite loci obtained from whole genome and other references.Agarose gel electrophoresis and polyacrylamide gel electrophoresis were used for PCR products, and better loci based on electrophoresis results were chosen.Then STR scan was used to select the microsatellite loci combination for genetic testing.Results Twenty-two microsatellite loci were selected with a significant Stutter peak on STR scanning.Comparing the alternative loci and ultimately selected loci, there were two loci available in the five alternative loci of T.glis.The coincidence rate between T.glis and T.b.chinensis was 40%.There were two loci available in the five alternative loci of T.minor, and the coincidence rate between T.minor and T.b.chinensis was 40%.There were two loci available in the three alternative loci of T.belangeri, and the coincidence rate between T.belangeri and T.b. chinensis was about 70%.Conclusions The 22 microsatellite loci screened in this study are well applied for genetic testing of Tupaia belangeri chinensis, therefore, provide a scientific basis for the genetic quality monitoring of tree shrews.

Objective To explore the metabolite profiles of mesenchymal stem cells(MSCs)underwent death using 9.4 T high resolution MR spectroscopy.Methods MSCs were cultured and treated for 6,12 and 24 hours in a stimulated condition which included hypoxia,serum deprivation and changes of microenvironment.Cell death and the mortality was detected by light microscopy,Hocchst staining and flow cytometry analyses.The morality of stem cells was analyzed using one-way analysis of variance (ANOVA).Cell metabolite extraction was prepared by methanol-chloroform(M/C) method and analyzed on a 9.4 T MR device.1H-MR spectroscopy was obtained and the metabolite concentration of each time point was calculated and compared using one way ANOVA,the difference between two groups was analyzed by SNK test.Results Necrosis was the major form of cell death in the built model.The morality of every time sets was 16±4(0 h),658±61 (6 h),1 571 ± 154(12 h) and 2 816± 178(24 h) respectively,and the difference between each groups were statistically significant (F=298.96,P＜0.01).After induced stem cells death for 6,12 and 24 h,the metabolite concentrations at 0.89 ppm was (1.48±0.69),(2.32±0.63)and (2.15±0.45)nmol/mg respectively,and increased compared to thc control[(1.41 ±0.25)nmol/mg]with statistical significance (F=329.57,P＜0.01).The metabolite concentrations at 1.28 ppm was (6.42±0.31),(7.26±0.32)and (7.01 ±0.61)nmol/mg,respectively,and increased compared to the control[(5.76 ±0.74)nmol/mg]with statistical significance (F=19.56,P＜0.01).The metabolite concentrations at 1.60 ppm was (2.36±0.31),(2.29±0.16)and (2.31 ± 0.24) nmol/mg respectively,and increased compared to the control[(1.96 ± 0.27)nmol/mg]with statistical significance (F=4.35,P＜0.05).After induced stem cells death for 12 hours,the metabolite concentrations at 0.89 ppm was increased compared to 6 hours with statistical significance (P＜0.05).The metabolite concentrations at 1.28 ppm was increased compared to 6 hours with statistical significance (P＜0.05).After induced stem cells death for 24 hours,the metabolite concentrations at 0.89 ppm was decreased compared to 12 hours with statistical significance (P＜0.05).Conclusions There are some specific characteristics on MRS of MSCs underwent death,and the fatty acid peak may serve as a biomarker for cell death.

