[Novel frameshift mutations in SALL4 in two Chinese families with Okihiro syndrome].

In the present study, clinical manifestations of two Chinese Okihiro syndrome families were analyzed, and genetic detections were performed on the two probands by exome sequencing and verified by Sanger sequencing for family members to determine the biological pathogenesis. Prenatal diagnoses were provided for three high-risk fetuses. The affected members exhibited a wildly spectrum of phenotypes, including ultrasound abnormalities of skeletal system (radius deformity and abnormal posture), and cardiac system (persistent common arterial trunk and ventricular septal defect) in the prenatal period of family 1, the severe phenotypes (grossly shortened and deformed forearm, Duane's anomaly and hearing loss), and the mild ones (usually only thenar dysplasia, or short radius styloid process). Two SALL4 variants, c.844delC p.(Q282Kfs*8) and c.2210delG p.(G737Vfs*23), have been identified respectively in two probands, and c.2210delG of SALL4 gene was unreported previously. The two variants were verified in all affected individuals, not in normal family members. Genotyping results of three fetuses indicated that one fetus was normal, and the two fetuses with heterozygous variation were affected. The two variants of SALL4 gene, c.844delC p.(Q282Kfs*8) and c.2210delG p.(G737Vfs*23), were the molecular pathological cause of Okihiro syndrome in the present study and enriched the spectrum of SALL4 variants. Our study provides accurate prenatal genetic diagnosis for the two families to avoid the birth of affected children.

[Clinical value of emergency endovascular embolization in the interventional treatment for oral hemorrhage caused by carcinoma].

To evaluate the clinical value of emergency endovascular embolization in the interventional treatment for oral hemorrhage caused by carcinoma, 32 patients with oral hemorrhage caused by carcinoma, who received emergency endovascular embolization due to unsatisfactory hemostatic effect of conventional conservative treatment in the First Affiliated Hospital of Zhengzhou University from January 2014 to December 2019, were included in this study and their clinical data, laboratory data and imaging information were retrospectively analyzed. There were 16 males and 16 females, aged (60.6±13.6) years (34-88 years). Technical successful rate of emergency endovascular embolization, immediate successful rate of controlling hemorrhage, blood pressure before and after operation, hemoglobin before and after operation, postoperative complications and recurrence rate of oral hemorrhage were statistically analyzed. Results showed that technical successful rate of operation and immediate successful rate of controlling oral hemorrhage are both 100% (32/32). Recurrent oral hemorrhage occurred in 4 patients (13%). The hemorrhagic shock symptoms of all patients were significantly improved after interventional therapy. After operation, local swelling happened in 34% (11/32) patients and intermittent local pain happened in 22% (7/32) within 24 hours; the swelling and the pain gradually disappeared from 2nd to 5th days. Mild complications of transient fever happened in 9% (3/32) patients and disappeared spontaneously in the short term. No serious complications such as blindness, cerebrovascular accident or central nervous system disturbance occurred in all patients after operations. During the whole follow-up period (1 to 12 months), a total of 8 patients died. The causes of death were progression and metastasis of carcinoma (n=4), heart failure (n=2), severe pneumonia (n=1) and respiratory failure caused by recurrent oral hemorrhage (n=1). Owing to the remarkable short-term curative effect, repeatable operation, low recurrence rate of oral hemorrhage and low incidence of complications, emergency endovascular embolization can be used in the clinical therapy and application of oral hemorrhage caused by carcinoma.

The protective effect of Metformin/Donepezil in diabetic mice brain: evidence from bioinformatics analysis and experiments.

