OBJECTIVE: To explore the genetic etiology for a child presenting with motor retardation, language delay, intellectual disability, and dysmorphic features. METHODS: A child presented at Linyi People's Hospital in June 2022 was selected as the study subject. Clinical data of the child was collected. Peripheral blood samples were obtained from the child and her parents. Following extraction of genomic DNA, whole-exome sequencing (WES) was carried out. Candidate variant was validated by Sanger sequencing. Amniotic fluid samples were obtained from the mother's subsequent pregnancies for prenatal diagnosis. This study has been reviewed and approved by the Medical Ethics Committee of Linyi People's Hospital (Ethics No.: 2019-134). RESULTS: The proband was a 2-year-old girl showing developmental delays in motor, language, and intellectual domains, strabismus, hypertelorism, hearing impairment, obesity, and brachymesophalangy of the fifth finger. Magnetic resonance imaging revealed abnormalities of the white matter. Chromosomal microarray analysis (CMA) identified a 15q26.3 duplication (chr15:101562020_102060896 × 3) inherited from her mother. WES has uncovered a heterozygous c.1931A>G (p.Tyr644Cys) variant in the FBXO11 gene. Sanger sequencing confirmed the variant to be de novo in origin. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as likely pathogenic. Prenatal diagnosis revealed that the fetuses from the mother's second and third pregnancies did not harbor the same variant. CONCLUSION The c.1931A>G (p.Tyr644Cys) variant of the FBXO11 gene probably underlay the abnormal phenotype in the child. Based on its genotype and phenotype, the proband was diagnosed with Intellectual developmental disorder with dysmorphic facies and behavioral abnormalities.
Humans ; Female ; Intellectual Disability/genetics* ; Child, Preschool ; F-Box Proteins/genetics* ; Protein-Arginine N-Methyltransferases/genetics* ; Exome Sequencing

OBJECTIVE: To explore the clinical presentation and genetic etiology of a case with Xeroderma pigmentosum in conjunct with basal cell carcinoma and melanoma. METHODS: A male patient with Xeroderma pigmentosum treated at Xinxiang Central Hospital in October 2022 was selected as study subject. Whole exome sequencing (WES) was carried out. Candidate variants were verified by Sanger sequencing of his family members. This study was approved by the Ethics Committee of the hospital (Ethics No.: 2021-167). RESULTS: Magnetic resonance imaging showed that the patient has a solid soft tissue mass in the anterior and lower part of his right eyeball and a small nodule on the left nasal wing. Histopathological biopsy showed that the periocular tumor was basal cell carcinoma in conjunct with malignant melanoma, and the nasal wing tumor was basal cell carcinoma. WES and Sanger sequencing revealed that he has harbored compound heterozygous variants of the XPC gene, namely c.2391delT (p.F797Lfs*11) and IVS1+1G>A, which were inherited from his father and mother, respectively. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variants were rated as likely pathogenic (PVS1+PM2_Supporting+PM3) and pathogenic (PVS1+PM2_Supporting+PM3+PP5), respectively. The c.2391delT variant was unreported previously. Bioinformatic analysis suggests that it could significantly affect the tertiary structure of XPC protein. CONCLUSION The c.2391delT(p.F797Lfs*11) and IVS1+1G>A compound heterozygous variants probably underlay the pathogenesis in this patient. The detection of the novel variant has enriched the mutational spectrum of the XPC gene.
