Objective To explore the relationship between the oral health status with the severity and outcome of pediatric bronchopneumonia.Methods A total of 150 children patients with bronchopneumonia admitted and treated in this hospital from January 2020 to August 2022 were selected as the observation group(n=150).According to the severity of the illness,the children patients were divided into the mild group(n=97)and severe group(n=53).In addition,150 children undergoing physical examination during the same pe-riod were selected as the control group(n=150).The changes in the decay missing filling surface(DMFS),simplified oral hygiene index(OHI-s)and plaque index(PLI)were analyzed.Spearman was used to analyze the relationship between the oral health status and the severity of bronchopneumonia.After one month of treatment,the changes in oral health status were analyzed under different outcome conditions.The receiver op-erating characteristic(ROC)curve was drawn to assess the predictive effect of the oral health status on the outcome of pediatric bronchopneumonia.Results The DMFS,OHI-s scores and PLI scores in the observation group were higher than those in the control group(P＜0.05).The DMFS,OHI-s score and PLIscores in the severe group were higher than those in the mild group(P＜0.05).Spearman analysis found that DMFS,OHI-s and PLI were positively correlated with the severity(r=0.441,0.253,0.328,P＜0.05).The levels of DMFS,OHI-s and PLI in the poor prognosis group were higher than those in the good prognosis group(P＜0.05).The area under the curve(AUC)of DMFS,OHI-s and PLI combination in predicting the outcome of bronchopneumonia was 0.851(0.703～0.999),with a sensitivity of 83.78％and a specificity of 92.92％.Con-clusion The oral health status is closely related to the severity of pediatric bronchopneumonia,which can be used to assess the outcome of the children patients.

Objective To explore the potential role and underlying mechanism of the functionally uncharacterized gene Gm49394 on regulating β-cell apoptosis under diabetic conditions.Methods The expression and translational activity of Gm49394 in pancreatic β-cell lines and non-β-cell lines were validated using RNA fluorescence in situ hybridization(RNA-FISH),quantitative real-time PCR(qPCR),Western blotting,and immunofluorescence(IF)assay.The β-cell lines(NIT-1/Min6)with Gm49394 overexpression or knockdown were constructed.The proliferation,apoptosis,mitochondrial function,as well as oxidative stress and endoplasmic reticulum stress markers in these β-cell lines under physiological homeostasis or pathological stress conditions,such as high glucose(30 mmol/L),inflammation(10 ng/mL IFN-γ alone or combined with 10 ng/mL IL-6),and hydrogen peroxide(100 μmol/L H2O2)were detected by flow cytometry and Western blotting.Results RNA-FISH and qPCR indicated that Gm49394 was specifically expressed in pancreatic β-cell lines and up-regulated under high glucose or inflammatory stimulation.IF assay and Western blotting showed that Gm49394 had protein-coding activity.Flow cytometry and Western blotting identified that Gm49394 overexpression did not affect β-cell proliferation,but promoted β-cell apoptosis and increased reactive oxygen species(ROS)and mitochondrial superoxide(MitoSOX)levels in β cells under physiological homeostasis or pathological stress conditions(P<0.05).Under physiological conditions,Gm49394 knockdown failed to induce significant alterations on β-cell apoptosis,ROS,or MitoSOX levels.Under pathological stress conditions,Gm49394 knockdown significantly suppressed β-cell proliferation,apoptosis,as well as oxidative and endoplasmic reticulum stress(P<0.05).Conclusion Gm49394 may promote β-cell apoptosis via oxidative stress and endoplasmic reticulum stress.