OBJECTIVE: To identify candidate differentially expressed genes (DEGs) and pathways in diabetic mice brain with metformin/donepezil, network pharmacology analysis was used and verified by experiments. MATERIALS AND METHODS: We analyzed GSE62013 microarray datasets derived from the Gene Expression Omnibus (GEO) database for diabetic brain. Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were performed using the Database. Subsequently, the protein-protein interaction network (PPI) and Cytoscape were used for visualizing the most significant module and hub genes. Metformin/donepezil were used to treat streptozotocin (STZ)-induced diabetic mice models. Blood glucose levels and Morris water maze test were measured. The apoptotic rate of diabetic brain tissue was analyzed using Annexin V/propidium iodide double staining. The levels of PI3K and AKT in the mice brain tissues were detected by Western blot. RESULTS: DEGs included 214 up-regulated genes and 100 down-regulated genes in diabetic brain tissues of mice. The enriched GO functions were multicellular organism development, negative regulation of transcription from RNA polymerase II promoter, and extracellular region. The enriched pathways were PI3K-Akt signaling pathway, Linoleic acid metabolism and Arachidonic acid metabolism. Blood glucose levels and apoptosis were reduced in STZ-induced diabetic mice following metformin/donepezil treatment. Metformin/donepezil could reverse this neurocognitive deficiency. Protein levels of PI3K and AKT were significantly increased in STZ-induced diabetic mice. CONCLUSIONS: Overall, we proposed that 10 genes (Cdc20, Fbxo32, Igtp, Atg7, Fbxo15, Trim37, Psmb8, Ifi47, Asb12, and Asb5) that might be novel hub genes strongly associated with diabetic mice brain. Metformin/donepezil ameliorates STZ-induced brain injury by activating the PI3K/AKT pathway and alleviating apoptosis.

[Clinical application and efficacy of TIPS combined with AngioJet mechanical thrombectomy for liver cirrhosis with extensive portal vein thrombosis].

Objective: To assess the safety and efficacy of transjugular intrahepatic portosystemic shunt (TIPS) combined with AngioJet mechanical thrombectomy for liver cirrhosis with extensive portal vein thrombosis. Methods: From March 2018 to April 2019, a total of 11 patients with liver cirrhosis and extensive portal vein thrombosis were treated by TIPS combined with AngioJet mechanical thrombectomy, including 6 males and 5 females, with the age of 37-71 (46±9) years old, 3 cases of Child-Pugh grade A, 8 cases of grade B and 0 cases of grade C. The intraoperative immediate thrombus clearance rate, perioperative complication rate, postoperative thrombus recurrence rate, rebleeding rate, the incidence of hepatic encephalopathy and the rate of stent patency of all cases were collected and analyzed. Results: All the patients were treated successfully. The immediate complete thrombus clearance (grade Ⅲ) rate of portal vein trunk was 9/11, and grade Ⅱ was 2/11, The average dose of urokinase was 30-60 (40±5) ten thousand U, slight puncture point bleeding occurred in 3 cases, and recurrence of PVT in portal vein trunk occurred in 1 case with Ⅱ grade clearance rate after operation, rebleeding occurred in 1 case, hepatic encephalopathy occurred in 2 cases, the primary patency rate of stents was 9 cases. Conclusion: TIPS combined with AngioJet mechanical thrombectomy can treat the liver cirrhosis with extensive portal vein thrombosis effectively and safely, and postoperative portal vein patency rate and intrahepatic shunt patency rate are high.

[Application of real-time image fusion technique in transjugular intrahepatic portosystemic shunt].

Objective: To assess the feasibility and value of real-time image fusion technique guiding the procedure of transjugular intrahepatic portosystemic shunt(TIPS). Methods: From July 2017 to May 2018,a total of 48 consecutive patients complicated by portal venous hypertension due to cirrhosis who underwent TIPS were prospectively allocated into two groups that 27 cases underwent normal TIPS and 21 cases underwent image fusion guided TIPS. There were 25 males and 23 females with a mean age of 29-74(51±10) years. The differences of portal vein(PV) between image fusion angiographyand digital subtraction angiography(DSA), and the times of puncture PV, X-ray exposure dose and exposure time and contrast agent amount of all cases were collected and analyzed. Results: The longitudinal and traverse difference of PV between image fusion angiography and DSA were 1.7-2.5(2.1±0.2) mm and 0.9-1.8(1.4±0.3) mm, respectively.The times of puncture PV, X-ray exposure time and dose, and contrast agent amount between normal TIPS group and image fusion guided TIPS group were 1-7(3.8±0.6) times vs 1-3(2.0±0.6) times, 41-63(53±8)min vs 27-42(35±5) min, 513-787(644±96) mGy vs 357-524(423±59) mGy,102-196(151±23) ml vs 87-145(105±14) ml(all P<0.05), respectively. Conclusions: There are minor differences between image fusion angiography of PV and DSA. Real-time image fusion guided TIPS is feasible and valuable to reduce intraprocedural X-ray exposure time and dose and contrast agent amount of TIPS.

[Effect of interleukin-7 on the anti-tumor function of CD8(+) T cells in patients with non-small cell lung cancer].