Humans ; Male ; Xeroderma Pigmentosum/genetics* ; Basal Cell Carcinoma/genetics* ; DNA-Binding Proteins/genetics* ; Melanoma/genetics* ; Mutation ; Skin Neoplasms/genetics* ; Middle Aged ; Exome Sequencing ; Pedigree

In order to establish a serological method for the detection of bovine parainfluenza virus type 3(BPIV3),the prokaryotic expression and purification of BPIV3 HN,NP,F,and P proteins were carried out,and the optimal protein-coated antigen was screened,and an indirect ELISA de-tection method was established.The results showed that the four recombinant proteins of BPIV3,rHN,rNP,rF,and rP were expressed,and the checkerboard titration results showed that rHN pro-tein had the highest P/N value as the coating protein,so it was used for the subsequent method es-tablishment.The optimal reaction conditions for indirect ELISA were found to be:the mass con-centration of the antigen coating was 0.5 mg/L,37 ℃ 1.5 h,5％skim milk,overnight blocking at 4 ℃,serum dilution at 1∶50,incubation at 37 ℃ 1 h,secondary antibody dilution at 1∶10 000 and incubation at 37℃ 0.5 h,substrate reaction conditions were 37℃ for 12 min.The results of speci-ficity experiments showed that the established method could specifically identify BPIV3 antibody-positive serum with a sensitivity of 1∶800,and the coefficient of variation in the detection of intra-and inter-assay repeatability was less than 10％,and the overall coincidence rate of the same batch of samples detected with the SVANOVIR kit was 92.22％.This method was used to detect 192 se-rum samples in Hebei Province,and the positive rate of BPIV3 antibody in serum was 66.15％.The indirect ELISA detection method of BP1V3 antibody constructed in this study is suitable for large-scale clinical serological investigations,and provides valuable data support for the research and de-velopment of BPIV3 antigen and antibody detection kits in China.

In order to investigate the role of sRNA OxyS in the pathogenicity of Salmonella typhi-murium infection,the OxyS gene deletion strain ATCC25241 △OxyS and the back-complemented strain ATCC25241 △OxyS/OxyS of Salmonella typhimurium ATCC25241 were constructed by using λRed homologous recombination technique.We investigated the effect of OxyS deletion on the biological characteristics and pathogenicity of Salmonella typhimurium ATCC25241.The re-sults showed that the deletion of OxyS did not affect the growth rate,the ability of biofilm forma-tion,and the ability of adhesion,invasion and intracellular survival of Salmonella typhimurium,but significantly reduced the motility of Salmonella typhimurium as well as its ability to survive in alkaline and oxidative environments.The results of mouse infection test showed that OxyS dele-tion caused a significant decrease in the virulence of Salmonella typhimurium in mice,and toxicity is reduced obviously.The qPCR results also showed that OxyS deletion could lead to changes in the transcript levels of a number of virulence-related genes of Salmonella typhimurium such as pipB,orf245,csgA,invH,tatA,sipA,sipB,and so on.The above results indicate that OxyS gene affects the biological characteristics and pathogenicity of Salmonella typhimurium and is an important virulence regulator of Salmonella typhimurium.

Objective To investigate the prognostic value of peripheral blood neutrophil-lymphocyte ratio(NLR)and thioredoxin reductase(TrxR)in patients with ovarian cancer receiving immunotherapy.Methods A total of 109 patients with advanced ovarian cancer treated in the Tumor Hospital Affiliated to Nantong University from January 2021 to December 2021 were selected as the research objects.The levels of NLR and TrxR in peripheral blood before immunotherapy were detected,and the evaluation value of NLR and TrxR on short-term efficacy,progression-free survival(PFS)and overall survival(OS)in ovarian cancer pa-tients receiving immunotherapy was explored.Results The optimal cut-off values of TrxR and NLR were 4.97 U/mL and 2.49%,respectively.According to the optimal cut-off value of TrxR and NLR,the patients were divided into the high level of TrxR group(69 cases,≥4.97 U/mL)and the low level of TrxR group(40 cases,<4.97 U/mL),the high level of NLR group(72 cases,≥2.49%)and the low level of NLR group(37 cases,<2.49%).The objective response rate(ORR)of the high level of NLR group was lower than that of the low level NLR group(P<0.05),and the disease progression rate(DPR)was higher than that of the low NLR group(P<0.05).The high level of TrxR group had a significantly lower ORR and a significantly higher DPR than the low level of TrxR group(P<0.05).The median PFS and OS of the high level of NLR group were 15.0 months and 16.0 months,respectively.The median PFS and OS of the low level of NLR group were 19.0 months and 21.0 months,respectively.The median PFS and OS of the high level of TrxR group were 15.0 months and 17.0 months,respectively.The median PFS was 18.0 months and the median OS was 21.0 months in the low level of TrxR group.NLR and TrxR were the influencing factors of PFS and OS in pa-tients with ovarian cancer immunotherapy(P<0.05).Conclusion The levels of NLR and TrxR in peripheral blood can be used as important prognostic indicators for advanced ovarian cancer patients receiving immuno-therapy.The lower the levels of NLR and TrxR,the better the prognosis of ovarian cancer patients.