Objective To investigate the expression of Cyclin O(CCNO)in cervical cancer and its molecular mechanism in the progression of cervical cancer.Methods The pathological sections of 60 patients with cervical cancer in the Department of Oncology and Gynecology of the First Affiliated Hospital of Bengbu Medical University from January 2018 to December 2023 were collected,and the expression of CCNO in cervical cancer was detected by immunohistochemistry.The cells were divided into Vector group,CCNO group,siNC group and siCCNO group by transfection technology.CCK-8 assay and colony formation assay were used to evaluate the ability of CCNO to affect cell proliferation.Transwell assay and wound healing assay were used to evaluate the effect of CCNO on cell invasion and migration.Glycolysis assay and Western blot assay were used to evaluate the effect and mechanism of CCNO on glycolysis of cervical cancer cells.Results The results of immunohistochemistry and WB showed that CCNO was highly expressed in cervical cancer(P<0.001).Overexpression of CCNO promoted the proliferation,invasion,migration and glycolysis of cervical cancer cells(P<0.05),whereas the opposite effect inhibited the proliferation,invasion,migration and glycolysis of cervical cancer cells(P<0.05).Bioinformatics analysis and WB results showed that CCNO overexpression may regulate the occurrence of cervical cancer by activating the PI3K/AKT signaling pathway(P<0.05).Conclusion Cyclin O may promote the proliferation and metastasis of cervical cancer cells by regulating glycolysis.

Objective To screen the quality evaluation indicators of Bushen Huoxue Prescription(BHP)in the treatment of diabetic retinopathy(DR)by network pharmacology and molecular docking technology,and to establish content determination of active components in BHP by ultra-high-performance liquid chromatography triple-quadrupole mass spectrometry(UHPLC-QqQ-MS/MS).Methods Network pharmacology was used to screen the disease-related targets and key components,followed by molecular docking to further verify the interaction between them and confirm the active ingredients in BHP as the quality evaluation indicators.A UHPLC-QqQ-MS/MS method for the content determination of the active ingredients in BHP was established.A ZORBAX SB-C18 column(2.1 mm×50 mm,1.8 μm)was used,and methanol(A)-0.1％formic acid solution(B)was used as mobile phase.Gradient elution was performed in multiple reaction monitor mode.The flow rate was 0.3 mL·min-1 and the injection volume was 1 μL.Results Network pharmacology and molecular docking revealed five core targets and 12 BHP-related components(verbascoside,echinacoside,isoacteoside,tanshinone ⅡA,cryptotanshinone,dihydrotanshinone I,ginsenoside Re,Rd and Rb1,puerarin,daidzin,biochanin A)for the treatment of DR.There was a strong binding affinity between them(binding energy≤-5.0 kcal·mol-1).The established quantitative method demonstrated each component presented a good linearity within the specified range(r>0.999 5).The average recovery was in the range of 97.57％～101.48％.The contents of 12 components in eight batches of BHP samples were 0.027 9％～0.050 6％,0.006 4％～0.022 0％,0.017 1％～0.041 5％,0.009 2％～0.015 4％,0.012 6％～0.020 5％,0.004 4％～0.007 6％,0.334％～0.643％,0.238％～0.530％,0.353％～0.693％,3.411％～6.048％,1.023％～1.352％,0.000 8％～0.001 8％,respectively.Conclusion Based on network pharmacology,molecular docking and UHPLC-QqQ-MS/MS,a method for rapid screening and determination of 12 active components of BHP in the prevention and treatment of DR was established.This study provided a reference for comprehensive assessment of the quality and effectiveness of BHP.

Objective To compare and analyze the differences in CD8+naive,effector,memory,exhausted and regulatory T cells in order to investigate the impact of gender on the differentiation fate of CD8+T cells in the context of type 1 diabetes (T1D)based on female and male non-obese diabetic (NOD)mice and healthy Institute for Cancer Research (ICR)mice.Methods The frequencies and phenotypes of CD8+T cell differentiation subsets including naive T cells (TN),central memory T cells (TCM),effector T cells(TEFF),effector precursor T cells (TEP),exhausted T cells (TEX),precursor exhausted T cells (TPEX)and regulatory T cells (Tregs)in the spleen,pancreatic draining lymph nodes (pLN)and pancreas infiltrating lymphocytes (PIL)of male and female NOD mice were detected by flow cytometry.Results The frequencies of IFN-γ+,CD107a+and CCL5+CD8+TEFF in pLN and PIL of female NOD mice were significantly higher than those of male NOD mice.However,the frequencies of CD8+TN,CD8+TCM,CD8+TEX,CD8+TPEX and CD122+CD8+Tregs subsets in the spleen were significantly decreased.While there were no significant differences in the above CD8+T cell subsets except CD8+Tregs between female and male ICR mice. Conclusion Androgen may inhibit the differentiation of memory T cells into effector T cells and promote the exhaustion of effector T cells,leading to the difference in morbidity between the male and female mice.

