Objective To examine the impact of different numbers of microsatellite markers on the analysis of population genetic diversity of Schistosoma japonicum, so as to provide insights into studies on the population genetic diversity of S. japonicum. Methods Oncomelania hupensis snails were collected from a wasteland in Gong’an County, Hubei Province, and 37 S. japonicum-infected O. hupensis snails were identified using the cercarial shedding method. A single cercaria released from each S. japonicum-infected O. hupensis snail was collected, and 10 cercariae were randomly collected from DNA extraction. Nine previously validated microsatellite loci and 15 additional microsatellite loci screened from literature review and the GenBank database and confirmed with stable amplification efficiency were selected as molecular markers. Genomic DNA from cercariae was subjected to three multiplex PCR amplifications of microsatellite markers with the Type-it Microsatellite PCR kit, and genotyped using capillary electrophoresis. The population genetic diversity of S. japonicum cercariae DNA was analyzed with observed number of alleles (Na), effective number of alleles (Ae), observed heterozygosity (Ho), expected heterozygosity (He), and polymorphism information content (PIC), and tested for Hardy-Weinberg equilibrium (HWE) and linkage disequilibrium (LD). To further investigate the impact of the number of microsatellite loci on the population genetic diversity of S. japonicum, the number of microsatellite markers was sequentially assigned from 1 to 24, and the mean and standard deviation of Na were calculated for S. japonicum populations at different locus numbers. In addition, the coefficient of variation (CV) of allelic number (defined as the ratio of the standard deviation to the mean) was determined, and the variation in Na with increasing microsatellite locus numbers was analyzed. Results Genomic DNA from 345 S. japonicum cercariae was selected for genotyping of 24 microsatellite markers, and all 24 microsatellite loci met linkage equilibrium (standardized linkage disequilibrium coefficient D′ < 0.7, r² < 0.3) and deviated from Hardy-Weinberg equilibrium (P < 0.001). The mean Na, Ae, Ho and He were 27.46 ± 2.18, 12.46 ± 0.95, 0.46 ± 0.03, and 0.91 ± 0.01 for 24 microsatellite loci in S. japonicum cercarial populations, respectively, and PIC ranged from 0.85 to 0.96, indicating high genome-wide representativeness of 24 microsatellite loci. The mean value of Na-Ae was higher in genotyping with 9 previously validated microsatellite loci (19.88 ± 8.43) than with all 24 loci (14.99 ± 8.09). As the number of microsatellite loci increased, the mean Na showed no significant variation; however, the standard deviation gradually decreased. Notably, if the locus number reached 18 or more, the variation in the standard deviation of Na remarkably reduced. In addition, the standard deviation of Na at 18 loci was less than 5% of the mean Na at 24 loci, with a CV of 4.6%. Conclusions The number of microsatellite loci significantly affects the population genetic diversity analysis of S. japonicum. Eighteen or more microsatellite loci are recommended for analysis of the population genetic diversity of S. japonicum under the current conditions of low-prevalence infection and unbalanced genetic distribution of S. japonicum.

