Objective To optimize the immune scheme of SARS-CoV-2 RBD recombinant protein vaccine based on P.pastoris,and investigate the effect of different adjuvants on neutralizating antibody(NAb)titer,in order to provide reference for the continuous optimization research of SARS-CoV-2 vaccine.Methods The RBD protein was selected and the corresponding gene fragment was synthesized,which was constructed into the pPICZαA plasmid,and the plasmid was integrated into the genome of P.pastoris after linear transforma-tion for recombinant expression.The obtained recombinant protein vaccine was combined with different adju-vants to immunize mice to evaluate its immunogenicity.Results Both the target proteins wtRBD and Delta RBD were able to achieve satisfactory overexpression through the P.pastoris system.Compared with the 42 d interval,the IgG antibody titer at the 28 d interval increased by 1.8 times(44 923 vs.80 507).After 3 doses of immunization at an interval of 28 d,the geometric mean titer of NAb for Delta variant was 2.5 times higher than that at an interval of 42 days(2 191 vs.891).After immunization with Delta RBD recombinant protein vaccine combined with aluminum adjuvant,the NAb geometric mean titer for Delta variants reached 32 255(2 167-88 084).When using 5 μg or 30 μg Delta RBD immunization,the NAb titers of the aluminum adju-vant+CpG adjuvant group were about 10 times higher than those of the aluminum adjuvant group alone.Af-ter the third immunization,there was no significant difference in Delta RBD specific IgG titers between the 5 μg antigen group and the 30 μg antigen group(P＞0.05).Conclusion Both wtRBD and Delta RBD prepared based on P.pastoris could be used as effective antigens,with three doses of vaccine administered at a 28 day in-terval being the most effective.The combined immunization of Delta RBD recombinant protein with aluminum adjuvant+CpG adjuvant could obtain higher titers of NAb to exert immune effects on SARS-CoV-2 and its va-riants,providing some reference for the continuous optimization research of SARS-CoV-2 vaccines.

Objective:To establish a scientific and standardized evaluation index system of neonatal breastfeeding nursing quality under the state of mother-infant separation, in order to provide a tool for the detection and management of neonatal breastfeeding nursing quality under mother-infant separation, so as to promote the implementation of breastfeeding in China.Methods:A systematic and comprehensive search of Chinese and English databases was conducted to collect guidelines on neonatal breastfeeding in the state of mother-infant separation. Based on the "structure-process - result" three-dimensional quality structure model, the preliminary draft of evaluation indexes of neonatal breastfeeding nursing quality in the state of mother-infant separation was formed. The evaluation indexes of neonatal breastfeeding nursing quality in the state of mother-infant separation were constructed by Delphi expert correspondence method from July to October, 2023.Results:The effective recovery rates were 95.83% (23/24) and 91.30% (21/23), respectively. The expert authority coefficients were 0.895 and 0.870, respectively. The Kendall harmony coefficients of the whole index were 0.134 and 0.178 (both P<0.01), respectively. The final evaluation index system of neonatal breastfeeding nursing quality under the state of mother-infant separation includes 3 first-level indicators, 18 second-level indicators and 82 third-level indicators. Conclusions:The evaluation index system of neonatal breastfeeding nursing quality in the state of mother-infant separation established in this study is highly scientific and reliable, highlighting the characteristics of neonatal breastfeeding nursing work in the state of mother-infant separation, and can provide a reference for the evaluation of neonatal breastfeeding nursing quality in the state of mother-infant separation.

Objective:To analyze the clinical phenotype and genetic characteristics of a patient with Isidor-Toutain spinal epiphyseal dysplasia (SEMD) due to variant of RPL13 gene. Methods:A pregnant woman at 18 weeks of gestation who had presented at Quzhou Maternal and Child Health Care Hospital on January 14, 2023 was selected as the study subject. Whole exome sequencing (WES) was carried out for the patient, and candidate variant was validated by Sanger sequencing and bioinformatic analysis.Results:The woman was 37 years old with extremely short stature (135 cm) and "O" shaped legs. WES revealed that she has harbored a c. 548G>C (p.Arg183Pro) missense variant of the RPL13 gene (NM_000977.4). The same variant was not found in her fetus. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was predicted to be likely pathogenic (PS4+ PM2_Supporting+ PP3+ PP4). Conclusion:Isidor-Toutain type SEMD due to variants of the RPL13 gene may have variable expressivity and diverse clinical phenotypes. Above finding has facilitated the differential diagnosis and genetic counseling for this family.

