Objective:To investigate the clinical value of the Nomogram model based on two-dimensional shear wave elastography(2D-SWE)combined with biochemical indicators in the diagnosis of moderate and severe metabolic-associated fatty liver disease(MAFLD).Methods:A total of 170 patients with MAFLD diagnosed by ultrosound image in the health management center of the Second People's Hospital of Wuhu from Jan 2023 to Dec 2023 were divided into mild,moderate and severe groups according to two-dimensional ultrasound images,and 111 healthy volunteers were recruited as control group in the same period.Multivariate logistic regression analysis was used to screen independent risk factors for moderate and severe MAFLD and construct a predictive model.The diagnostic efficacy of the ROC model was plotted,and the Nomogram model was used to conduct internal verification.Results:Multivariate logistic regression analysis showed that liver stiffness measurement(LSM),platelet count(PLT)and hepatic steatosis index(HSI)were the independent risk factors for the diagnosis of moderate and severe MAFLD.The area under the curve(AUC)of LSM,PLT and HSI was 0.940(95%CI:0.911-0.969),with a sensitivity was 87.5%and a specificity was 88.1%.The internal validation showed that the model had high accuracy and stability.Conclusion:The Nomogram model based on LSM combined with PLT and HSI can effectively diagnose moderate and severe MAFLD,providing a reliable evidence for early clinical intervention and adjustment of therapeutic measures.

Objective:The aim of this study is to analyze the clinical characteristics and genetic variants of a late-onset auditory neuropathy pedigree caused by maternally inherited- mitochondrial mutation.Methods:A male proband who presented with bilateral sensorineural hearing loss at Henan Children′s Hospital in September 2023 was chosen, along with his family members (4 generations, 20 individuals) as the study subjects. Data from this pedigree were collected, organized, and analyzed for clinical genetic characteristics. Medical histories were obtained from family members, pedigree charts were drawn, audiological, imaging, and physical examinations were conducted. Pathogenic genes and mutations were screened using high-throughput sequencing. Sanger sequencing was employed for variant confirmation and segregation validation in the family.Results:In this family, a total of 12 members (10 members collected) had sensorineural hearing loss, characterized by late-onset hearing impairment with an onset age ranging from 9 to 30 years. The patients exhibited poor speech recognition rates, and audiometric examinations are consistent with auditory neuropathy. There was no history of ototoxic drug use. High-throughput sequencing identified the variant NC_012920.1:m.7471dup in the mitochondrial MT-TS1 gene as the pathogenic variant. Sanger sequencing results confirmed that the pathogenic gene mutation site perfectly co-segregated with the auditory neuropathy phenotype in this family. According to the classification criteria and guidelines for genetic variations by the American College of Medical Genetics and Genomics, the variant was classified as a pathogenic mutation. Conclusion:The mitochondrial MT-TS1 gene mutation m.7471dup is considered to be the pathogenic cause in this late-onset auditory neuropathy pedigree.

Objective:To investigate the clinical value of the Nomogram model based on two-dimensional shear wave elastography(2D-SWE)combined with biochemical indicators in the diagnosis of moderate and severe metabolic-associated fatty liver disease(MAFLD).Methods:A total of 170 patients with MAFLD diagnosed by ultrosound image in the health management center of the Second People's Hospital of Wuhu from Jan 2023 to Dec 2023 were divided into mild,moderate and severe groups according to two-dimensional ultrasound images,and 111 healthy volunteers were recruited as control group in the same period.Multivariate logistic regression analysis was used to screen independent risk factors for moderate and severe MAFLD and construct a predictive model.The diagnostic efficacy of the ROC model was plotted,and the Nomogram model was used to conduct internal verification.Results:Multivariate logistic regression analysis showed that liver stiffness measurement(LSM),platelet count(PLT)and hepatic steatosis index(HSI)were the independent risk factors for the diagnosis of moderate and severe MAFLD.The area under the curve(AUC)of LSM,PLT and HSI was 0.940(95%CI:0.911-0.969),with a sensitivity was 87.5%and a specificity was 88.1%.The internal validation showed that the model had high accuracy and stability.Conclusion:The Nomogram model based on LSM combined with PLT and HSI can effectively diagnose moderate and severe MAFLD,providing a reliable evidence for early clinical intervention and adjustment of therapeutic measures.