Objective:To explore the genetic etiology for a child presenting with motor retardation, language delay, intellectual disability, and dysmorphic features.Methods:A child presented at Linyi People′s Hospital in June 2022 was selected as the study subject. Clinical data of the child was collected. Peripheral blood samples were obtained from the child and her parents. Following extraction of genomic DNA, whole-exome sequencing (WES) was carried out. Candidate variant was validated by Sanger sequencing. Amniotic fluid samples were obtained from the mother′s subsequent pregnancies for prenatal diagnosis. This study has been reviewed and approved by the Medical Ethics Committee of Linyi People′s Hospital (Ethics No.: 202402-H-034).Results:The proband was a 2-year-old girl showing developmental delays in motor, language, and intellectual domains, strabismus, hypertelorism, hearing impairment, obesity, and brachymesophalangy of the fifth finger. Magnetic resonance imaging revealed abnormalities of the white matter. Chromosomal microarray analysis (CMA) identified a 15q26.3 duplication (chr15: 101562020_102060896×3) inherited from her mother. WES has uncovered a heterozygous c. 1931A>G (p.Tyr644Cys) variant in the FBXO11 gene. Sanger sequencing confirmed the variant to be de novo in origin. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as likely pathogenic. Prenatal diagnosis revealed that the fetuses from the mother′s second and third pregnancies did not harbor the same variant. Conclusion:The c. 1931A>G (p.Tyr644Cys) variant of the FBXO11 gene probably underlay the abnormal phenotype in the child. Based on its genotype and phenotype, the proband was diagnosed with Intellectual developmental disorder with dysmorphic facies and behavioral abnormalities.

Objective:To investigate the effects of Jianpi Rougan Xifeng Decoction on the behaviors and the Ca 2+ -CaM-CaMK Ⅱ signaling pathway in the striatum of rats with tic disorder models. Methods:Seventy-two SPF-grade SD male rats were randomly divided into blank group, model group, tiapride group (15.93 mg/kg, intragastric administration), low-dose (4.32 g/kg, intragastric administration), medium-dose (8.64 g/kg, intragastric administration) and high-dose (17.28 g/kg, intragastric administration) Jianpi Rougan Xifeng Decoction groups, with 12 rats in each group. Rats in the blank group and model group were gavaged with 0.5 mg/kg of distilled water while rats in other 4 groups were gavaged with corresponding drugs, all rats were gavaged once a day for 28 days.The evaluations of motor behavior and stereotyped behavior were conducted using the Kadasah scoring method and the Diamond scoring method. Calcium content in the striatum was detected using a calcium assay kit. Immunohistochemical analysis was employed to detect the expression of dopamine transporter (DAT) and inositol 1, 4, 5-triphosphate (IP3) in the striatum of rats. Western blot was used to assess the expression of calmodulin (CaM) and calcium/calmodulin-dependent protein kinase Ⅱ (CaMK Ⅱ) in the striatum. RT-PCR was utilized to detect the expression of CaM and CaMK Ⅱ mRNA in the striatum. All data were processed using SPSS 26.0 software, and comparisons among multiple groups were performed using one-way ANOVA and Kruskal-Wallis test.Results:(1) After four weeks of pharmacological intervention, statistically significant differences were observed in the locomotor activity scores and stereotyped scores among the six groups of rats ( H=41.20, 44.24, both P<0.01). Specifically, the locomotor activity scores(3.00(3.00, 3.25), 1.00(0.75, 1.25), 1.00(0.75, 2.00), 1.00(0, 1.00)) and stereotyped scores(3.00(3.00, 4.00), 1.00(0.75, 2.00), 2.00(0.75, 2.00), 1.00(0, 1.25)) in the tiapride group and the medium- and high-dose Jianpi Rougan Xifeng Decoction groups were significantly lower than those in the model group (all P<0.01). (2) Results from the calcium assay kit revealed statistically significant differences in striatal calcium content among the six groups of rats ( F=146.67, P<0.01). The calcium content in the tiapride group and the medium- and high-dose Jianpi Rougan Xifeng Decoction groups was significantly lower than that in the model group (all P<0.01). Additionally, the calcium content in the medium-dose ((0.40±0.02)mmol/g) and high-dose ((0.30±0.03)mmol/g) Jianpi Rougan Xifeng Decoction groups was lower than that in the low-dose group ((0.48±0.02)mmol/g) (both P<0.01). (3) Immunohistochemical