Objective:To analyze the characteristics of cervical lymph node metastasis in tongue squamous cell carcinoma (TSCC).Methods:A retrospective study was conducted. A total of 329 patients with TSCC who underwent en bloc resection of primary tumor and neck dissection in the Second Xiangya Hospital of Central South University from June 2010 to March 2018 were included. There were 283 males and 46 females, aged from 26 to 80 years. All patients underwent the modified neck dissection. The main difference between the modified neck dissection and the traditional neck dissection lay in the managements of unconventional lymph nodes. The lymphatic adipose tissues adjacent to the superior thyroid artery, the base of facial artery and the branches of external carotid artery were thoroughly dissected. The primary tumor as well as lingual artery, tissues along the lingual artery and lymph nodes in the mouth floor were resected. χ 2 test was used for comparison of count data, and linear regression model was used for multivariate analysis. Results:Cervical lymph node metastases were found in 136 patients (41.3%). Among 142 patients (T1-2cN0) with supraomohyoid neck dissection, 22 patients had pathologically occult lymph node metastases (15.5%), with a 5-year overall survival rate of 90.2%, which was similar to the 5-year overall survival rate of 92.1% in 120 patients without lymph node metastasis (χ 2=0.156, P=0.693). Multivariate linear regression analysis showed that T stage, clinical stage and unconventional lymph node metastasis were important factors for cervical lymph node metastasis in tongue cancer patients ( P<0.05). Unconventional lymph node metastases occurred in 30 patients (9.1%), including the metastases of lymph nodes in the floor of mouth (3.0%), the lingual artery (2.4%), the base of the external maxillary artery (2.1%), the superior thyroid artery (0.9%), and the external carotid artery (0.6%). There were significant differences in the unconventional lymph node metastasis rates between patients with negative and positive conventional lymph node metastases [4.9%(10/203) vs. 15.9%(20/126), χ 2=11.242, P=0.001] and also between patients with depth of invasion ≤5 mm, 5 mm 10 mm [3.1%(2/64) vs. 5.7%(6/106) vs. 13.8%(22/159), χ 2=7.907, P=0.005]. Conclusion:Supraomohyoid neck dissection can achieve reliable control efficacy in patients with cN0 tongue cancer. All patients with lymph node dissection should undergo unconventional lymph node dissection. Unconventional lymph node dissection is strongly recommended for patients with conventional lymph node metastasis.

ObjectiveTo observe the effect of Simiaowan on the intestinal ATP-binding cassette superfamily G （White） member 2 （ABCG2） expression and the intestinal uric acid excretion in hyperuricemia rats. MethodA total of 48 SD male rats were randomized into the normal， model， benzbromarone （4.7 mg·kg^-1）， and high-， medium-， low-dose Simiaowan groups （2 260.6， 1 130.3， 565.2 mg·kg^-1， respectively）， with 8 rats in each group. Potassium oxonate and hypoxanthine was employed to induce hyperuricemia in rats （21 days）. On the 8^th day， administration began （once a day for 14 days）. Rats were killed on the 21^st day， and serum uric acid， serum creatinine， blood urea nitrogen， and intestinal uric acid were detected. The protein expression of ABCG2 in the small intestine was detected by Western blot. The ABCG2 protein expression and localization in intestinal tissues were determined by immunohistochemistry. The ABCG2 mRNA expression in small intestine was measured by quantitative real-time PCR. ResultThe levels of serum uric acid， serum creatinine， and blood urea nitrogen in the model group were higher than those in the normal group （P<0.01）. Low level of serum uric acid in the three Simiaowan groups and benzbromarone group （P<0.01）， high level of intestinal uric acid in medium-dose and low-dose Simiaowan groups （P<0.05， P<0.01）， high level of serum creatinine in benzbromarone group （P<0.01）， and low level of blood urea nitrogen in low-dose Simiaowan group （P<0.05） were observed as compared with those in the model group. Serum uric acid showed insignificant difference between the low-dose Simiaowan group and benzbromarone group. The expression of ABCG2 protein in the model group was lower than that in the normal group （P<0.05）. The expression of ABCG2 protein in the medium-dose and low-dose Simiaowan groups （P<0.05， P<0.01）， the high-dose Simiaowan group， and benzbromarone group increased as compared with that in the model group. ABCG2 mRNA expression was insignificantly different between the model group and the normal group， while the expression in the medium-dose and low-dose Simiaowan groups was higher than that in the model group （P<0.05）. ABCG2 protein was mainly distributed in intestinal villi， and ABCG2 protein expression demonstrated no significant difference between the model group and the normal group. The ABCG2 protein expression in the three Simiaowan groups increased as compared with that in the model group （P<0.05）. ConclusionSimiaowan can significantly reduce the serum uric acid level in hyperuricemia rats. Particularly， the low-dose Simiaowan shows similar efficacy to benzbromarone in lowering uric acid and protects renal function. The mechanism is the likelihood that it up-regulates intestinal ABCG2 expression to promote intestinal excretion of uric acid.