Increased intestinal barrier permeability, leaky gut, has been reported in patients with autism. However, its contribution to the development of autism has not been determined. We selected dextran sulfate sodium (DSS) to disrupt and metformin to repair the intestinal barrier in BTBR T⁺tf/J autistic mice to test this hypothesis. DSS treatment resulted in a decreased affinity for social proximity; however, autistic behaviors in mice were improved after the administration of metformin. We found an increased affinity for social proximity/social memory and decreased repetitive and anxiety-related behaviors. The concentration of lipopolysaccharides in blood decreased after the administration of metformin. The expression levels of the key molecules in the toll-like receptor 4 (TLR4)-myeloid differentiation factor 88 (MyD88)-nuclear factor kappa B (NF-κB) pathway and their downstream inflammatory cytokines in the cerebral cortex were both repressed. Thus, "leaky gut" could be a trigger for the development of autism via activation of the lipopolysaccharide-mediated TLR4-MyD88-NF-κB pathway.
Mice ; Animals ; NF-kappa B ; Myeloid Differentiation Factor 88/metabolism* ; Lipopolysaccharides/pharmacology* ; Toll-Like Receptor 4/metabolism* ; Autistic Disorder/metabolism* ; Signal Transduction/physiology*

Objective: To investigate the efficacy of different bleaching methods on white-spot lesions of the enamel using optical coherence tomography and to evaluate its feasibility for monitoring the therapeutic effects on white-spot lesions. Methods: Forty-eight sound premolars extracted for orthodontic reasons were selected and cut for 4 mm×4 mm×2 mm enamel blocks in buccal surfaces of the crowns. The samples were covered with acid-resistant varnish (except for the buccal surfaces) and immersed in demineralization solution for 18 days to establish the white-spot lesion models of the enamels. Samples were randomly divided into four groups (n=12). Group A was given demineralization only. Specimens in Group B, C and D were treated with 40% hydrogen peroxide, resin infiltration and 40% hydrogen peroxide combined with resin infiltration, respectively. Eight samples in each group were randomly selected. OCT was applied to observe the optical changes of the enamel surface and according to the OCT scanning results, the demineralization depth of enamel samples in each group was calculated. Then, the enamel blocks were embedded in epoxy resins, except the buccal surfaces, and measured for the microhardness values of the enamel surface by a microindentation hardness tester. Four samples in each group were cut longitudinally, and the ultrastructural changes of enamel samples in each group were observed by scanning electron microscope. Results: OCT showed that the light scattering characteristics of enamel surface changed in all groups, and the bright layer was formed, but the thickness of bright layer in Group C and D was significantly lower than that in Group A and B (P<0.05). The microhardness values (kg/mm2) of the samples in Group A-D were (214.99±31.70), (250.66±33.64), (312.42±18.01) and(286.53±26.65), respectively. The microhardness of enamel surfaces in Group C and D was significantly higher than that in Group A and B (P<0.05), and the ultrastructure of enamel surfaces in Group C and D were more flat and dense in SEM observation (P<0.05). Conclusion The methods of resin infiltration therapy or 40% hydrogen peroxide combined with resin infiltration could effectively improve white-spot lesions of the enamel and the non-invasive OCT can be used as a better evaluation method for the diagnosis and treatment of white-spot lesions of the enamel.

Abstract: To investigate the value of whole-body dynamic18F-flurodeoxygluse positron emission tomography with computed tomography(18F-FDG PET/CT) Patlak imaging for evaluating the radiosensitization early efficacy in C6 rat glioma. Methods: Twenty-four C6 rat glioma models were established and randomly divided into control group, Oleanolic acid(OA) group, radiotherapy group and OA+radiotherapy group, with 6 rats in each group. 75 min whole-body dynamic18F-FDG PET/CT Patlak imaging was performed before and 48 h after treatment. The left ventricular time-activity curve(TAC) of tumor-bearing rats was used as input function for Patlak analysis to obtain Ki images. The last frame of the dynamic PET scan was selected for static analysis, and the changes of maximum standardized uptake value(SUVmax), Ki values and tumor volume were observed, and the survival time of tumor-bearing rats was recorded. Then, the tumor samples were dissected for Ki-67 and glucose transporter-1(GLUT-1) immunohistochemical staining. Results: Before treatment, there were no significant differences in SUVmaxand Ki values among the 4 groups(F=0.909, 0.858,P>0.05). 48 h after treatment, SUVmaxand Ki values of control group and OA group increased significantly(t=-10.697,-17.516,P<0.01;t=-19.321,-9.887,P<0.01), SUVmaxand Ki values of radiotherapy group and OA+radiotherapy group decreased(t=5.629, 6.916,P<0.01;t=7.241, 10.490,P<0.01), there was no significant difference in SUVmaxbetween radiotherapy group and OA+radiotherapy group 48 h after treatment(t=1.233,P=0.246), while the Ki values in OA+radiotherapy group was significantly lower than that in radiotherapy group, the difference was statistically significant(t=3.051,P<0.05). The immunohistochemistry staining results suggested that the expression of Ki-67 and GLUT-1 in OA+radiotherapy group was lower than that in the other 3 groups(F=63.887, 40.720,P<0.05). In addition, Ki values showed positive correlations with the expression of Ki-67 and GLUT-1(r=0.931, 0.936,P<0.01). Kaplan-Meier analysis showed that the survival time of tumor-bearing rat in OA+radiotherapy group was longer than that in the other three groups, and the difference was statistically significant(χ2=19.817,P<0.01). Conclusion OA has radiosensitization effect on C6 rat glioma. Whole-body dynamic18F-FDG PET/CT Patlak multi-parameter imaging can evaluate the radiosensitization early efficacy in C6 rat glioma, which is more sensitive than PET/CT static imaging.