results showed that there were statistically significant differences in the mean optical density values of DAT and IP3 in the striatum among the six groups of rats ( F=25.57, 154.98, both P<0.01). The IP3 mean optical density in the tiapride group and the Jianpi Rougan Xifeng Decoction groups with medium and high doses exhibited lower values compared to the model group (all P<0.05), whereas the DAT mean optical density displayed higher values in these groups compared to the model group (all P<0.05). The low-dose Jianpi Rougan Xifeng Decoction group also exhibited a lower optical density value of IP3 compared to the model group( P<0.05). The optical density values of IP3 (2.68±0.21, 2.40±0.22) in the medium-dose and high-dose Jianpi Rougan Xifeng Decoction groups were lower than that in the low-dose group (4.27±0.23) (both P<0.01). (4) Western blot results indicated that there were statistically significant differences in the protein expression levels of CaM and CaMK Ⅱ in the striatum among the six groups of rats ( F=233.03, 118.60, both P<0.01). The protein expression levels of CaM and CaMK Ⅱ in the tiapride group and the low-dose, medium-dose, and high-dose Jianpi Rougan Xifeng Decoction groups were lower than those in the model group (all P<0.01). The protein expression levels of CaM (1.02±0.06, 0.84±0.02) and CaMK Ⅱ (0.48±0.03, 0.40±0.02) in the medium-dose and high-dose Jianpi Rougan Xifeng Decoction groups were lower than those in the low-dose group (1.21±0.03, 0.57±0.02)) (all P<0.05). Additionally, the protein expression level of CaM in the high-dose Jianpi Rougan Xifeng Decoction group was lower than that in the medium-dose group( P<0.05).(5) The RT-PCR results indicated significant variations in the mRNA expression levels of CaM and CaMK Ⅱ within the striatum across the six groups rats ( F=30.54, 20.78, both P<0.01). The mRNA expression levels of CaM and CaMK Ⅱ in the tiapride group and the high-dose Jianpi Rougan Xifeng Decoction group were lower than those in the model group (both P<0.05). The mRNA expression levels of CaMK Ⅱ (1.38±0.17) in the high-dose Jianpi Rougan Xifeng Decoction group were lower than those in the low-dose group (1.99±0.27) ( P<0.01). Conclusion:Jianpi Rougan Xifeng Decoction can improve locomotor activity and stereotyped behavior in rats with tic disorder model. The mechanism may be through inhibiting the Ca 2+ -CaM-CaMK Ⅱ signaling pathway and regulating the expression of DAT and IP3, thereby modulating the release and recovery of dopamine and reducing the occurrence of tic symptoms.

In order to establish a serological method for the detection of bovine respiratory syncytial virus,the prokaryotic expression of four proteins of BRSV,G,F,P,and M was carried out,and the most suitable coating antigen was screened to establish an indirect ELISA detection method.The results showed that the four recombinant proteins of BRSV,rG,rF,rP and rM were successfully expressed.The results of checkerboard screening showed that the P/N value of rG protein was the largest,which was determined to be the best coating antigen established by indirect ELISA meth-od.The optimal reaction conditions for indirect ELISA were as follows:the mass concentration of rG protein coating was 1 mg/L,37℃ for 2 h;3％BSA 37℃ block for 1 h;Serum was diluted 1∶50 and incubated at 37℃ for 1h;Secondary antibody 1∶5 000 dilution,37℃ for 30min;The color development conditions of the substrate were 37℃ for 15 min;Thirty negative sera were selected,and the cut-off value was determined to be 0.63 by the established indirect ELISA method.The re-sults of the specificity test showed that the indirect ELISA method established in this test only recognized BRSV-positive serum,and did not react with IBRV,BCoV,and BPIV3-positive serum.The results of repeatability test showed that the method had good repeatability,and the coefficient of variation within and between batches was less than 10％.The results of the sensitivity test showed that the BRSV-positive serum was still positive when diluted to 1∶8 192.The indirect ELISA method established in this experiment was used to detect 100 clinical serum samples at the same time,and the total coincidence rate of the two reached 90.48％,the positive coincidence rate was 93.42％,and the negative coincidence rate was 82.75％.The indirect ELISA established in this test can be used for the detection of bovine respiratory syncytial virus in clinical practice.