Objective:To understand the epidemiological characteristics and spatiotemporal distribution of viral encephalitis in children and adolescents in Henan Province from 2012 to 2023.Methods:The information about viral encephalitis cases from October 1, 2012 to July 26, 2023 were collected from Zhengzhou Children's Hospital (National Children's Regional Medical Center),Henan Provincial Children's Hospital for the analyses on temporal distribution the cases, the severe illness rate, age distribution, pathogen type and imaging findings of the cases.Results:A total of 6 276 cases of viral encephalitis were included in this study after excluding cases with incomplete information. The cases mainly originated from Zhengzhou (38.96%), followed by Zhoukou (9.93%), Xuchang (8.68%), Zhumadian (7.90%) and Pingdingshan (7.39%). The cases in boys accounted for 62.13% and the cases in girls accounted for 37.87%. Most cases (72.45%) occurred in age group 7-13 years. The overall rate of severe illness cases was 4.51% from 2012 to 2023. There were significant differences in severe illness cases among different areas and years ( χ2=5.33, P=0.021; χ2=48.14, P<0.001). Enteroviruses were mainly detected (31.57%), in which Coxsackie virus was predominant (58.37%). Imaging findings showed that cerebral hemisphere damage was most common in children and adolescents with viral encephalitis (54.93%). Conclusions:From 2012 to 2023, more cases of viral encephalitis occurred in boys in Henan. Children and adolescents aged 7-13 years were the main affected group. The prevention of enteroviruses infection, especially Coxsackie virus, needs to be strengthened. Special attention should be paid to the prevention of cerebral hemisphere damage after viral encephalitis diagnosis.

Objective:To evaluate the clinical value of sonography based volume computer aided display heart (SonoVCADheart) in the display of key diagnostic elements in basic fetal echocardiographic views.Methods:4D volume data based on fetal four-chamber view of 80 singleton fetuses (including 57 normal fetuses and 23 fetuses with heart abnormalities) were collected by using a volumetric probe from the Sir Run Run Shaw Hospital, Zhejiang University College of Medicine from January 8-22, 2019. Four to five volume datasets based on four-chamber view of the heart were rapidly acquired from each fetus. Three doctors (Doctor A: Engaged in fetal echocardiography diagnosis for more than 15 years, Doctor B and C: 6 months of regular fetal echocardiography training but lack of experience) with different fetal echocardiographic experiences performed off-line processing using SonoVCADheart combined tomographic ultrasound imaging (TUI) at different times to obtain eight standard echocardiographic diagnostic views, and to score the elementary contents of each echocardiographic view. The scores of the same doctor at different times, the scores of less experienced doctors and experienced doctor, and the scores between normal and abnormal fetuses, and the time required for analysis and diagnosis among all doctors were analyzed and compared, respectively. The diagnostic coincidence rates of SonoVCADheart for fetuses with cardiac malformations were also assessed.Results:A total of 279 volume datasets obtained from 57 normal fetuses, an average of 4.89/fetus, and 109 volume datasets obtained from 23 fetuses with cardiovascular abnormalities, an average of 4.74/fetus, and all volume datasets were used for SonoVCADheart analysis. The volume percentage of all the elements in the 8 diagnostic views with image quality≥2 points shown by SonoVCADheart in the normal fetuses was about 70.61%-74.91%, in the abnormal fetuses was about 53.21%-55.96%. There were no significant differences in the scores between the same doctor at different times, the scores between inexperienced doctors, and the scores among experienced and less experienced doctors(all P>0.05). There were significant differences in the scores between normal and abnormal fetuses except for all of the superior and inferior vena cava view and the aorticarch view of doctors A and C(all P<0.05). The time required for experienced doctor A and inexperienced doctors B and C to obtain 8 diagnostic views and to complete the diagnosis was statistically significant ( P<0.05). There was no significant difference in the time required for the inexperienced doctors B and C to obtain 8 diagnostic sections and to complete the diagnosis ( P>0.05). There was statistically significant difference in the diagnosis time required for doctor A using SonoVCADheart and two-dimensional echocardiography in fetuses with cardiovascular malformations ( P<0.05). However, there was no statistically significant difference in the time required in the normal fetus between SonoVCADheart and two-dimensional echocardiography ( P>0.05). The diagnostic coincidence rate of SonoVCADheart for fetuses with cardiac malformations is about 89.91%-90.83%. Conclusions:SonoVCADheart is a repeatable and stable novel fetal heart processing tool enabling displaying eight standard diagnostic sections of the fetal heart, and has potential clinical application value in the standardization of image acquisition and sequence display.