Objective:To explore the clinical characteristics,molecular characteristics,treatment and prognosis of pediatric Philadelphia chromosome-like acute lymphoblastic leukemia(Ph-like ALL)with a therapeutic target.Methods:A total of 27 patients of Ph-like ALL with targeted drug target were initially diagnosed in Children's Hospital of Soochow University from December 2017 to June 2021.The data of age,gender,white blood cell(WBC)count at initial diagnosis,genetic characteristics,molecular biological changes,chemotherapy regimen,different targeted drugs were given,and minimal residual disease(MRD)on day 19,MRD on day 46,whether hematopoietic stem cell transplantation(HSCT)were retrospective analyed,and the clinical characteristics and treatment effect were summarized.Survival analysis was performed by Kaplan-Meier method.Results:The intensity of chemotherapy was adjusted according to the MRD level during induced remission therapy in 27 patients,10 patients were treated with targeted drugs during treatment,and 3 patients were bridged with HSCT,1 patient died and 2 patients survived.Among the 24 patients who did not receive HSCT,1 patient developed relapse,and achieved complete remission(CR)after treatment with chimeric antigen receptors T cells(CAR-T).The 3-year overall survival,3-year relapse-free survival and 3-year event-free survival rate of 27 patients were(95.5±4.4)％,(95.0±4.9)％and(90.7±6.3)％respectively.Conclusion:Risk stratification chemotherapy based on MRD monitoring can improve the prognosis of Ph-like ALL in children,combined with targeted drugs can achieve complete remission as soon as possible in children whose chemotherapy response is poor,and sequential CAR-T and HSCT can significantly improve the therapeutic effect of Ph-like ALL in children whose MRD is continuously positive during induced remission therapy.

Objective: To investigate the clinicopathological features and differential diagnosis of NTRK3 gene rearrangement thyroid papillary carcinoma (PTC). Methods: The PTC cases without BRAF V600E mutation were collected at Fujian Provincial Hospital South Branch from January 2015 to January 2020. The cases of NTRK3 gene rearrangement PTC were examined using immunohistochemistry and fluorescence in situ hybridization (FISH). The clinical data, histopathological characteristics, immunohistochemical features and molecular pathological changes were retrospectively analyzed. Data from the TCGA PTC dataset and the literature were also studied. Results: A total of 3 PTC cases harboring NTRK3 gene rearrangement were confirmed. All the patients were female, aged from 26,49,34 years. Histologically, two of them demonstrated a multinodular growth pattern. Only one case showed prominent follicular growth pattern; the other two tumors showed a mixture of follicular, papillary and solid growth patterns. All tumors showed a typical PTC nuclear manifestation, with some nuclear pleomorphism, vacuolated foci and oncocytic features. The characteristic formation of glomeruloid follicular foci was present in two cases which also showed psammoma bodies, and tumoral capsular or angiolymphatic invasion. The background thyroid parenchyma showed chronic lymphocytic thyroiditis. Mitotic rates were low, and no cases had any tumor necrosis. The pan-TRK and TTF1 testing was both positive in 3 cases, while S-100 and mammaglobin were both negative in them. FISH studies confirmed the NTRK3 gene rearrangement in all 3 cases. Studies on the TCGA datasets and literature revealed similar findings. Conclusions: NTRK3 gene rearrangement PTC is rare. It may be easily misdiagnosed due to the lack of histological and clinicopathological characteristics. Molecular studies such as pan-TRK immunostaining, FISH and even next-generation sequencing are needed to confirm the diagnosis. Immunohistochemistry of pan-TRK performed in the PTC cases without BRAF V600E mutation can be used as a good rapid-screening tool. With the emergence of pan-cancer tyrosine receptor kinase inhibitors, proper diagnosis of these tumors can help determine appropriate treatments and improve their outcomes.
Biomarkers, Tumor ; Female ; Gene Rearrangement ; Humans ; In Situ Hybridization, Fluorescence ; Mutation ; Proto-Oncogene Proteins B-raf/genetics* ; Receptor, trkC ; Retrospective Studies ; Thyroid Cancer, Papillary/genetics* ; Thyroid Neoplasms/genetics*