Objective:To investigate the mechanisms by which calpain small subunit 1(CAPN4)regulates cisplatin resistance and cancer stem cell(CSC)stemness in lung adenocarcinoma,and to provide experimental evidence for reversing drug resistance through targeting CSC stemness.Methods:Tissue samples were collected from 10 lung adenocarcinoma patients who underwent surgical resection at Fujian Cancer Hospital from January 2023 to January 2024.Immunohistochemistry(IHC)stining was used to detect the differential expression of CAPN4 in five cisplatin-resistant and five cisplatin-sensitive lung adenocarcinoma tissues,followed with a histological scoring(H-score).CAPN4 gene expression-related survival analysis in lung cancer patients was conducted using The Cancer Genome Atlas(TCGA)database and Gene Expression Profiling Interactive Analysis(GEPIA)platform.Additionally,tissue samples from two cisplatin-resistant and two cisplatin-sensitive lung adenocarcinoma cases were collected to establish lung adenocarcinoma organoid(PDO)models.H-E and IHC staining were used to assess the morphological consistency between PDOs and the primary tumors.CAPN4 gene expression was silenced using lentivirus-mediated shRNA transduction.The expression levels of stem cell markers ALDH1A1,CD133,Nanog,and SOX9 were detected at both the gene and protein levels using quantitative polymerase chain reaction(qPCR)and Western blotting(WB),respectively.The sensitivity of CAPN4-knockdown PDOs to cisplatin was evaluated using the adenosine triphosphate(ATP)assay,and the apoptosis was assessed using the caspase-3 assay.Results:IHC results showed that CAPN4 protein expression was significantly upregulated in cisplatin-resistant lung adenocarcinoma tissues(P<0.05).TCGA cohort analysis revealed that high CAPN4 expression was significantly associated with poor prognosis(reduced OS)in lung adenocarcinoma patients(HR=1.4,P<0.05).PDOs derived from cisplatin-resistant patients exhibited significant upregulation in CAPN4 protein and stemness markers at both gene and protein levels(all P<0.05).Cisplatin sensitivity assays demonstrated that PDOs derived from cisplatin-resistant patients had significantly higher IC50 values than those from cisplatin-sensitive patients(P<0.05).After CAPN4 knockdown,the expression of stem cell makers in PDOs derived from cisplatin-resistant patients were significantly reduced,and their sensitivity to cisplatin was enhanced(P<0.05).Conclusion:Knockdown of CAPN4 reduces stem cell marker expression and enhances cisplatin sensitivity in lung adenocarcinoma PDOs,providing a potential therapeutic target for reversing cisplatin resistance in lung cancer.