Uncovering the risk factors of pulmonary hypertension and its mechanisms is crucial for the prevention and treatment of the disease.In the current study,we showed that experimental periodontitis,which was established by ligation of molars followed by orally smearing subgingival plaques from patients with periodontitis,exacerbated hypoxia-induced pulmonary hypertension in mice.Mechanistically,periodontitis dysregulated the pulmonary microbiota by promoting ectopic colonization and enrichment of oral bacteria in the lungs,contributing to pulmonary infiltration of interferon gamma positive(IFNγ+)T cells and aggravating the progression of pulmonary hypertension.In addition,we identified Prevotella zoogleoformans as the critical periodontitis-associated bacterium driving the exacerbation of pulmonary hypertension by periodontitis,and the exacerbation was potently ameliorated by both cervical lymph node excision and IFNγ neutralizing antibodies.Our study suggests a proof of concept that the combined prevention and treatment of periodontitis and pulmonary hypertension are necessary.

AIM:To explore the effect of polyphenolic compound 3,5-dihydroxy-4-methoxybenzyl alcohol(DHMBA)on hypoxia/reoxygenation(H/R)injury of human umbilical vein endothelial cells(EA.hy926 cells)and its po-tential mechanisms.METHODS:To construct an H/R model,the EA.hy926 cells were cultured in an acidic hypoxia buffer while in an anaerobic workstation.The cells were divided into control,H/R,H/R+different doses of DHMBA,H/R+edaravone(antioxidant)and H/R+reactive oxygen species(ROS)inducer oligomycin A+DHMBA groups.Cell viability was measured by CCK-8 assay,and tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)levels in cells were mea-sured by ELISA.Phosphorylation of endothelial nitric oxide synthase(eNOS)and nuclear factor-κB(NF-κB)p65 were measured by Western blot.Intracellular NO levels were determined by laser confocal microscopy.Glutathione(GSH)/glu-tathione disulfide(GSSG)oxidation balance was determined by the dinitrobenzoic acid chromogenic method.Intracellular ROS levels were measured by flow cytometry.Lactate dehydrogenase(LDH)leakage was determined using nitro blue tet-razolium staining.Scratch assays were performed to assess cell migration.RESULTS:DHMBA exhibited no significant cytotoxicity between 125 and 1 000 μmol/L.In H/R-injured human umbilical vein endothelial cells,DHMBA improved cell survival,inhibited phosphorylation of NF-κB p65,reduced the content of TNF-α and IL-6,and increased phosphory-lation of eNOS and NO levels.DHMBA also suppressed ROS overload and restored the ratio between GSH and oxidized GSH,decreased in LDH leakage and increased cell migration in H/R-injured human umbilical vein endothelial cells.CONCLUSION:DHMBA can alleviate H/R-induced oxidative stress,inflammation,cellular damage,and dysfunction,which are associated with the ability of DHMBA to inhibit ROS production in human umbilical vein endothelial cells.