BACKGROUND: Pulmonary vein (PV) occlusion generally depends on repetitive contrast agent injection when cryoballoon ablation for atrial fibrillation (AF). The present study was to compare the effect of cryoballoon ablation for AF guided by transesophageal echocardiography (TEE) vs. contrast agent injection. METHODS: Eighty patients with paroxysmal AF (PAF) were enrolled in the study. About 40 patients underwent cryoballoon ablation without TEE (non-TEE group) and the other 40 underwent cryoballoon ablation with TEE for PV occlusion (TEE group). In the TEE group during the procedure, PVs were displayed in 3-dimensional images to guide the balloon to achieve PV occlusion. The patients were followed up at regularly scheduled visits every 2 months. RESULTS: No differences were identified between the groups in regard to the procedure time and cryoablation time for each PV. The fluoroscopy time (6.7 ± 4.2 min vs. 17.9 ± 5.9 min, P < 0.05) and the amount of contrast agent (3.0 ± 5.1 mL vs.18.1 ± 3.4 mL, P < 0.05) in the TEE group were both less than the non-TEE group. At a mean of 13.0 ± 3.3 mon follow-up, success rates were similar between the TEE group and non-TEE group (77.5% vs. 80.0%, P = 0.88). CONCLUSIONS Cryoballoon ablation with TEE for occlusion of the PV is both safe and effective. Less fluoroscopy time and a lower contrast agent load can be achieved with the help of TEE for PV occlusion during procedure.
Aged ; Atrial Fibrillation ; diagnostic imaging ; surgery ; Contrast Media ; Cryosurgery ; methods ; Echocardiography, Three-Dimensional ; methods ; Echocardiography, Transesophageal ; methods ; Female ; Humans ; Male ; Middle Aged ; Pulmonary Veins ; diagnostic imaging ; surgery ; Treatment Outcome

OBJECTIVE: To evaluate the clinical efficacy of decitabine combined with arsenious acid in the treatment of patients with higher-risk myelodysplastic syndromes(MDS) and chronic myelomonocytic leukemia(CMML). METHODS: Totally 39 patients with MDS and 8 patients with CMML received the treatment of decitabine and arsenious acid from April 2016 to December 2018. Decitabine [20 mg/(m~2·d)] and arsenious acid [0.15 mg/(m~2·d)] were administered intravenously for 5 consecutive days every 4-6 weeks. Patients who achieved complete or partial remission entered into the consolidation cycle. Efficacy and influencing factor were analyzed. RESULTS: Clinical response were observed in 31 patients after a median of 2 courses(ranging 1-12) of treatment. The overall response rate(ORR) was 66.0%. The median duration of response was 16 weeks(ranging 2-52 weeks). There were 8 cases(17.0%) of complete remission(CR), 10 cases(21.3%) of partial remission(PR),12 cases(25.5%) of hematological improvement(HI), 1 case(2.1%) of marrow complete remission(mCR), 8 cases(17.0%) of stable disease(SD), and 1 case(2.1%) of progressive disease(PD). By next generation sequencing, 25 genes mutated with 70 times in 33 cases. The mutation frequency of epigenetic regulators(57.6%) was higher than splicing factors(33.5%), transcription factors and kinase signaling(54.5%),and TP53(21.2%)(P<0.01). There was no significant difference in response rates among these patients(47.4%, 54.5%, 50.0% and85.7%, P=0.977). Gene mutation frequency(VAF) of patients who responded to the regimen declined significantly(16.67% vs. 10.26%,P=0.014). CONCLUSION: Decitabine combined with arsenious acid has significant effect in the treatment of patients with higher-risk MDS and CMML and is well-tolerated. Gene mutation test results by next generation sequencing might be related to clinical response.