Objective:To investigate the effects of Jianpi Rougan Xifeng Decoction on the behaviors and the Ca 2+ -CaM-CaMK Ⅱ signaling pathway in the striatum of rats with tic disorder models. Methods:Seventy-two SPF-grade SD male rats were randomly divided into blank group, model group, tiapride group (15.93 mg/kg, intragastric administration), low-dose (4.32 g/kg, intragastric administration), medium-dose (8.64 g/kg, intragastric administration) and high-dose (17.28 g/kg, intragastric administration) Jianpi Rougan Xifeng Decoction groups, with 12 rats in each group. Rats in the blank group and model group were gavaged with 0.5 mg/kg of distilled water while rats in other 4 groups were gavaged with corresponding drugs, all rats were gavaged once a day for 28 days.The evaluations of motor behavior and stereotyped behavior were conducted using the Kadasah scoring method and the Diamond scoring method. Calcium content in the striatum was detected using a calcium assay kit. Immunohistochemical analysis was employed to detect the expression of dopamine transporter (DAT) and inositol 1, 4, 5-triphosphate (IP3) in the striatum of rats. Western blot was used to assess the expression of calmodulin (CaM) and calcium/calmodulin-dependent protein kinase Ⅱ (CaMK Ⅱ) in the striatum. RT-PCR was utilized to detect the expression of CaM and CaMK Ⅱ mRNA in the striatum. All data were processed using SPSS 26.0 software, and comparisons among multiple groups were performed using one-way ANOVA and Kruskal-Wallis test.Results:(1) After four weeks of pharmacological intervention, statistically significant differences were observed in the locomotor activity scores and stereotyped scores among the six groups of rats ( H=41.20, 44.24, both P<0.01). Specifically, the locomotor activity scores(3.00(3.00, 3.25), 1.00(0.75, 1.25), 1.00(0.75, 2.00), 1.00(0, 1.00)) and stereotyped scores(3.00(3.00, 4.00), 1.00(0.75, 2.00), 2.00(0.75, 2.00), 1.00(0, 1.25)) in the tiapride group and the medium- and high-dose Jianpi Rougan Xifeng Decoction groups were significantly lower than those in the model group (all P<0.01). (2) Results from the calcium assay kit revealed statistically significant differences in striatal calcium content among the six groups of rats ( F=146.67, P<0.01). The calcium content in the tiapride group and the medium- and high-dose Jianpi Rougan Xifeng Decoction groups was significantly lower than that in the model group (all P<0.01). Additionally, the calcium content in the medium-dose ((0.40±0.02)mmol/g) and high-dose ((0.30±0.03)mmol/g) Jianpi Rougan Xifeng Decoction groups was lower than that in the low-dose group ((0.48±0.02)mmol/g) (both P<0.01). (3) Immunohistochemical results showed that there were statistically significant differences in the mean optical density values of DAT and IP3 in the striatum among the six groups of rats ( F=25.57, 154.98, both P<0.01). The IP3 mean optical density in the tiapride group and the Jianpi Rougan Xifeng Decoction groups with medium and high doses exhibited lower values compared to the model group (all P<0.05), whereas the DAT mean optical density displayed higher values in these groups compared to the model group (all P<0.05). The low-dose Jianpi Rougan Xifeng Decoction group also exhibited a lower optical density value of IP3 compared to the model group( P<0.05). The optical density values of IP3 (2.68±0.21, 2.40±0.22) in the medium-dose and high-dose Jianpi Rougan Xifeng Decoction groups were lower than that in the low-dose group (4.27±0.23) (both P<0.01). (4) Western blot results indicated that there were statistically significant differences in the protein expression levels of CaM and CaMK Ⅱ in the striatum among the six groups of rats ( F=233.03, 118.60, both P<0.01). The protein expression levels of CaM and CaMK Ⅱ in the tiapride group and the low-dose, medium-dose, and high-dose Jianpi Rougan Xifeng Decoction groups were lower than those in the model group (all P<0.01). The protein expression levels of CaM (1.02±0.06, 0.84±0.02) and CaMK Ⅱ (0.48±0.03, 0.40±0.02) in the medium-dose and high-dose Jianpi Rougan Xifeng Decoction groups were lower than those in the low-dose group (1.21±0.03, 0.57±0.02)) (all P<0.05). Additionally, the protein expression level of CaM in the high-dose Jianpi Rougan Xifeng Decoction group was lower than that in the medium-dose group( P<0.05).(5) The RT-PCR results indicated significant variations in the mRNA expression levels of CaM and CaMK Ⅱ within the striatum across the six groups rats ( F=30.54, 20.78, both P<0.01). The mRNA expression levels of CaM and CaMK Ⅱ in the tiapride group and the high-dose Jianpi Rougan Xifeng Decoction group were lower than those in the model group (both P<0.05). The mRNA expression levels of CaMK Ⅱ (1.38±0.17) in the high-dose Jianpi Rougan Xifeng Decoction group were lower than those in the low-dose group (1.99±0.27) ( P<0.01). Conclusion:Jianpi Rougan Xifeng Decoction can improve locomotor activity and stereotyped behavior in rats with tic disorder model. The mechanism may be through inhibiting the Ca 2+ -CaM-CaMK Ⅱ signaling pathway and regulating the expression of DAT and IP3, thereby modulating the release and recovery of dopamine and reducing the occurrence of tic symptoms.