Objective:To investigate the value of 18F-FDG total-body PET/CT dynamic imaging in evaluating the efficacy of early radiotherapy for MDA-MB-231 breast cancer bearing nude mice. Methods:MDA-MB-231 breast cancer bearing nude mice were established and divided into control group and radiotherapy group based on the random number table method ( n=10 for each group). 18F-FDG total-body PET/CT dynamic imaging was performed before and after the radiotherapy. The SUV max and the maximum tracer uptake net influx constant ( Kimax) of tumors, and the target-to-background ratio (TBR) were calculated. The change of tumor volume was recorded. The value of 18F-FDG total-body PET/CT dynamic imaging in evaluating the efficacy of radiotherapy was accessed using pathological findings as the reference. Paired t test, independent-sample t test and Pearson correlation analysis were used for data analysis. Results:After radiotherapy, SUV max and Kimax(4.66±0.46 and 0.14±0.03) were reduced in the radiotherapy group compared with those before radiotherapy (5.30±0.52 and 0.19±0.03; t values: 4.61, 8.31, P values: 0.001, <0.001), while the SUV max (5.94±0.74 vs 5.24±0.50) and Kimax (0.23±0.03 vs 0.19±0.02) were increased compared with baseline in the control group ( t values: 4.77, 6.87, P values: 0.001, <0.001). TBR Ki was significantly higher than TBR suv based on all images of the 2 groups (14.11±5.58 vs 5.91±1.60; t=8.92, P<0.001). The tumor volume in the radiotherapy group decreased compared with that before radiotherapy, but the difference was not statistically significant ((0.74±0.12) vs (0.81±0.08) cm 3; t=2.24, P=0.052). The results of immunohistochemistry showed that glucose transport protein (Glut)1 was highly expressed in tumors, and the Glut1 positive cell percentage of the radiotherapy group was significantly lower than that of the control group ((38.30±6.18)% vs (69.78±5.37)%; t=12.17, P<0.001). The expression of Glut1 was significantly positively correlated with SUV max and Kimax(the control group: rsuv=0.75, P=0.012; rKi=0.77, P=0.010; the radiotherapy group: rsuv=0.67, P=0.035; rKi=0.77, P=0.010). The apoptosis index (AI) of tumor cells in the radiotherapy group was higher than that in the control group ((24.15±4.00)% vs (10.15±3.05)%; t=8.85, P<0.001). Conclusion:18F-FDG total-body PET/CT dynamic imaging can sensitively monitor the effect of early radiotherapy in MDA-MB-231 breast cancer bearing nude mice.

Objective The purpose of this study was to evaluate whether whole-tumor histogram-based analysis of readout-segmented echo-planar diffusion-weighted imaging (RESOLVE-DWI) ADC map can help in the discrimination of parotid gland tumors(pleomorphic adenoma, Warthin tumor, malignant parotid gland tumor). Methods The MR images(pre / post-contrast enhanced MRI, RESOLVE-DWI, ADC map) of 47 patients with a biopsy-or surgery-proven pleomorphic adenomas, 25 patients with Warthin tumors and 36 patients with malignant parotid gland tumors were retrospectively analyzed. Histogram-based analysis was performed with the software MaZda. ROIs were drawn on every section of the ADC map containing the tumor, then 12 Parameters(Area, MinNorm, MaxNorm, Mean,Variance, Skewness, Kurtosis, Perc.1％, Perc.10％, Perc.50％, Perc.90％, Perc.99％) derived from histogram were calculated. Statistical analysis among the three groups (One-way ANOVA or Kruskal-Wallis test) were performed to find out the statistical significance of each histogram parameter. Then LSD test or Mann-Whitney U test was used for pairwise comparison between groups. And the differential efficiency of each parameter was determined using ROC analysis. Results Overall, 9 parameters (MinNorm, MaxNorm, Mean, Variance, Skewness, Perc. 10％, Perc. 50％, Perc. 90％, Perc. 99％) among three groups were shown to be statistically significant (P<0.05). Between the pleomorphic adenomas and Warthin tumors, these 9 parameters were of statistical significance. Perc.50％revealed the highest diagnostic efficiency, followed by Mean and Perc.10％. Between the pleomorphic adenomas and malignant parotid gland tumors, also all these 9 parameters were of statistical significance. Mean was revealed to have the highest diagnostic efficiency, followed by Perc. 10％ and Perc. 90％. However, between the Warthin tumors and malignant parotid gland tumors, only 5 parameters (MinNorm, Mean, Skewness, Perc. 10％, Perc. 50％) were statistically significant. MinNorm was revealed to have the highest diagnostic efficiency, followed by Perc. 50％ and Perc. 10％. Generally, Mean, Perc. 10％ and Perc. 50％ were more effective in the differential diagnosis of these three types common parotid neoplasms. Conclusion Whole-tumor histogram analysis of ADC maps are effective in differentiating common parotid neoplasms.

