Objective:To establish a TaqMan-MGB probe-based method for detecting the polymorphic loci C677T and A1298C of the MTHFR gene.Methods:Specific primers and TaqMan-MGB probes targeting the C677T and A1298C polymorphic loci of the MTHFR gene were designed and optimized based on the gene sequence information.A real-time quantitative PCR detection system was established.Gradient dilution experiments were conducted to determine the limit of detection,and reproducibility experiments were performed to evaluate detection consistency.Specificity was validated using wild-type and mutant plasmid templates.The method was applied to detect 56 clinical samples,and its accuracy and practicality were assessed through comparison with traditional Sanger sequencing.Results:The TaqMan-MGB probe method demonstrated high specificity for detecting the C677T and A1298C loci,with no cross-reactivity between wild-type and mutant probes,enabling accurate genotype differentiation.Sensitivity experiments revealed detection limits of 1.13 × 103 copies/μL for C677T and 8.39 × 101 copies/μL for A1298C.Reproducibility experiments showed coefficients of variation below 1％,indicating stable and reliable results.Among the 56 clinical samples,the overall detection rate for the C677T locus was 86.99％,and for the A1298C locus,it was 97.92％.The TaqMan-MGB method exhibited good concordance with Sanger sequencing results.Conclusion:The TaqMan-MGB method exhibits high specificity,sensitivity,and excellent reproducibility in detecting the polymorphic loci C677T and A1298C of the MTHFR gene,making it suitable for rapid detection in large-scale clinical samples.This method provides an effective molecular diagnostic tool for the early diagnosis and prevention of folate-related diseases.

OBJECTIVE: To analyze a patient with infertility due to complex chromosome rearrangement by optical genome mapping (OGM). METHODS: A female patient who was diagnosed with "primary infertility" at Shenzhen Longhua District Maternal and Child Health Care Hospital in April 2024 was selected as the study subject. Clinical data of the patient was collected. Chromosome G banding karyotyping analysis was carried out for the patient and her parents, in addition with OGM and copy number variation sequencing (CNV-seq). This study was approved by the Medical Ethics Committee of the Hospital (Ethics No.: 2023052504). RESULTS: The patient, a 33-year-old female, had infertility for the past 5 years. OGM revealed formation of two derivative chromosomes through rearrangement of chromosomes 5 and 18. A loss of heterozygosity on chromosome 5 was also detected by OGM and CNV-seq techniques. Both of her parents had a normal karyotype. CONCLUSION The OGM technique can refine the position of chromosomal breakpoints and determine the direction and position of insertional fragment. Combined with karyotype analysis, the OGM can accurately determine the chromosomal karyotype of the patient and facilitate genetic counseling.
Humans ; Female ; Adult ; Karyotyping ; DNA Copy Number Variations/genetics* ; Chromosome Mapping/methods* ; Chromosome Aberrations ; Infertility, Female/diagnosis*

This study was aimed at establishing a method of ultra-fast quantitative PCR for Brucella detection.We used an exogenous recombinant plasmid as the internal reference and targeted the T4SS secretion system,an important Brucella viru-lence factor,to design specific primers and probes.The sensitivity,specificity,and repeatability of this method were evaluated,and a standard curve was constructed.The coincidence rate of detection findings with this method versus quantitative PCR was determined.This method markedly decreased the detection time to only 10 minutes.The standard curve demonstrated a good linear relationship(Y=-3.410 7x+38.357,R2=0.998 5)with a low minimum detection limit of 10 copies/μL.The method exhibited good specificity and did not specifically amplify several common clinical bacteria other than Brucella.The de-tection of three concentrations of positive plasmids yielded coefficients of variation(CVs)of 0.20％to 0.91％,thus demonstra-ting the method's excellent repeatability.Furthermore,140 clinical samples were analyzed concurrently with the fluorescence PCR method,which yielded a 100％compliance rate and consistent results.Our findings indicated that the Brucella ultra-fast quantitative PCR was ultrafast;had high sensitivity,high specificity,and good specificity;and can be used for the clinical de-tection of Brucella and emergency investigation of epidemics.Therefore,this method is valuable for the early diagnosis of Bru-cella.