Objective:The aim of this study is to analyze the clinical characteristics and genetic variants of a late-onset auditory neuropathy pedigree caused by maternally inherited- mitochondrial mutation.Methods:A male proband who presented with bilateral sensorineural hearing loss at Henan Children′s Hospital in September 2023 was chosen, along with his family members (4 generations, 20 individuals) as the study subjects. Data from this pedigree were collected, organized, and analyzed for clinical genetic characteristics. Medical histories were obtained from family members, pedigree charts were drawn, audiological, imaging, and physical examinations were conducted. Pathogenic genes and mutations were screened using high-throughput sequencing. Sanger sequencing was employed for variant confirmation and segregation validation in the family.Results:In this family, a total of 12 members (10 members collected) had sensorineural hearing loss, characterized by late-onset hearing impairment with an onset age ranging from 9 to 30 years. The patients exhibited poor speech recognition rates, and audiometric examinations are consistent with auditory neuropathy. There was no history of ototoxic drug use. High-throughput sequencing identified the variant NC_012920.1:m.7471dup in the mitochondrial MT-TS1 gene as the pathogenic variant. Sanger sequencing results confirmed that the pathogenic gene mutation site perfectly co-segregated with the auditory neuropathy phenotype in this family. According to the classification criteria and guidelines for genetic variations by the American College of Medical Genetics and Genomics, the variant was classified as a pathogenic mutation. Conclusion:The mitochondrial MT-TS1 gene mutation m.7471dup is considered to be the pathogenic cause in this late-onset auditory neuropathy pedigree.

The main functional ingredients of hot water extract of Chlorella pyrenoidosa (CPE) were investigated through a bioassay-guided fractionation based on free radical scavenging and macrophage proliferation effects. The main functional ingredients of CPE were polysaccharides (PS) that were isolated by high pressure extraction, Sevag method, ethanol precipitation and ultrafiltration separation. Crude polysaccharides were further separated and purified by ion exchange chromatography DEAE52 and size exclusion chromatography Sephadex G-100. The purified fractions were analyzed by gel permeation chromatography. Molecular weights of the purified fractions PS-1-4-2, PS-1-3-2 and PS-2-3-3 were 3.97×10⁴, 2.28×10⁴ and 4.1×10³ Da, respectively. Bioassay-guided fractionation results indicated that CPE could remove free radicals and promote Ana-1 cells proliferation, mainly due to its various components working together. The components of free radicals scavenging mainly concentrated in PS-1-3, PS-1-4, PS-2-3 and PS-2-4. The components of Ana-1 proliferation mainly concentrated in PS-1-3, PS-1-4 and PS-2-3. This study established the activity screening method of main functional component from CPE, and got three new functional ingredients. It can be used to guide the development of high value products, further promote the industrialization process of microalgae energy, and realize microalgae 'high value products, microalgae energy and microalgae carbon' integration of exemplary role.

This research was aimed at establishing the pilot-scale purification technology of lipopeptide from marine-derived Bacillus marinus. We studied lipopeptide surfactivity interferences on scale-up unit technologies including acid precipitation, methanol extraction, solvent precipitation, salting out, extraction, silica gel column chromatography and HZ806 macroporous absorption resin column chromatography. Then, the unit technologies were combined in a certain order, to remove the impurities gradually, and to gain purified lipopeptide finally, with high recovery rate throughout the whole process. The novel pilot-scale purification technology could effectively isolate and purify lipopeptide with 87.51% to 100% purity in hectograms from 1 ton of Bacillus marinus B-9987 fermentation broth with more than 81.73% recovery rate. The first practical hectogram production of highly purified lipopeptide derived from Bacillus marinus was achieved. With this new purification method, using complex media became possible in fermentation process to reduce the fermentation cost and scale-up the purification for lipopeptide production. For practicability and economy, foaming problem resulting from massive water evaporation was avoided in this technology.

Objective To establish a real-time fluorescence quantitative PCR method for detection of the different expression level of FPGS in methotrexate enantiomer-resistant A549 cell lines,and observe FPGS mRNA expression in patients with leukemia.Methods A real-time fluorescence quantitative PCR method for detection FPGS mRNA was established using SYBR Green Ⅰ as fluorescence and β-actin as reference.The method was evaluated by Ct,correlation coefficient,slope,repeatability curve,melting curve and amplification efficiency curve.The expression levels of FPGS gene in methotrexate enantiomer-resistant A549 cell lines and methotrexate resistant leukemia cells in bone marrow were detected by the method.Results The standard curves had a high linear relationship between cycle threshold and template concentration.The correlation coefficients of FPGS and β-actin were 0.996 8 and 0.998 7,and the slopes were -3.595 and -3.740,respectively.The inter-coefficient of variation was from 1.27% to 2.95%.The intra-coefficient of variation was 3.82%.The method was characterized with specific melting curve and similar amplification efficiency(slope was 0.021 7).The relative contents of FPGS mRNA were(3.51 ±0.66),(0.16 ±0.01) and(1.00 ±0.31) in L-(+)-MTX/A549 cells(L),D-(-)-MTX/A549 cells(D)and A549 parent cells,and there was statistically difference among the three groups(F = 64.45 ,P＜ 0.01)Statistical difference was observed between L and D(q =9.29,P＜0.01).After treated with MTX,the expression level of FPGS mRNA was(0.35 ± 0.04) in methotrexate resistant leukemia patients,compared with(1.00 ± 0.44) before treatment.Statistical difference was observed(t = 8.83 ,P＜ 0.01).Conclusions The real-time fluorescence quantitative PCR is suitable for the quantification of FPGS.The expression levels of FPGS in methotrexate resistant leukemia cells in bone marrow and drug resistant cells are different.Two enantiomer forms of methotrexate may play different roles in drug resistance mechanisms.

