Objective To study the in vitro expression of three phenylalanine hydroxylase(PAH)mutants(p.R243Q,p.R241C,and p.Y356X)and determine their pathogenicity.Methods Bioinformatics techniques were used to predict the impact of PAH mutants on the structure and function of PAH protein.Corresponding mutant plasmids of PAH were constructed and expressed in HEK293T cells.Quantitative reverse transcription polymerase chain reaction was used to measure the mRNA expression levels of the three PAH mutants,and their protein levels were assessed using Western blot and enzyme-linked immunosorbent assay.Results Bioinformatics analysis predicted that all three mutants were pathogenic.The mRNA expression levels of the p.R243Q and p.R241C mutants in HEK293T cells were similar to the mRNA expression level of the wild-type control(P>0.05),while the mRNA expression level of the p.Y356X mutant significantly decreased(P<0.05).The PAH protein expression levels of all three mutants were significantly reduced compared to the wild-type control(P<0.05).The extracellular concentration of PAH protein was reduced in the p.R241C and p.Y356X mutants compared to the wild-type control(P<0.05),while there was no significant difference between the p.R243Q mutant and the wild type control(P>0.05).Conclusions p.R243Q,p.R241C and p.Y356X mutants lead to reduced expression levels of PAH protein in eukaryotic cells,with p.R241C and p.Y356X mutants also affecting the function of PAH protein.These three PAH mutants are to be pathogenic.[Chinese Journal of Contemporary Pediatrics,2024,26(2):188-193]

Zearalenone(ZEN) is a toxic metabolite produced by Fusarium culmorum, F. graminearum, F. tricinctum, and other fungi, with estrogenic characteristics. Exposure to or ingestion of ZEN during pregnancy can cause reproductive dysfunction, miscarriage, stillbirth, and malformation, and seriously endanger human life and health. The detection methods for ZEN in the Chinese Pharmacopoeia(2020 edition) are liquid chromatography(LC) and liquid chromatography-mass spectrometry(LC-MS), and it is stipulated that ZEN should not exceed 500 μg in 1 000 g of Coicis Semen. Although these detection methods by instruments can achieve the qualitative and quantitative analysis of ZEN in Coicis Semen, their high detection cost and long periods hinder the rapid screening of a large number of samples in the field. In this study, the synthesized ZEN hapten was conjugated with bovine serum albumin(BSA) and ovalbumin(OVA) to obtain the complete ZEN antigen. By virtue of antibody preparation techniques, ZEN monoclonal antibody 4F6 was prepared, which showed 177.5%, 137.1%, and 109.7% cross-reactivity with ZEN structural analogs zearalanol, zearalenone, and α-zearalenol, respectively, and no cross-reactivity with other fungal toxins such as aflatoxin. Direct competitive enzyme-linked immunosorbent assay(dcELISA) based on ZEN monoclonal antibody 4F6 was developed for the determination of ZEN in Coicis Semen with an IC_(50) of 1.3 μg·L~(-1) and a detection range of 0.22-21.92 μg·L~(-1). The recoveries were 83.91%-105.3% and the RSD was 4.4%-8.0%. The established dcELISA method was used to determine the ZEN residuals in nine batches of Coicis Semen samples, and the results were validated by LC-MS. The correlation between the two detection methods was found to be 0.993 9, indicating that the established dcELISA could be used for the rapid qualitative and quantitative detection of ZEN residuals in Coicis Semen.
Humans ; Female ; Pregnancy ; Zearalenone ; Coix ; Enzyme-Linked Immunosorbent Assay ; Mycotoxins ; Antibodies, Monoclonal

Objective: To further elucidate the clinical efficacy and safety of a combination regimen based on the BTK inhibitor zebutanil bridging CD19 Chimeric antigen receptor T cells (CAR-T cells) in the treatment of relapsed/refractory diffuse large B-cell lymphoma (r/r DLBCL) . Methods: Twenty-one patients with high-risk r/r DLBCL were treated with a zanubrutinib-based regimen bridging CAR-T between June 2020 and June 2023 at the Department of Hematology, Tongji Hospital, Tongji University and the Second Affiliated Hospital of Zhejiang University, and the efficacy and safety were retrospectively analyzed. Results: All 21 patients were enrolled, and the median age was 57 years (range: 38-76). Fourteen patients (66.7%) had an eastern cooperative oncology group performance status score (ECOG score) of ≥2. Eighteen patients (85.7%) had an international prognostic index (IPI) score of ≥3. Three patients (14.3%) had an IPI score of 2 but had extranodal infiltration. Fourteen patients (66.7%) had double-expression of DLBCL and seven (33.3%) had TP53 mutations. With a median follow-up of 24.8 (95% CI 17.0-31.6) months, the objective response rate was 81.0%, and 11 patients (52.4%) achieved complete remission. The median progression-free survival (PFS) was 12.8 months, and the median overall survival (OS) was not reached. The 1-year PFS rate was 52.4% (95% CI 29.8% -74.3%), and the 1-year OS rate was 80.1% (95% CI 58.1% -94.6%). Moreover, 18 patients (85.7%) had grade 1-2 cytokine-release syndrome, and two patients (9.5%) had grade 1 immune effector cell-associated neurotoxicity syndrome. Conclusion: Zanubrutinib-based combination bridging regimen of CAR-T therapy for r/r DLBCL has high efficacy and demonstrated a good safety profile.
Humans ; Middle Aged ; Receptors, Chimeric Antigen/therapeutic use* ; Retrospective Studies ; Immunotherapy, Adoptive/adverse effects* ; Lymphoma, Large B-Cell, Diffuse/drug therapy* ; Cell- and Tissue-Based Therapy ; Antigens, CD19/adverse effects*