Objective:To establish a TaqMan-MGB probe-based method for detecting the polymorphic loci C677T and A1298C of the MTHFR gene.Methods:Specific primers and TaqMan-MGB probes targeting the C677T and A1298C polymorphic loci of the MTHFR gene were designed and optimized based on the gene sequence information.A real-time quantitative PCR detection system was established.Gradient dilution experiments were conducted to determine the limit of detection,and reproducibility experiments were performed to evaluate detection consistency.Specificity was validated using wild-type and mutant plasmid templates.The method was applied to detect 56 clinical samples,and its accuracy and practicality were assessed through comparison with traditional Sanger sequencing.Results:The TaqMan-MGB probe method demonstrated high specificity for detecting the C677T and A1298C loci,with no cross-reactivity between wild-type and mutant probes,enabling accurate genotype differentiation.Sensitivity experiments revealed detection limits of 1.13 × 103 copies/μL for C677T and 8.39 × 101 copies/μL for A1298C.Reproducibility experiments showed coefficients of variation below 1％,indicating stable and reliable results.Among the 56 clinical samples,the overall detection rate for the C677T locus was 86.99％,and for the A1298C locus,it was 97.92％.The TaqMan-MGB method exhibited good concordance with Sanger sequencing results.Conclusion:The TaqMan-MGB method exhibits high specificity,sensitivity,and excellent reproducibility in detecting the polymorphic loci C677T and A1298C of the MTHFR gene,making it suitable for rapid detection in large-scale clinical samples.This method provides an effective molecular diagnostic tool for the early diagnosis and prevention of folate-related diseases.

Nasojejunal feeding tubes are widely used in surgical,intensive care,and older patients.Manual blind insertion of nasojejunal feeding tubes is technically challenging,associated with a high failure rate,and prone to complications.The primary causes of suboptimal placement outcomes are the uncertainty and weak controllability of the interaction forces during the coordination between manual posterior advancement and the patient's physiological state.While current auxiliary techniques such as X-ray,ultrasound,and endoscopy can improve the success rate of nasojejunal tube placement and reduce complications to some extent,the accuracy and safety of placement remain constrained by challenges in controlling insertion forces and achieving precise positional localization.Robotic technology holds promise for addressing the uncertainties and controllability issues inherent in the placement process.By leveraging precise sensing,real-time navigation,and efficient control,robots can achieve intelligent positioning and precise control over the direction and location of the catheter tip during nasojejunal intubation.However,current research on robotic applications for nasojejunal feeding tube placement is still in an early stage,facing challenges such as high costs,operational complexity,and concerns over safety and reliability.Herein,we analyzed the limitations and causes of failure in existing placement methods and explored the application prospects of robotic technologies for precise control and intelligent positioning in nasojejunal feeding tube placement.The paper provides new insights for developing nursing techniques that enable safer and more effective,comfortable,and rapid intubation.Future efforts should focus on deepening the integration of artificial intelligence and robotics,optimizing drive technologies,and accelerating the translation of these technologies from the laboratory to clinical practice.This will drive the advancement of nasojejunal feeding tube placement techniques towards intelligent,precise,and accessible solutions.

Objective To establish a stable,reliable,and clinically relevant porcine model of endotoxin-induced acute respiratory distress syndrome(ARDS).Methods Ten 8-month-old male Bama minipigs were deeply sedated,followed by invasive mechanical ventilation and electrocardiographic monitoring.Lipopolysaccharide(LPS)was intravenously pumped at 600 μg/(kg·h)for 3 hours,then maintained at 15 μg/(kg·h)thereafter.Dynamic monitoring was performed at five time points after LPS injection(LPS 0,1,3,5,and 8 h),including arterial blood gas analysis and chest computed tomography(CT)scans.Pathological examination of lung tissues obtained via bronchoscopic biopsy(HE staining and transmission electron microscopy)was conducted.These indicators were comprehensively used to evaluate the success of the animal model.Results At 5 hours after LPS administration,8 minipigs developed symptoms such as skin cyanosis,elevated body temperature,and respiratory distress.The oxygenation index decreased to<300 mmHg.Chest CT scans showed diffuse pulmonary infiltrates.Histopathology revealed alveolar edema and hyaline membrane formation.Transmission electron microscopy demonstrated disruption of pulmonary blood-air barrier,depletion of lamellar bodies in type Ⅱ pneumocytes,inflammatory cell infiltration,and exudation of plasma proteins and fibrin.Compared with LPS 0 h,at LPS 8 h,the oxygenation index and arterial blood pH were significantly decreased(P<0.001),while blood lactic acid and serum potassium were significantly increased(P<0.05);serum calcium and base excess were significantly decreased(P<0.05),and the lung injury score based on HE-stained lung sections was significantly increased(P<0.01).Conclusion The porcine ARDS model established by continuous LPS injection can dynamically simulate the pathophysiological characteristics and typical pathological manifestations of clinical septic ARDS,making it an effective tool to study the pathogenesis,prevention,and treatment strategies of septic ARDS.