This study aimed to determine whether aldosterone could induce vascular cell apoptosis in vivo. Thirty-two male rats were randomly divided into 4 groups: vehicle (control), aldosterone, aldosterone plus eplerenone or hydralazine. They were then implanted with an osmotic mini-pump that infused either aldosterone or the vehicle. Systolic blood pressure (SBP) was measured weekly by the tail-cuff method. After 8 weeks, plasma aldosterone concentration (PAC) and renin activity (PRA) were determined by radioimmunoassay. Aortic apoptosis was examined by TUNEL assay. The levels of cytochrome c and caspase-3 were determined by Western blotting and the expression of Bax and Bcl-2 was detected by immnuohistochemistry and Western blotting. The results showed that as compared with control group, aldosterone-infused rats exhibited: (1) an increase in SBP; (2) significantly elevated PAC with depressed PRA; (3) elevated aortic vascular cell apoptosis accompanied with higher levels of cytochrome c and activated caspase-3; and (4) significantly up-regulated Bax protein with down-regulated Bcl-2. These effects of aldosterone were significantly inhibited after co-administration with eplerenone but not with hydralazine. It was concluded that aldosterone induced vascular cell apoptosis by its direct effect on the aorta via mineralocorticoid receptors and independently of blood pressure, which may contribute to aldosterone-mediated vascular injury.

This study aimed to determine whether aldosterone could induce vascular cell apoptosis in vivo.Thirty-two male rats were randomly divided into 4 groups: vehicle(control),aldosterone,aldosterone plus eplerenone or hydralazine.They were then implanted with an osmotic mini-pump that infused either aldosterone or the vehicle.Systolic blood pressure(SBP)was measured weekly by the tail-cuff method.After 8 weeks,plasma aldosterone concentration(PAC)and renin activity(PRA)were determined by radioimmunoassay.Aortic apoptosis was examined by TUNEL assay.The levels of cytochrome c and caspase-3 were determined by Western blotting and the expression of Bax and Bcl-2 was detected by immnuohistochemistry and Western blotting.The results showed that as compared with control group,aldosterone-infused rats exhibited:(1)an increase in SBP;(2)significantly elevated PAC with depressed PRA;(3)elevated aortic vascular cell apoptosis accompanied with higher levels ofcytochrome c and activated caspase-3; and(4)significantly up-regulated Bax protein with down-regulated Bcl-2.These effects of aldosterone were significantly inhibited after co-administration with eplerenone but not with hydralazine.It was concluded that aldosterone inducedvascular cell apoptosis by its direct effect on the aorta via mineralocorticoid receptors and independently of blood pressure,which may contribute to aldosterone-mediated vascular injury.

Objective:To investigate the effect of traditional Chinese medicine (TCM) icariin (ICA) on metastatic phenotype of methotrexate (MTX)-resistant lung cancer A549 cells and elucidate the action mechanism and therapeutic value of ICA. Methods:The half inhibition concentration(IC_(50))value of ICA in inhibiting the growth of A549/MTX cells was measured by MTT assay. The colony formation rates and the morphology of cell cluster of A549/MTX and ICA-treated A549/MTX cells were determined by double-layer soft agar colony formation assay. The migration abilities of A549/MTX and ICA-treated A549/MTX cells were evaluated by cell scratch assay. The invasion ability of cells was tested by using Transwell chamber assay. Results:MTT assay showed that the IC_(50) value of non-toxic ICA plus MTX was reduced compared with that induced by equal dose of MTX (35.50±1.85 μmol/L vs 52.17±2.25 μmol/L). The colony formation rate of ICA-treated A549/MTX cells was 0.94±0.09, less than that of A549/MTX cells (1.56±1.07, P<0.05). Cell scratching assay demonstrated that the migration capability of A549/MTX cells was stronger than that of ICA-treated A549/MTX cells at 72 h (P<0.05). Transwell experiment revealed that more A549/MTX cells passed through artificial basement membrane than ICA-treated A549/MTX cells (P<0.05), indicating that the invasion capability of ICA-treated A549/MTX cells was weaker than that of A549/MTX cells.Conclusion:TCM ICA can reverse the metastatic phenotype of MTX-resistant A549 cells.