Objective: This study systematically explore the efficacy and safety of fourth-generation chimeric antigen receptor T-cells (CAR-T), which express interleukin 7 (IL7) and chemokine C-C motif ligand 19 (CCL19) and target CD19, in relapsed or refractory large B-cell lymphoma. Methods: Our center applied autologous 7×19 CAR-T combined with tirelizumab to treat 11 patients with relapsed or refractory large B-cell lymphoma. The efficacy and adverse effects were explored. Results: All 11 enrolled patients completed autologous 7×19 CAR-T preparation and infusion. Nine patients completed the scheduled six sessions of tirolizumab treatment, one completed four sessions, and one completed one session. Furthermore, five cases (45.5%) achieved complete remission, and three cases (27.3%) achieved partial remission with an objective remission rate of 72.7%. Two cases were evaluated for disease progression, and one died two months after reinfusion because of uncontrollable disease. The median follow-up time was 31 (2-34) months, with a median overall survival not achieved and a median progression-free survival of 28 (1-34) months. Two patients with partial remission achieved complete remission at the 9th and 12th months of follow-up. Therefore, the best complete remission rate was 63.6%. Cytokine-release syndrome and immune effector cell-associated neurotoxicity syndrome were controllable, and no immune-related adverse reactions occurred. Conclusion: Autologous 7×19 CAR-T combined with tirelizumab for treating relapsed or refractory large B-cell lymphoma achieved good efficacy with controllable adverse reactions.
Humans ; Antibodies, Monoclonal/therapeutic use* ; Antigens, CD19 ; Chemokine CCL19 ; Immunotherapy, Adoptive ; Interleukin-7 ; Lymphoma, Large B-Cell, Diffuse/therapy* ; Programmed Cell Death 1 Receptor ; Receptors, Chimeric Antigen

Objective: To explore the heterogeneity and correlation of clinical phenotypes and genotypes in children with disorders of sex development (DSD). Methods: A retrospective study of 1 235 patients with clinically proposed DSD in 36 pediatric medical institutions across the country from January 2017 to May 2021. After capturing 277 DSD-related candidate genes, second-generation sequencing was performed to analyzed the heterogeneity and correlation combined with clinical phenotypes. Results: Among 1 235 children with clinically proposed DSD, 980 were males and 255 were females of social gender at the time of initial diagnosis with the age ranged from 1 day of age to 17.92 years. A total of 443 children with pathogenic variants were detected through molecular genetic studies, with a positive detection rate of 35.9%. The most common clinical phenotypes were micropenis (455 cases), hypospadias (321 cases), and cryptorchidism (172 cases) and common mutations detected were in SRD5A2 gene (80 cases), AR gene (53 cases) and CYP21A2 gene (44 cases). Among them, the SRD5A2 mutation is the most common in children with simple micropenis and simple hypospadias, while the AMH mutation is the most common in children with simple cryptorchidism. Conclusions: The SRD5A2 mutation is the most common genetic variant in Chinese children with DSD, and micropenis, cryptorchidism, and hypospadias are the most common clinical phenotypes. Molecular diagnosis can provide clues about the biological basis of DSD, and can also guide clinicians to perform specific clinical examinations. Target sequence capture probes and next-generation sequencing technology can provide effective and economical genetic diagnosis for children with DSD.
3-Oxo-5-alpha-Steroid 4-Dehydrogenase/genetics* ; Child ; China/epidemiology* ; Cryptorchidism/genetics* ; Disorders of Sex Development/genetics* ; Female ; Genital Diseases, Male ; Genotype ; Humans ; Hypospadias/genetics* ; Male ; Membrane Proteins/genetics* ; Penis/abnormalities* ; Phenotype ; Retrospective Studies ; Steroid 21-Hydroxylase/genetics*