Chronic prostatitis,a common condition in andrology,is clinically characterized by a prolonged course,resistance to treatment,and frequent recurrence.In traditional Chinese medicine,it is classified under the categories of"turbid sperm,""overstrain strangury,"and"vaginal pain."Based on the"host-guest interaction-collateral disease"theory,we believe that healthy qi deficiency,latent pathogenic factors,and collateral obstruction are the primary pathological factors of this disease,which run through the entire process of chronic prostatitis occurrence and development.Accordingly,we propose that obstruction of collaterals and apathesia of the semen chamber are the core pathogenesis.The disease progression can be divided into three pathological stages:"deficiency,depression,and blood stasis."Spleen and kidney deficiency and malnutrition of collaterals form the pathological foundation.In the deficiency stage,treatment strategies involve reinforcing qi and nourishing the collaterals,using Fuzheng Yangrong Decoction.During disease progression,dampness and heat invasion,as well as collateral stagnation qi,are key contributors to disease progression.Thus,treatment focuses on clearing heat and dampness,promoting qi flow,and smoothing the collaterals,achieved with a modified Qiantongding Decoction.In the final stage,blood stasis and collateral obstruction dominate,warranting therapeutic strategies aimed at tonifying and removing blood stasis,addressing both the body and the collaterals simultaneously using the modified Guiling Huayu Decoction.Overall,the clinical treatment generally focuses on the concept of function through free flow,combination of unblocking and tonifying.This study provides a novel perspective and reference for clinical differentiation and treatment of chronic prostatitis.

AIM To investigate effects of petroleum ether fraction from Derris eriocarpa How on glucose and lipid metabolism in a mouse model of metabolic syndrome(MS).METHODS KM mice were fed a high-fat diet and administered streptozotocin intraperitoneally to establish MS models.The MS mice were then randomly assigned to the model group,the metformin hydrochloride group,the lovastatin group,the ursolic acid group,and the high-,medium-and low-dose D.eriocarpa petroleum ether fraction groups,with 10 mice in each group.Ten additional mice maitained on a normal diet served as the normal control group.After 4 weeks of intragastric administration,glucose and lipid metabolism indicators were measured.Hepatic pathological changes were assessed using HE staining and oil red O staining.Liver tissue mRNA expressions of ATF3,PEPCK,FXR,CYP7A1,HNF4ɑ,CYP8B1 and SRB1 were quantified by RT-qPCR.Hepatic protein expressions of ATF3,HNF4ɑ,PEPCK,FXR and CYP7A1 was analyzed by Western blot in MS mice.RESULTS Compared to the model group,the high-dose D.eriocarpa petroleum ether fraction group exhibited significant glucose tolerance improvement(reduced OGTT-AUC,P＜0.01);favorable serum lipid modulation in terms of increased HDL-C levels(P＜0.01)and decreased TG,TC,LDL-C(P＜0.01);reduced renal biomarkers(BUN,SCR)and hepatotoxic indicators of TBA,AST and ALT activities(P＜0.01);alleviated hepatic lipid accumulation and histopathological damage;downregulated mRNA and protein expressions of ATF3,HNF4ɑ and PEPCK,as well as CYP8B1 mRNA expression(P＜0.01);and upregulated mRNA and protein expressions of FXR and CYP7A1,along with SRB1 mRNA expression(P＜0.01).CONCLUSION D.eriocarpa petroleum ether fraction ameliorates glucose and lipid metabolism dysregulation in MS mice by modulating the ATF3/HNF4ɑ/CYP7A1 signaling pathway,consequently eliciting hypoglycemic,hypolipidemic,hepatoprotective and nephroprotective effects.

A 58-year-old female patient with breast cancer received treatment with pegylated recombinant human granulocyte colony-stimulating factor(PEG-rhG-CSF)injection after adjuvant chemotherapy.The serum creatinine level of the patient gradually increased from the normal baseline value to 199.3 μmol·L-1.The patient was diagnosed as acute kidney injury(AKI),after stopping the medication and providing symptomatic treatment,the patient's renal function gradually improved.The patient completed subsequent chemotherapy as planned without reusing PEG-rhG-CSF injection,and other medications and dosages remained unchanged.Renal function remained stable during follow-up.Naranjo's Assessment Scale was used to evaluate the association between PEG-rhG-CSF injection and AKI,the result was"probable."There are few reports of AKI occuring with PEG-rhG-CSF injection,and this case provides evidence for clinical safe medication.