BACKGROUND:Inflammation affects the osteogenic differentiation of periodontal ligament stem cells,and the osteogenic ability and autophagy level of periodontal ligament stem cells are closely related.However,there are no relevant reports on whether inflammation affects the osteogenic ability and autophagy level of periodontal ligament stem cells at different stages of osteogenic differentiation. OBJECTIVE:To explore alkaline phosphatase expression and autophagy periodontal ligament stem cells levels in periodontitis and normal conditions. METHODS:Periodontal ligament stem cells from normal and periodontitis patients were isolated and cultured,and underwent Vimentin,pan-CK,and Stro-1 fluorescence staining.At 3,7,and 14 days of osteogenic differentiation,western blot assay was used to detect the protein expression levels of alkaline phosphatase,LC3B,Beclin1,and ATG5 in normal and inflammatory periodontal ligament stem cells.The mRNA expression levels of alkaline phosphatase,bone sialoprotein,osteocalcin,Runx2,LC3B,Beclin1,and ATG5 were detected by real-time PCR. RESULTS AND CONCLUSION:(1)Stro-1 was positive,Vimentin was positive,and pan CK was negative in periodontal ligament stem cells.(2)At 3,7,and 14 days after osteogenic differentiation,compared with normal periodontal ligament stem cells,the mineralization nodules formed by periodontal ligament stem cells from inflammatory sources were significantly reduced(P<0.01);the expression of alkaline phosphatase protein and mRNA was significantly lower(P<0.05);the mRNA expression levels of bone sialoprotein,osteocalcin,and Runx2 were significantly decreased(P<0.05).(3)At 7 and 14 days after osteogenic differentiation,compared with normal periodontal ligament stem cells,the expression levels of ATG5,LC3B,and Beclin1 proteins and mRNA of periodontal ligament stem cells were downregulated(P<0.05).These findings suggest that inflammation reduces the activity of periodontal ligament stem cells in mineralizing nodule formation and the expression of alkaline phosphatase and weakens the autophagy potential of periodontal ligament stem cells at 7 and 14 days after osteogenic differentiation.