Objective: To investigate the effect of interleukin (IL)-7 on anti-tumor activity of CD8(+) T cells in non-small cell lung cancer (NSCLC) patients. Methods: Twenty-seven NSCLC patients and ten healthy controls were included from Zhengzhou Central Hospital between January 2017 and July 2017. Plasma, peripheral blood mononuclear cells (PBMCs), and bronchial alveolar lavage fluid (BALF) (both tumor site and non-tumor site) were collect from NSCLC patients, while plasma and PBMCs were also collected from healthy controls. IL-7 level and IL-7 receptor α (CD127) mRNA relative expression was measured. Purified CD8(+) T cells and primary NSCLC cells were stimulated with recombinant IL-7. Cellular proliferation, cytokines secretion, and protein expression in IL-7 signaling pathway were investigated. Direct/indirect contact coculture system of CD8(+) T cells and primary NSCLC cells was also used to assess the cytolytic and noncytolytic activity after IL-7 stimulation. Results: Plasma IL-7 level was significantly reduced in NSCLC patients compared with normal controls[1 731 (1 364, 2 536) vs 2 686 (1 692, 4 786) ng/L, P=0.023). IL-7 level in BALF isolated from tumor site was also remarkably down-regulated compared with non-tumor site in NSCLC patients[1 045 (562, 1 550) vs 1 599 (1 166, 2 107) ng/L, P=0.006 9). There was no statistical difference of CD127 mRNA between the two groups. Recombinant IL-7 stimulation did not affect cellular proliferation and cytokines production in primary NSCLC cells, however, remarkably increased cellular proliferation, interferon (IFN)-γ/tumor necrosis factor (TNF)-α secretion, and signal transducers and activators of transcription 5 (STAT5) phosphorylation and suppressor of cytokine signaling 3 (SOCS3) expression of CD8(+) T cells from NSCLC patients. IL-7 stimulation also significantly enhanced the cytolytic and noncytolytic function of CD8(+) T cells on primary NSCLC cells. Conclusion: IL-7 enhances anti-tumor activity of CD8(+) T cells in NSCLC patients.

Effect of dietary energy restriction and subsequent compensatory feeding on testicular transcriptome in developing rams.

Nutritional intake and reproductive allocation are strongly associated and dietary energy restriction (ER) or surpluses can affect reproductive capacity. The objective of this study was to investigate the effect of energy levels on sheep testicular development. Three-month old Hu sheep were assigned to four groups, and fed diets containing different levels of energy (Control, maintenance energy; ER1, 85% maintenance energy; ER2, 70% maintenance energy; ER3, 55% maintenance energy). Two months later, half the sheep in each group were euthanized, whereas the remaining sheep were euthanized after a further 3 months feeding on a compensatory energy diet. The testicular weight and reproductive hormone levels of the Hu sheep were investigated. Differences in the testes of ER3 and control group sheep were investigated at the transcriptional level using high-throughput sequencing. The results showed that the testicular weights had decreased in the energy-restricted rams compared with the controls, and that the testosterone concentration in ER3 group rams was significantly lower than that in other compared groups (P < 0.05). After the period of compensatory feeding, however, ER3 sheep testicular weight and testosterone concentrations were similar to those of the control group sheep. In addition, the RNA sequencing results revealed that 81 genes were upregulated and 180 genes were downregulated in the ER3 group compared with the control group. Moreover, based on the enriched steroidogenesis, meiosis and kinases pathways, a number of candidate genes potentially involved in the regulation of testicular development or reproduction of Hu sheep, including CYP11A1, ALDH3B1, FDFT1, WNT2, PGR and INSR, were screened. Quantitative real-time polymerase chain reaction analysis results correlated well with the sequencing data. Taken together, this study provides a first insight into the development of the testis with dietary energy restriction in sheep and shows that these changes are associated with alterations in transcriptomic. The sheep testis mRNA database were extended in this study will provides novel candidate regulators for future genetic and molecular studies on sheep testicular development associated with energy restriction, which will contribute to improving the reproductive performance of sheep.

Calpain-activated mTORC2/Akt pathway mediates airway smooth muscle remodelling in asthma.