OBJECTIVE: To study role of TLR4/NF-κB pathway for early change of synovial membrane in knee osteoarthritis rats. METHODS: Eighteen male SD rats weighted (200±20) g were randomly divided into 2 groups, namely control and model group, and 9 in each group. Knee OA model group was established by using modified Hulth method in model group. Control group was not treated. Synovial tissue and serum was extracted at 4 and 21 d after operation. Expression of CD14, TLR4, IL-1β, TNF-α, ADAMTS-4, MMP-13 were detected by real-time PCR respectively. NF-κB p65 protein was detected by Western-blot; serum concentrations of haluronic acid (HA), N-propeptide of type III procollagen(PIIINP) was detected by Elisa. RESULTS: Expression of CD14, ADAMTS-4, and NF-κB p65 in model group were higher than that of control group at 4 and 21 days after operation, while expression of TLR4, IL-1β, TNF-α and MMP-13 were higher than that of control group at 21 days after operation(<0.01). Concentration of PIIINP and HA in model group were higher than that of control group at 4 days after operation, while there was no significant difference at 21 days after operation. CONCLUSIONS NF-κB pathway could mediate occurrence of KOA by early activating and triggeringg synovial increasingly secreting inflammatory secretion CD14, TLR4, IL-1β, TNF-α, ADAMTS-4, MMP-13, PIIINP and HA.
Animals ; Male ; NF-kappa B ; Osteoarthritis, Knee ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Synovial Membrane ; Toll-Like Receptor 4

Objective To compare the clinical efficacy and safety of insulin glulisine (GLU) and insulin aspam(ASP) combined with insulin glargine (GLA) in type 2 diabetes patients (T2DM) with secondary failure to oral hypoglycemic agents.Methods One hundred and twenty cases of T2DM with secondary failure to oral hypoglycemia agents were randomly divided into control group (n =60 cases) and treatment group (n =60 cases).The control group received subcutaneous injection of ASP before 0-10 min three meals daily,and the treatment group received subcutaneous injection of GLU before 0-10min three meals daily.The two groups were treated with subcutaneous injection of GLA,once daily.The initial dose of ASP and GLU were 0.8 U · kg-1 · d-1,and the initial dose of GLA was 0.2 U · kg-1 · d-1.Two groups of patients were hospitalized for 2 weeks.The changes of blood glucose,cases and days of blood glucose of achieving target were compared after treatment of hospitalization.The improvement of HbAlc and the frequency of hypoglycemia were observed after treatment of 12 weeks.Results The days of 2-hour postprandial blood glucose(2 h PG) of achieving target in the treatment group and the control group were (10.01 ± 1.99)d and (10.93 ± 1.52)d with 57 cases and 47 cases,and the days of fasting plasma glucose,postprandial blood glucose and 2 h PG of achieving target were (10.31 ± 1.04) d and (11.03 ± 1.38) d with 48 cases and 40 cases(all P ＜ 0.05).After treatment 12 weeks,the HbAlc in treatment and control groups were (6.78 ±0.59)％ and (7.07 ±0.49)％ without significant difference (P ＞ 0.05).During treatment of 12 weeks,the adverse drug reactions in the two groups were mainly hypoglycemic events,the incidence of hypoglycemia in the treatment group was 30.00％,and 25.00％ in the control group,the difference was not statistically significant (P ＞ 0.05).Conclusion Both GLU combined with GLA or ASP combined with GLA have similar efficacy and safety for T2DM with secondary failure to oral hypoglycemic agents.GLU combined with GLA is more rapid in controlling 2 h PG of these patients.

OBJECTIVETo examine the role of Cd-induced reactive oxygen species (ROS) generation in the apoptosis of neuronal cells.

METHODSNeuronal cells (primary rat cerebral cortical neurons and PC12 cells) were incubated with or without Cd post-pretreatment with rapamycin (Rap) or N-acetyl-L-cysteine (NAC). Cell viability was determined by MTT assay, apoptosis was examined using flow cytometry and fluorescence microscopy, and the activation of phosphoinositide 3'-kinase/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) and mitochondrial apoptotic pathways were measured by western blotting or immunofluorescence assays.

RESULTSCd-induced activation of Akt/mTOR signaling, including Akt, mTOR, p70 S6 kinase (p70 S6K), and eukaryotic initiation factor 4E binding protein 1 (4E-BP1). Rap, an mTOR inhibitor and NAC, a ROS scavenger, blocked Cd-induced activation of Akt/mTOR signaling and apoptosis of neuronal cells. Furthermore, NAC blocked the decrease of B-cell lymphoma 2/Bcl-2 associated X protein (Bcl-2/Bax) ratio, release of cytochrome c, cleavage of caspase-3 and poly(ADP-ribose) polymerase (PARP), and nuclear translocation of apoptosis-inducing factor (AIF) and endonuclease G (Endo G).