In order to promote the innovative application of Sanjiao theory and Yingwei theory， this paper tries to apply the ''Sanjiao-Yingwei'' Qi transformation theory to the treatment of tumor diseases， integrating it with T cell exhaustion mechanism to elaborate on its scientific connotation and using network pharmacology and bioinformatics to elucidate the correlation between the anti-tumor mechanism of ''Sanjiao-Yingwei'' Qi transformation and T cell exhaustion. The ''Sanjiao-Yingwei'' Qi transformation function is closely related to the immunometabolic ability of the human body， and the ''Sanjiao-Yingwei'' Qi transformation system constitutes the immunometabolic exchange system within and outside the cellular environment. Cancer toxicity is generated by the fuzzy Sanjiao Qi， and the long-term fuzzy Sanjiao Qi is the primary factor leading to T cell exhaustion， which is related to the long-term activation of T cell receptors by the high tumor antigen load in the tumor microenvironment. Qi transformation malfunction of the Sanjiao produces phlegm and collects stasis， which contributes to T cell exhaustion and is correlated with nutrient deprivation， lipid accumulation， and high lactate levels in the immunosuppressed tumor microenvironment， as well as with the release of transforming growth factor-β and upregulated expression of programmed death receptor-1 by tumor-associated fibroblasts and platelets in the tumor microenvironment. Ying and Wei damage due to Sanjiao Qi transformation malfunction is similar to the abnormal manifestations such as progressive loss of exhausted T cell effector function and disturbance of cellular energy metabolism. Guizhi decoction， Shengming decoction， and Wendan decoction can correct T cell exhaustion and exert anti-tumor effects through multi-target and multi-pathways by regulating ''Sanjiao-Yingwei'' Qi transformation， and hypoxia inducible factor-1α （HIF-1α） may be one of the main pathways to correct T cell exhaustion. It was found that HIF-1α may be one of the important prognostic indicators in common tumors by bioinformatics. The use of the ''Sanjiao-Yingwei'' Qi transformation method may play an important part in improving the prognosis of tumor patients in clinical practice.

Incorporation of multiple functions into one nanoplatform can improve cancer diagnostic efficacy and enhance anti-cancer outcomes. Here, we constructed doxorubicin (DOX)-loaded silk fibroin-based nanoparticles (NPs) with surface functionalization by photosensitizer (N770). The obtained nanotheranostics (N770-DOX@NPs) had desirable particle size (157 nm) and negative surface charge (-25 mV). These NPs presented excellent oxygen-generating capacity and responded to a quadruple of stimuli (acidic solution, reactive oxygen species, glutathione, and hyperthermia). Surface functionalization of DOX@NPs with N770 could endow them with active internalization by cancerous cell lines, but not by normal cells. Furthermore, the intracellular NPs were found to be preferentially retained in mitochondria, which were also efficient for near-infrared (NIR) fluorescence imaging, photothermal imaging, and photoacoustic imaging. Meanwhile, DOX could spontaneously accumulate in the nucleus. Importantly, a mouse test group treated with N770-DOX@NPs plus NIR irradiation achieved the best tumor retardation effect among all treatment groups based on tumor-bearing mouse models and a patient-derived xenograft model, demonstrating the unprecedented therapeutic effects of trimodal imaging-guided mitochondrial phototherapy (photothermal therapy and photodynamic therapy) and chemotherapy. Therefore, the present study brings new insight into the exploitation of an easy-to-use, versatile, and robust nanoplatform for programmable targeting, imaging, and applying synergistic therapy to tumors.

Objective: To explore the feasibility of Sonography based Volume Computer Aided Display Heart (SonoVCADheart) in the display of the fetal ventricular outflow views, and compare diameters of fetal aorta (AO) and pulmonary artery (PA) measured by two-dimensional echocardiography (2DE) and SonoVCADheart. Methods: Eighty singleton fetuses in the second and third trimesters of pregnancy in January 2019 in Sir Run Run Shaw Hospital, Zhejiang University College of Medicine were enrolled. Conventional 2DE examinations were performed. The volume datasets were analyzed offline using the new automatic image processing software SonoVCADheart. The diameters of AO and PA were measured by 2DE and SonoVCADheart, respectively. Pearson correlation analysis was used to evaluate the correlation between the two methods. The consistency of the two methods was verified by Bland-Altman analysis, and he reliability of SonoVCADheart was assessed using the intraclass correlation coefficient (ICC). Results: Fetal ventricular outflow views were successfully obtained using SonoVCADheart in 73(91.2%) of 80 fetuses. There were good correlations between the two methods for measuring the diameters of AO and PA (r=0.953, 0.971; all P<0.001). The 95% agreement limits of AO and PA were (-0.669, 0.568)mm and (-0.632, 0.580)mm, respectively. ICC demonstrated that SonoVCADheart achieved great repeatability both between and within observers. Conclusions SonoVCADheart may have potentials for the quantitative evaluation of fetal ventricular outflow tracts with its good repeatability and reliability.