BACKGROUND: Allergic asthma is characterized by inflammation and airway remodelling. Airway remodelling with excessive deposition of extracellular matrix (ECM) and larger smooth muscle mass are correlated with increased airway responsiveness and asthma severity. Calpain is a family of calcium-dependent endopeptidases, which plays an important role in ECM remodelling. However, the role of calpain in airway smooth muscle remodelling remains unknown. OBJECTIVE: To investigate the role of calpain in asthmatic airway remodelling as well as the underlying mechanism. METHODS: The mouse asthma model was made by ovalbumin sensitization and challenge. Calpain conditional knockout mice were studied in the model. Airway smooth muscle cells (ASMCs) were isolated from smooth muscle bundles in airway of rats. Cytokines IL-4, IL-5, TNF-α, and TGF-ß1, and serum from patients with asthma were selected to treated ASMCs. Collagen-I synthesis, cell proliferation, and phosphorylation of Akt in ASMCs were analysed. RESULTS: Inhibition of calpain using calpain knockout mice attenuated airway smooth muscle remodelling in mouse asthma models. Cytokines IL-4, IL-5, TNF-α, and TGF-ß1, and serum from patients with asthma increased collagen-I synthesis, cell proliferation, and phosphorylation of Akt in ASMCs, which were blocked by the calpain inhibitor MDL28170. Moreover, MDL28170 reduced cytokine-induced increases in Rictor protein, which is the most important component of mammalian target of rapamycin complex 2 (mTORC2). Blockage of the mTORC2 signal pathway prevented cytokine-induced phosphorylation of Akt, collagen-I synthesis, and cell proliferation of ASMCs and attenuated airway smooth muscle remodelling in mouse asthma models. CONCLUSIONS AND CLINICAL RELEVANCE: Our results indicate that calpain mediates cytokine-induced collagen-I synthesis and proliferation of ASMCs via the mTORC2/Akt signalling pathway, thereby regulating airway smooth muscle remodelling in asthma.

Clinical study of the correlation between complement factor H polymorphism and age-related macular degeneration.

This study aimed to investigate the correlation between age-related macular degeneration (AMD) of the liver-kidney yin-deficiency type and complement factor H (CFH) polymorphism, and to determine whether the C allele of the T1277C (Y402H) variant is a risk factor for this condition. We performed a case-control investigation of 60 patients with liver-kidney yin-deficiency AMD and 60 normal control subjects. Peripheral blood was collected from each participant for DNA extraction. Following amplification by polymerase chain reaction, the DNA samples were sequenced, and polymorphism of the CFH gene was examined. Data were analyzed with the chi-square test, with P < 0.05 signifying statistical significance. The frequency of the C allele was significantly higher in the wet than in the dry AMD group (P = 0.044). In addition, the TC and CC genotypes were markedly more common in the former than in the control group (P = 0.013), and there was a significant difference in the distribution of the T and C alleles between wet AMD patients and control subjects (P < 0.05). Based on this, we conclude that liver-kidney yin-deficiency AMD is associated with the C allele and TC and CC genotypes of the CFH Y402H polymorphism. Among patients with this condition, CFH genotypes were normally distributed. The principal CFH genotypes that induce liver-kidney yin-deficiency AMD are the mutant homozygote CC and heterozygote TC forms. Moreover, C allele carriers are at higher risk of developing this disease.

[Combination of B-type brain natriuretic peptide with left ventricular diastolic dysfunction for prediction of mortality in the patients with septic shock].

OBJECTIVE: To evaluate the prognostic value of combination of plasma brain natriuretic peptide(BNP) with the ratio of early diastolic mitral inflow velocity to early diastolic mitral annulus velocity(E/E') in the patients with septic shock. METHODS: From June 2012 to December 2013 , 127 patients with septic shock were consecutively recruited and underwent trans-thoracic echocardiography examination within 6 h after admission to Intensive Care Unit(ICU), Zhejiang Provincial People's Hospital. Plasma BNP concentration was measured using ELISA method. All Clinical, laboratory, and survival data were prospectively collected. RESULTS: Of 127 patients enrolled, mean values for age were(59.9±17.3) years and APACHE Ⅱ score(16.8±5.8), respectively. 95 patients(74.8%) took mechanical ventilation. 28- , 60-day mortality rate was 36.3% and 42.3%, respectively. Univariate Cox regression analysis showed that age, coronary artery disease, serum creatinine and lactate, plasma BNP, left ventricular ejection fraction(LVEF), E/E' and APACHE Ⅱ score were significantly(P≤0.05) associated with 60-day mortality. Multivariate analysis revealed that serum lactate, plasma BNP(χ(2)=9.4, P=0.002) , E/E'(χ(2)=4.89, P=0.02) and APACHE Ⅱ score(χ(2)=10.6, P=0.001) remained independent predictors for 60-day mortality. ROC curve analysis showed that the optimal plasma BNP and E/E' cutoff values identified were 338.8 pg/ml and 10.8, and the areas under ROC curve were 0.89(sensitivity: 83.7%; specificity: 81.4%)and 0.83(sensitivity: 76.7%; specificity: 72.9%)for 60-day mortality, respectively. In addition to plasma BNP and clinical predictors, the E/E' could provide in independent and incremental prognostic value of 60-day mortality(χ(2)=59.3 vs 47.8, P<0.001). CONCLUSION: Plasma BNP and E/E' are independent predictors for 60-day mortality, and combination of plasma BNP and E/E' could improve risk stratification in patients with septic shock.

