Objective Based on real-time PCR technique and ring primers,to establish a simple,accurate,cost-effective and easily standardized quantitative assay for quantification of HER2 mRNA,and apply to provide medication guidance for clinical tumor personalized molecular targeted therapy.Methods Screening reference gene which was stable expression in breast cancer,and optimizing the PCR reaction system.Then a real-time PCR with Eva Green for quantification of the mRNA expression levels of HER2 gene was developed.The specificity,sensitivity and reproducibility of the method were evaluated 87 specimens including 55 liquid nitrogen-frozen breast cancer tissues and 32 normal tissues were detected by the real-time quantitative reverse transcription (FQ RT)-PCR and immunohistochemistry(IHC).Results The standard curve of the method indicated a good linear relationship between the Ct value and the template concentration with the correlation coefficient being 0.997.The linear range of the system was from 101 to 106 copies/μl and the lower detection limit was 101 copies/μl.It had a high sensitivity and good specificity.The inter-assay coefficients of variation of HER and RPL37A genes were (5.93 ± 0.57)％ and (5.11 ± 0.59)％,(2.49 ±0.81)％ and (2.98 ±0.97)％ respectively.The intra-assay coefficients of variation were (5.76 ±0.58)％and (7.71 ±0.61)％,(3.75 ±0.76)％ and (4.40 ±0.96)％ respectively.Using the optimized FQ RTPCR system,HER2 gene of 87 specimens was quantificated.The sensitivity of the assay was 96.36％ (53/55),the specificity was 78.13％ (25/32),the positive predictive value was 88.33％ (53/60),the negative predictive value was 92.59％ (25/27),and the total coincidence rate between FQ RT-PCR and IHC was 89.66％ (78/87).The correlation of the results between the FQ RT-PCR and IHC was good (Kappa =0.770,P ＞ 0.05).Conclusions The method can quantify the mRNA expression levels of HER2 gene rapidly and cost-effictively with high sensitivity and specificity.It can be applied to clinical molecular diagnosis with attractive prospect.

Purpose Sustained attention dysfunction is a common symptom of patient with attention-deficit/hyperactivity disorder (ADHD).To reveal the neural mechanism of the abnormality of sustained attention of patients with ADHD,the cerebral blood flow (CBF) abnormalities in patients were studied by perfusion imaging.Materials and Methods Sixteen children with ADHD and twenty normal controls treated at the 401st Hospital of PLA from June 2013 to September 2015 underwent 3D arterial spin labeling (3D-ASL)scanning with GE 3.0T MRI scanner.The participants were performed four continuous sections of sustained attention to response task (SART) for 20 minutes in the scanner.Using SPM 8 toolkit,the local CBF values of both groups were compared in a voxel-wise manner,and their correlations with response time and target accuracy of SART were analyzed.Results When performing the SART,the patients with ADHD showed significantly inhibited trend of increasing CBF in the anterior cortex like dorsal cortex,medial prefrontal cortex,and motor area;however,they presented enhanced trend of increasing CBF in the posterior cortex such as posterior cingulate cortex and parietooccipital sulcus (P＜0.01);the change of CBF in the dorsal prefrontal cortex and that in the precentral and postcentral gyrus had significant correlation with response time of SART task and targeting ratio (dorsal prefrontal cortex:r=0.745,P＜0.001;r=0.591,P＜0.001;r=-0.521,P＜0.001.Precentral and postcentral gyrus:r=0.579,P＜0.001).Conclusion Patients with ADHD show different CBF redistribution between anterior and posterior cerebral cortex in performing SART,and the abnormal CBF shows significant correlations with behavioral metrics,which reflects the mechanism of sustained attention dysfunction of patients with ADHD.

Objective To investigate the characteristics of adverse drug event (ADE) related to tacrolimus (Tac) in pediatric solid organ transplant recipients. Methods The data were retrieved from the US Food and Drug Administration Adverse Event Reporting System database from the first quarter of 2004 to the second quarter of 2023. The ADE data of pediatric organ transplant recipients with Tac as the primary suspected drug were extracted. The relationship between Tac and ADE was quantitatively analyzed by proportional imbalance method. Basic characteristics and signal strength of ADE related to Tac were analyzed. ADE related to Tac in children of different ages and different types of organ transplantation were analyzed. Results A total of 1 443 children's ADE reports involving Tac were screened, including 188 cases (13.0%) of heart transplantation, 668 cases (46.3%) of liver transplantation, 531 cases (36.8%) of kidney transplantation and 56 cases (3.9%) of lung transplantation. The median age of children was 10 years old. The top three countries with ADE reporting were the United States, France and the United Kingdom. China reported 26 cases, accounting for 1.8%. Infection and infectious diseases accounted for the highest proportion (20.96%) in ADE related to Tac, including EB virus and cytomegalovirus infection, etc. Infection and infectious diseases occupied the largest proportion of ADE related to Tac in children of different ages, whereas the pathogen types were different. Rejection, unstable immunosuppression level and renal function damage were also common ADE related to Tac in children of all ages. Nervous system disease was the main ADE in heart transplant recipients, while infection and infectious diseases were more common in liver and kidney transplant recipients. Rejection was the most common ADE in lung transplant recipients. Conclusions ADE related to Tac possess different distribution characteristics in different types of organ transplantation. Extensive attention should be paid to individualized drug monitoring and risk assessment in pediatric organ transplant recipients, thereby optimizing Tac treatment and reducing the risk of ADE.

Objective To explore the molecular mechanisms and cell-cell interactions in the injury process of pancreatic islet transplantation. Methods Single-cell transcriptome data from mouse islets treated with inflammatory factors were used, and data processing was performed using the Seurat package, with integrated data to remove batch effects. Cell subpopulations were annotated based on known markers. Cell-cell interactions in the inflammatory factor-treated group were analyzed using the CellChat package, and inferred based on the expression of cell surface receptors and ligands. Gene set enrichment analysis was used to clarify the biological processes enriched in β-cells after treatment with inflammatory factors. Finally, differentially expressed transcription factors were identified and verified using microarray datasets of donor islet ischemic injury and Western blotting. Results A total of 7 different cell subpopulations were found in mouse islets, with β-cells being the most abundant. Cell-cell interaction network analysis showed that the number and strength of interactions between ductal cells and other cells were the highest. Gene set enrichment analysis showed that after treatment with inflammatory factors, the immune response was positively enriched in β-cells, while peptide hormone metabolism, bile acid metabolism, and ion homeostasis were downregulated. The common differential transcription factors identified in the mouse single-cell transcriptome and the microarray dataset of donor islet ischemic injury were early growth response 1 (EGR1), nuclear factor-κB inhibitor α (NFKBIA), and activating transcription factor 3 (ATF3). Among them, NFKBIA and ATF3 were upregulated, while EGR1 was downregulated. The expression of EGR1 protein was downregulated after 24 h, 48 h, and 72 h of cold ischemia. Conclusions EGR1 is a transcription factor closely related to islet cold ischemia, and future research should focus on the specific mechanisms of EGR1 and its downstream target genes, in order to provide more effective strategies for clinical treatment of islet transplantation.