ObjectiveTo evaluate the safety and comfort of hepatic artery infusion chemotherapy （HAIC） via distal transradial access （dTRA） in patients with liver cancer. MethodsPatients with advanced liver cancer who received HAIC via dTRA or transfemoral access （TFA） at the Department of Interventional Radiology， the First Affiliated Hospital of Sun Yat-sen University， were enrolled. The patients underwent dTRA or TFA for onetime and crossed-over subsequently. The patients received HAIC using FOLFOX4 regimen. Postcatheterization questionaire was used to compare the effects of the two vessel accesses on patients’ quality of life. Procedure-related adverse events were also recorded. ResultsAmong the 18 cases enrolled for HAIC， 9 underwent crossover from dTRA to TFA and the the remaining 9 from TFA to dTRA. During HAIC via dTRA， we only found grade 1 or grade 2 procedure-related adverse events such as 2 access site hematoma， 3 persistent pain at access site and 1 left palm numbness. No grade 3 or grade 4 procedure-related adverse event was found. Post dTRA ultrasound revealed no proximal radial artery occlusion. Significant difference in catheterization time between dTRA and TFA accesses was found （4 min vs. 3 min， P < 0.05）. All comfort scores were higher with dTRA as compared to TFA and patients preferred dTRA （7.89 vs. 2.72， P < 0.001）. The compression time for dTRA access was significantly shorter than TFA access （2 h vs. 7 h， P < 0.05）. ConclusionsdTRA approach is safe and tolerable， which is beneficial to improve the quality of life and comfort of patients with liver cancer when undergoing HAIC.

OBJECTIVE: To assess the inhibitory effect of the extract of Xanthoceras sorbifolium Bunge flower against benign prostatic hyperplasia (BPH) and explore its possible mechanism. METHODS: MTT assay was used to examine the effect of the extract of Xanthoceras sorbifolium Bunge flower on proliferation of benign prostatic hyperplasia cells (BPH-1), and cell apoptosis and cell cycle changes following the treatment were analyzed using annexin V/PI double staining and flow cytometry. The protein expression levels of Bcl-2, Bax, caspase-3, PI3K and AKT in the treated cells were detected using Western blotting. A rat model of BPH established by subcutaneous injection of testosterone propionate was treated with the flower extract for 28 days, and pathological changes in the prostate tissue were observed with HE staining. The protein expression levels of Bcl-2, Bax, caspase3 and PI3K/AKT in the prostate tissue were detected with Western blotting. RESULTS: Within the concentration range of 125-1000 µg/mL, the flower extract of Xanthoceras sorbifolium Bunge significantly inhibited the proliferation of BPH-1 cells and caused obvious cell cycle arrest at G0/G1 phase; the apoptotic rate of the cells was positively correlated with the concentration of the flower extract (P < 0.05). Bcl-2, p-PI3K and p-AKT expression levels were significantly down-regulated and Bax and caspase-3 expression levels were significantly increased in the cells after treatment with the flowers extract (P < 0.05). In the rat models of BPH, the rats treated with the flowers extract at moderate and high doses showed obviously decreased expressions of p-AKT and Bcl-2 and an increased expression of Bax in the prostate tissue; a significantly lowered p-AKT expression was observed in the prostate tissue of rats receiving the low-dose treatment (P < 0.05). CONCLUSION The flower extract of Xanthoceras sorbifolium Bunge has a inhibitory effect on BPH both in vitro and in rats, suggesting its potential value in the development of medicinal plant preparations for treatment of BPH.
Humans ; Male ; Rats ; Animals ; Prostatic Hyperplasia/drug therapy* ; Caspase 3 ; Phosphatidylinositol 3-Kinases/metabolism* ; bcl-2-Associated X Protein ; Proto-Oncogene Proteins c-akt ; Rats, Sprague-Dawley ; Plant Extracts/pharmacology* ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Apoptosis ; Flowers/metabolism* ; Sapindaceae/metabolism*

The "student-centered" flipped classroom teaching model can improve students' academic achievement, improve cognition, and cultivate innovation ability. However, it has obvious deficiencies in the integrity and systematization of knowledge as well as education. The traditional teaching concept based on " teacher-centered" has the unique advantages of systematization of knowledge learning and education. Therefore, we integrated the merits of these two different teaching models and introduced the semi-flipped classroom teaching model into the Biochemistry teaching of 2020 stomatology, pharmacy and preventive medicine majors in Cheeloo Medical College, Shandong University, compared with the traditional teaching of 2019 same majors. The data on the improvement of students ' academic achievement and self-cognition were analyzed. The results showed that the students' achievements of the semi-flipped classroom teaching model were better than those of the traditional teaching (P<0. 01). The students' self-cognitions were significantly improved after the implementation of semi-flipped classroom teaching (P<0. 01 or P<0. 05). This study provides a reference for the related teaching and research work in medical colleges.

Professor
Acupuncture ; Acupuncture Points ; Acupuncture Therapy ; Catgut ; Humans ; Panic Disorder/therapy* ; Perimenopause