Clinical effect of intravitreous injection of triamcinolone acetonide in treating cystoid macular edema.

Cystoid macular edema (CME), a commonly seen sign for multiple fundus diseases, is able to induce visual deterioration. The incidence rate of CME is constantly increasing; however, the existing clinical treatments cannot achieve satisfactory curative effects. To explore the curative effect of intravitreous injection of triamcinolone acetonide (TA) in treating CME, this study carried out a clinical test on 39 patients (42 eyes) from The First Affiliated Hospital of Zhengzhou University who developed CME induced by central retinal vein occlusion (CRVO). All 42 eyes received intravitreous injection of 40 mg/ml TA (0.1 ml) and then were followed up for 11-23.5 months. Eyes were examined by slit-lamp microscope, fundus fluorescein angiography (FFA) and optical coherence tomography (OCT) and best corrected visual acuity (BCVA), and intraocular pressure (IOP) of those eyes were detected before and after treatment. Average vision of eyes was 0.1 before treatment, and the vision improved in one month (vision ≥ 0.2: 100%; vision ≥ 0.5: 42.9%) and three months (vision ≥ 0.2: 64.3%; vision ≥ 0.5: 21.4%) after treatment; but as time went on, the vision of some patients declined; at the last follow-up, patients with vision ≥ 0.2 accounted for 28.6% and those with vision ≥0.5 accounted for 7.1%; compared to before treatment, 71.4% patients had improved vision and the remaining 28.6% had declined vision. Some patients were observed with high IOP during treatment, and 7 eyes were found with secondary cataract in posterior capsule of lens at the last follow-up. Intravitreous injection of triamcinolone acetonide proved to have significant short-term curative effect on CEM which is non-sensitive to conventional therapies, but it is likely to induce high IPO and posterior capsular opacification.

Microdialysis: a technique for pharmacokinetic-pharmacodynamic studies of oncological drugs.

Solid tumors have several barriers that may limit drug penetration and provide inherent mechanisms of resistance. Therefore, the evaluation of antineoplastic agents should base on tumor drug exposure and antitumor activity. Owing to selective access to the extracellular space of the tumor, which acts as target site for most antineoplastic drugs, monitoring drug disposition and corresponding antitumor activity, microdialysis (MD), a probe-based sampling technique, satisfies regulatory requirements for pharmacokinetic- pharmacodynamic (PK-PD) studies. MD is a powerful tool for PK-PD studies of oncological drugs, which would allow better understanding of exposure-response relationships and contributes to the research and development of oncological drugs. Recent progresses in the development of MD for estimating PK-PD studies of oncological drugs are summarized in this review. Special preclinical application examples of MD for PK-PD studies of oncological drugs organized by types of antineoplastic drugs and informations collected are also described. In conclusion, the role of MD in preclinical PK-PD studies of oncological drugs has been confirmed and it has provided a strong foundation for further clinical research.

Clonality in wild rice (Oryza rufipogon, Poaceae) and its implications for conservation management.

Correlations were examined between habitat characters and clonal structures determined by the RAPD (random amplified polymorphic DNA) assay in five populations of Oryza rufipogon in China. Nine of 175 decameric primers were used in the study because they reproducibly amplified polymorphisms. The extent of clonality together with the clonal and sexual reproductive strategies varied greatly among the five populations and correlated with the habitats where they occur. The populations under serious disturbance or seasonal drought tended to have small clones with relatively high clonal diversity caused by sexual reproduction, whereas the populations with little disturbance and sufficient supply of water were prone to have large clones with relatively low clonal variation and low sexual reproduction. Therefore, the dynamics of sexual vs. clonal reproduction of this species depended mainly on environmental factors, such as external disturbance and water supply, rather than latitudes indicated by previous study. These results have important implications for in situ conservation of O. rufipogon. Adequate external disturbance and water supply control are essential for maintaining high clone diversity of in situ conserved populations. According to the extent of clonality of the populations examined, we recommend that an interval of >12 m should be required for collecting samples for ex situ conservation and for population genetic studies to capture possible genetic diversity for O. rufipogon in China.