Objective: To evaluate the role of prognostic nutritional index (PNI) in the radiotherapy for elderly patients with esophageal squamous cell carcinoma. Methods: Clinical data of 108 elderly patients (aged>65 years) with esophageal squamous cell carcinoma who underwent radical radiotherapy were retrospectively analyzed. The PNI value of each patient was calculated, and the optimal cutoff value of PNI before treatment was determined by establishing the receiver operating characteristic curve (ROC curve). All patients were divided into the low and high PNI value groups. The overall survival rate was calculated by Kaplan-Meier method. Log-rank test, univariate and Cox’s multivariate prognosis analyses were performed. Results: The ROC curve demonstrated that the optimal cutoff value of PNI was 50.1(n=52 in high PNI group and n=56 in low PNI group). Age, gender and treatment did not significantly differ between two groups, whereas the TNM staging significantly differed (P=0.022). The effective rate of radiotherapy in the high PNI group was 96%, significantly higher than 73% in the low PNI group (P=0.001). In the high PNI group, the 1-, 2-, and 3-year overall survival rates were 94%, 69%, and 62%, significantly higher compared with 70%, 32% and 27% in the low PNI group (all P<0.001). Univariate analysis showed that PNI, T staging, N staging and TNM staging were significantly correlated the overall survival of patients (all P<0.01). Cox’s multivariate analysis revealed that N staging (RR=1.94, 95%CI=1.29-2.94, P=0.002) and PNI (RR=0.83, 95%CI=0.77-0.90, P<0.001) were independent risk factors affecting overall survival. Conclusions PNI before treatment has a good correlation with the prognosis and radiotherapy efficacy of patients, which can be used as a pivotal index to predict the clinical benefit of radiotherapy for elderly patients with esophageal squamous cell carcinoma.

Objective To study the value of whole tumors T2WI gray histogram analysis for differential diagnosis of three commonly presented pediatric posterior fossa tumors. Methods A retrospective analysis was conducted to compare brain MRI examination and pathology diagnosis of 133 cases of posterior fossa tumors in pediatric patients in our hospital.Among them, there were 49 cases of ependymoma(there were 25 males and 24 females), 36 cases of astrocytoma(WHO grade Ⅰ)(there were 19 males and 17 females), 48 cases of medulloblastoma(there were 36 males and 12 females),aged 1 to 14 years old, with an average (7±2) years old. Respectively draw the region of interest (ROI) in the T2 MR transaxial images of three groups on each layer of tumor level by using Mazda software and analyze the whole tumors gray histogram, including mean,variance, kurtosis,skewness, perc.01%,perc.10%,perc.50%,perc.90% and perc.99%.The histogram parameters of the three groups were not normally distributed. The Kruskal-Wallis test was used to compare the three groups. Bonferroni test was used to compare the two groups to find out statistical significance of each parameter. Results Through histogram analysis of 9 parameters, these 9 parameters were statistically significant (all P <0.05),including mean,variance,kurtosis,skewness, perc.01%,perc.10%,perc.50%,perc.90% and perc.99%. The sensitivity of skewness to differentiate ependymoma and the astrocytoma was 69.4％, the specificity was 77.8％, the area under the curve was 0.744, and the best cut-off value was-0.08. The sensitivity of kurtosis to differentiate astrocytoma and the medulloblastoma was 95.8％, the specificity was 97.2％ and the area under the curve was 0.972, the cut-off was 1.68. The sensitivity of kurtosis to differentiate ependymoma and the medulloblastoma was 91.7％, the specificity was 89.8％and the area under the curve was 0.932,the cut-off was 1.90.They had a high identification efficiency. Conclusions The T2WI gray histogram analysis based whole tumors is helpful for the dignosis of three types of pediatric posterior fossa tumors. Skewness has a high diagnostic efficiency to differentiate ependymoma and the astrocytoma. Kurtosis has a high diagnostic efficiency to differentiate ependymoma and the medulloblastoma. Kurtosis has a high diagnostic efficiency to differentiate ependymoma and medulloblastoma.