Objective:The human peripheral blood lymphocyte cell line 8E5 is capable of secreting non-infectious HIV-1 viral particles. By targeting the POL region of the HIV-1 proviral gene integrated into the genome of 8E5 cell line and constructing a monoclonal cell line containing a drug resistance mutation site in the POL region using CRISPR/Cas9 point mutation technology, safe and stable HIV-1 genotypic drug resistance test quality control materials can be prepared.Methods:8E5 cells were co-transfected with sgRNA (single guide RNA) and Cas9 coexpression vector and Donor ssODN (donor single-stranded oligonucleotides) carrying the target mutation sites. The positive monoclonal cell lines were obtained through flow microtiter plate sorting, and the editing efficacy of the targeted mutations was validated by Sanger sequencing. Sanger sequencing was performed to verify the editing effect of the targeted mutations on the HIV virus particles secreted into the supernatant of the monoclonal cell lines cultured to the 3rd, 5th and 7th generations.Results:A double sgRNA and Cas9 coexpression vector was successfully constructed and co-transfected with a Donor ssODN carrying the drug-resistant mutation site Q151M to the 8E5 cell line, resulting in the desired outcome. The sequencing result of the target site confirmed the successful mutation at the resistance site and the establishment of a monoclonal homozygous cell line. The Q151M mutation site was detected in non-infectious HIV-1 virus particles secreted by the 8E5 Q151M cell line after transmission. Conclusions:The cell line 8E5 Q151M was successfully constructed using CRISPR/Cas9 point mutation technology to stably carry the Q151M drug resistance mutation site, which provides a new technological platform for the preparation of quality-control materials for testing HIV-1 genotypic drug resistance.

OBJECTIVE: To explore the clinical and genetic characteristics of four patients with medium-chain acyl-CoA dehydrogenase deficiency (MCADD). METHODS: Four children who had presented at the Children's Hospital Affiliated to Zhengzhou University between August 2019 and August 2021 were selected as the study subjects. Clinical data of the children were collected. The children were subjected to whole exome sequencing (WES). RESULTS: All of the four children were diagnosed with MCADD. Blood amino acid and ester acyl carnitine spectrum test showed that the concentration of octanoyl carnitine (C8) was significantly increased. The main clinical manifestations included poor mental response (3 cases), intermittent diarrhea with abdominal pain (1 case), vomiting (1 case), increased transaminase (3 cases), and metabolic acidosis (2 cases). Five variants were identified by genetic testing, among which c.341A>G (p.Y114C) was unreported previously. Three were missense variants, one was frameshift variant and one was splicing variant. CONCLUSION The clinical heterogeneity of MCADD is obvious, and the severity of the disease may vary. WES can assist with the diagnosis. Delineation of the clinical symptoms and genetic characteristics of the disease can facilitate early diagnosis and treatment of the disease.
Child ; Humans ; Acyl-CoA Dehydrogenase/genetics* ; Carnitine ; Genetic Testing ; Lipid Metabolism, Inborn Errors/genetics* ; Neonatal Screening

OBJECTIVE: To explore the clinical features and genetic basis of a child with Galactosemia. METHODS: A child who had presented at the Children's Hospital Affiliated to Zhengzhou University on November 20, 2019 was selected as the study subject. Clinical data of the child was collected. Whole exome sequencing was carried out for the child. Candidate variants were validated by Sanger sequencing. RESULTS: Clinical manifestations of the child have included anemia, feeding difficulty, jaundice, hypomyotonia, abnormal liver function and coagulation abnormality. Tandem mass spectrometry showed increased citrulline, methionine, ornithine and tyrosine. Urine organic acid analysis showed increased phenyllactic acid, 4-hydroxyphenylacetic acid, 4-hydroxyphenyllactic acid, 4-hydroxyphenylpyruvate and N-acetyltyrosine. Genetic testing revealed that the child has harbored compound heterozygous variants of the GALT gene, namely c.627T>A (p.Y209*) and c.370G>C (p.G124R), which were respectively inherited from her healthy parents. Among these, c.627T>A (p.Y209*) was known as a likely pathogenic variant, while c.370G>C (p. G124R) was unreported previously and also predicted as a likely pathogenic variant(PM1+PM2_Supporting+PP3_Moderate+PPR). CONCLUSION Above discovery has expanded the spectrum of the GALT gene variants underlying Galactosemia. Patients with thrombocytopenia, feeding difficulties, jaundice, abnormal liver function and coagulation abnormality without obvious causes should be analyzed by screening of metabolic diseases in combination with genetic testing.
Child ; Female ; Humans ; Galactosemias/genetics* ; Genetic Testing ; Health Status ; Methionine ; Muscle Hypotonia ; Mutation