[Effects of hypoxic acclimation on oxygen consumption of skeletal muscles in rats with frostbite].

AIM AND METHODS: Oxygen consumption of gastrocnemius in rats of frostbite at normoxia (FN), frostbite during acute hypoxia (FAH) and frostbite during hypoxia after hypoxic acclimation (FHAC) groups was investigated with a system of oxygen consumption measurement before and after the rats' right hind legs were frozen in order to provide theoretical data for exploring the reasons that hypoxic acclimation could aggravate tissue damage of animals wth frostbite. RESULTS: Before freezing, oxygen consumption of gastrocnemius in rats of FHAC group decreased by 26.2%. At the first day after freezing, oxygen consumption of gastrocnemius in rats of FN, FAH and FHAC groups was 5.93 +/- 0. 66, 4.74 +/- 1.87 and 0.76 +/- 0.39 [kPa/(g x min)], and decreased by 76.3%, 77.9% and 95.9% respectively as compared with themselves before freezing, and that of FHAC group was much lower than those of FN and FAH groups. At the fifth day after freezing, oxygen consumption of gastrocnemius of the three groups returned to 62.8%, 44.9% and 28.8% of those themselves before freezing respectively, and that of FHAC group was much smaller than those of FN and FAH groups, showing that post-frostbite metabolic rate of skeletal muscle of rats acclimated to hypoxia decreased further and restored much slowly. CONCLUSION: Both hypoxic acclimation and frostbite could decrease the oxygen consumption of gastrocnemius in rats, the complex of those two effects on oxygen consumption made the oxygen consumption to decrease further, suggesting that the changes in tissue metabolism could be one of the main factors that affected the degree of tissue damage and recovery.

[The change of vWF in vascular endothelial cells under different stress].

AIM: To correlate the injury of vascular endothelial cells during various pathological conditions with the change of vWF (von Willebrand Factor) in different VEC lines. METHODS: Flow cytometer(FCM) were used to defect the immunoflourescent stained vWF in pulmonary artery endothelial cells (PAEC) of pig and aortic endothelial cells(AEC) of rats. RESULTS: The positive rates of vWF in PAEC of pigs is similar with that in AEC of rats under normal condition, but it decreased differently after hypoxic or cold injury. It was very interesting that the mean fluorescence intensity of positive PAEC or AEC exposed to hypoxia or cold elevated significantly compared with those of control. CONCLUSIONS: The change of vWF in VEC can be used to evaluate the function of VEC under different stress.

Monoclonal antibody 3F3 against conformational epitope of Torpedo acetylcholinesterase.

AIM: To study the type of epitope of native Torpedo acetylcholinesterase (AChE) directed by its monoclonal antibody (McAb) 3F3. METHODS: Enzyme-linked immunosorbent assay (ELISA) was used for the assay of the reaction between antigen and antibody. RESULTS: McAb 3F3 immunoreacted well with the native AChE, but not with the reduced- and alkylated-AChE (RA-AChE) at all. Soman did not interfere the binding of 3F3 with AChE molecule. The synthesized 24-peptide containing the active serine residue of the AChE active center did not react with McAb 3F3. CONCLUSION: 3F3 is a monoclonal antibody against the conformational epitope of Torpedo AChE active center, but dose not occupy the active serine residue of the enzyme.

Anionic subsite of active center of Torpedo acetylcholinesterase constructs a part of its conformational epitope.

AIM: To study the structure-activity relationship of Torpedo acetylcholinesterase (AChE) and explore whether the anionic subsite of the active center is a constituent of the conformational epitope of enzyme. METHODS: Using ELISA and enzyme inhibition test to examine the effect of 1-methyl-2-hydroxyiminomethylpyridium chloride (2-PAM), an anionic subsite probe of AChE, on the immunoreactivity between Torpedo AChE and its monoclonal antibody (McAb) 3F3. RESULTS: McAb 3F3 did not react with 2-PAM-AChE complex. 2-PAM decreased the inhibitory rate of McAb 3F3 on AChE in a concentration-dependent fashion, but did not dissociate the McAb 3F3-AChE complex. CONCLUSION: Anionic subsite of the active center of Torpedo AChE constructs a part of its conformational epitope.