CONCLUSIONCd-induced ROS generation activates Akt/mTOR and mitochondrial pathways, leading to apoptosis of neuronal cells. Our findings suggest that mTOR inhibitors or antioxidants have potential for preventing Cd-induced neurodegenerative diseases.

Animals ; Apoptosis ; drug effects ; Cadmium ; toxicity ; Caspases ; metabolism ; Mitochondria ; drug effects ; Neurons ; drug effects ; PC12 Cells ; Proto-Oncogene Proteins c-akt ; metabolism ; Rats ; Rats, Sprague-Dawley ; Reactive Oxygen Species ; metabolism ; Signal Transduction ; drug effects ; TOR Serine-Threonine Kinases ; metabolism

OBJECTIVETo compare Cirsium japonicum characteristics with C. leo and C. leducei, along with the content of buddleoside and pectolinarin, and lay the foundation for the quality control of C. japonicum.

METHODSamples were collected and the relevant drugs were bought. The samples were divided into root, stem, leaf and flower, and the content of buddleoside and pectolinarin was determine by the HPLC. Chromatographic column: Waters XBridge C18 (4.6 mm x 250 mm), mobile phase: methanol-water (45: 55), measurement wavelength: 326 nm, flow rate: 0.8 mL x min(-1), column temperature: 30 degrees C. RESULT AND CONDUSION: Standard curve equation of buddleoside: Y = 74 064X-47 748, R2 = 0.991. Standard curve equation of pectolinarin: Y = 1 711 64X - 180 707, R2 = 0.999. The content of buddleoside: C. japonicum leaf was 1.987 3%, C. leo leaf 1.412 2%, C. leducei leaf 0.149 2%. The content of buddleoside was lower in root and stem. Pectolinarin was not detected in the C. japonicum and C. leo. The pectolinarin content was 0.069 0% in C. leducei leaf.

Chromatography, High Pressure Liquid ; Chromones ; analysis ; chemistry ; Cirsium ; chemistry ; Drugs, Chinese Herbal ; analysis ; chemistry ; Reproducibility of Results ; Solubility ; Species Specificity

BACKGROUNDAblation of complex fractionated atrial electrograms (CFAE) is an important adjunctive therapy in atrial fibrillation (AF). The present study was to elucidate the substrate underlying CFAE.

METHODSNine adult mongrel dogs were involved in the present study. AF was induced through rapid atrial pacing with vagosympathetic nerve stimulation. CFAE was recorded during AF. Ablation was performed at CFAE sites. Based on the location of the ablation scar, the atrial specimens were divided into CFAE and non-CFAE sites. Serial sections of the atrium were stained respectively with hematoxylin-eosin (HE) and the general neural marker protein gene product 9.5 (PGP9.5). We compared the characteristics of the myocardium and the ganglionated plexus (GPs) distribution between the CFAE and non-CFAE sites.

RESULTSThe myocardium of non-CFAE sites was well-organized with little intercellular substance. However, the myocardium in the CFAE site was disorganized with more interstitial tissue ((61.7 ± 24.3)% vs. (34.1 ± 9.2)%, P < 0.01). GPs in the CFAE site were more abundant than in non-CFAE sites ((34.45 ± 37.46) bundles/cm(2) vs. (6.73 ± 8.22) bundles/cm(2), P < 0.01).

CONCLUSIONThe heterogeneity of the myocardium and GPs distribution may account for the substrate of CFAE and serve as a potential target of ablation.

Animals ; Atrial Fibrillation ; pathology ; Dogs ; Electrophysiologic Techniques, Cardiac ; methods ; Myocardium ; pathology