Identification of hub genes and pathways in the development of gastric cancer by gene co­expression network analysis.

There are many risk factors for gastric cancer (GC), including chronic atrophic gastritis, which involves multiple genes and signaling pathways. Weighted gene co-expression network analysis (WGCNA) was performed on GSE111762 to construct free-scale gene co-expression networks and identified four significant modules that consisted of blue, dark orange, dark red and dark violet. In each module, genes with the most connectivity were selected as hub genes, including G antigen 12J (GAGE12J) in blue, proline, histidine and glycine rich 1 (PHGR1) in dark orange, DNA polymerase gamma 2, accessory subunit (POLG2) in dark red and collagen type XXI alpha 1 chain (COL21A1) in dark violet. The transcription level of COL21A1 and GAGE12J was up-regulated in atrophic gastritis vs normal gastric mucosa, but down-regulated in GC vs atrophic gastritis. PHGR1 was consistently down-regulated from normal gastric mucosa to GC, while POLG2 was up-regulated. Gene set enrichment analysis (GSEA) was then conducted to study the biological functions of hub genes in the development of GC. It showed that multiple tumorigenesis-related pathways were enriched, including peroxisome, DNA repair and KRAS signaling pathway in COL21A1, IL6-JAK-STAT3, epithelial mesenchymal transition (EMT) and TNFα-NF-κB signaling pathway in PHGR1, MYC targets, E2F targets and angiogenesis in POLG2 and peroxisome, Notch signaling pathway and androgen response in GAGE12J. The identified four genes, especially for COL21A1, PHGR1 and POLG2, were important in GC tumorigenesis and affected many cancer-related pathways.

[Clinical study on the effects of global end-diastolic volume index-directed fluid resuscitation on the prognosis of chronic heart failure patients with septic shock].

Objective: To investigate the effect of global end-diastolic volume index (GEDI)-guided fluid resuscitation on the prognosis of patients with chronic heart failure and septic shock. Methods: This study was a prospective randomized controlled study. Consecutive eligible patients were divided into 2 groups according to the random number table method: control group (n=21) and experimental group (n=20). On the basis of routine treatment, patients in the control group received early goal-directed therapy until the central venous pressure (CVP) reaching 8-12 mmHg (1 mmHg=0.133 kPa), mean arterial pressure reaching over 65 mmHg, urine volume reaching over 0.5 ml·kg(-1)·h(-1), and central venous oxygen saturation reaching more than 70%. On the basis of routine treatment, patients in the experimental group were monitored continuously on cardiac output with pulse indication and fluid resuscitation guided by volume index GEDI. The GEDI should be maintained on the range of 680-800 ml/m(2). The remaining resuscitation goals were the same as control group. General clinical data of the two groups were collected at admission. Negative fluid balance onset time, duration of mechanical ventilation, ICU mortality and 28-day mortality were compared between the two groups. The outcomes were recorded as listed: start time of negative fluid balance, duration of mechanical ventilation, mortality in ICU and 28-day mortality. Results: There was no significant difference in age, sex, weight, APACHE Ⅱ score, SOFA score and NYHA functional class score between the two groups (all P>0.05). The negative liquid balance onset time in the control group was 3.5 (2.5, 4.0) days, which was significantly longer than that in the experimental group (2.6 (2.0, 3.0) days,U=115.0, P=0.012). The duration of mechanical ventilation was 355 (118, 552) hours in the control group, which was significantly longer than that in the experimental group (132 (36.75, 233.3) hours, U=130, P=0.038). The ICU mortality was 38.1% (8/21) in the control group, tended to be higher than that in the experimental group (20.0%(4/20), χ(2)=1.620, P=0.203). The 28-day mortality was 42.9% (9/21) in the control group, similar as in the experimental group (25.0%(5/20), χ(2)=1.482,P=0.477). Conclusion: Fluid resuscitation guided by volume index (GEDI) may improve the prognosis of patients with chronic heart failure complicated with septic shock.

Antioxidants add protection to a broad-spectrum sunscreen.

BACKGROUND: Exposure of human skin to ultraviolet radiation (UVR) results in erythema, pigment darkening, skin cancer and photoageing. In addition to conventional organochemical and the physical-mineral type sunscreens (SS), other non-SS protective strategies have been investigated, including antioxidants (AOx) and topical DNA repair enzymes. AIM: To investigate whether AOx could improve the protection provided by a broad-spectrum sunscreen (SS) preparation. METHODS: Volunteers were exposed to repetitive solar-simulated (ss)UVR at 1.5 times minimal erythema dose for four consecutive days. Thirty minutes before each exposure and 6, 24 and 48 h after the last exposure, the test materials [vehicle, SS (sun protection factor 25) alone, AOx alone and SS plus AOx] were applied to four different sites. Another two sites received ssUVR only, or SS plus AOx only, and a third site was left untreated (neither ssUVR or product). Erythema and pigmentation were measured using a Mexameter. Biopsy specimens were taken 72 h after the last irradiation. The thickness of the stratum corneum and epidermis were measured by microscopy. Expression of cytokeratins (CKs), matrix metalloproteinases (MMPs) and CD1a-positive Langerhans cells (LCs) analysed by immunohistochemical staining, and relative expression levels were compared between all seven sites. RESULTS: AOx alone did not reduce erythema. There was a significant reduction in pigmentation, and the product almost completely protected against LC depletion. AOx plus SS gave better protection against pigment formation and CK5/6 induction than SS alone. AOx alone protected against ssUVR-induced hyperproliferation, as shown by epidermal thickness and CK16 biomarkers, and was better than SS alone. Interestingly, although protection against induction of MMP-9, a marker of photoageing, did not reach significance when either SS or AOx were applied separately, there was complete protection against MMP-9 induction when these were combined. CONCLUSIONS: Non-SS materials such as AOx can contribute significantly to sun protection when added to a broad-spectrum SS and applied topically to human skin in vivo.

[Analysis of risk factors of early enteral nutrition intolerance in extremely severe burn patients].

Objective: To explore the risk factors of early enteral nutrition intolerance in extremely severe burn patients. Methods: A retrospective case-control study was performed. From January 2018 to December 2020, seventy-six adult patients with extremely severe burns who met the inclusion criteria were admitted to Hwa Mei Hospital of University of Chinese Academy of Sciences, including 55 males and 21 females, aged (45±11) years with burns of 62% (52%, 82%) total body surface area. Depending on the patient's tolerance to early enteral nutrition, they were divided into tolerance group (47 patients) and intolerance group (29 patients), and their clinical data were statistically analyzed, including age, sex, body mass index (BMI), underlying disease, total burn area, full-thickness burn area, abbreviated burn severity index (ABSI) score, implementation of mechanical ventilation on the day of admission, stable shock state, vomiting before feeding. The following data were recorded including the onset time, duration length, and frequency of enteral nutrition intolerance of patients in intolerance group, and the number of operations, the length of hospitalization, the occurrence of sepsis within 2 weeks after injury, the outcome, as well as the serum hypersensitive C-reactive protein (hs-CRP), albumin, fasting blood glucose, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and γ-glutamyl transpeptidase (γ-GT) on post burn day (PBD) 1, 5, 9, and 13 of patients in the two groups. Data were statistically analyzed with independent sample t test, Mann-Whitney U test, and chi-square test to screen the related factors of early enteral nutrition intolerance of the patients. Binary univariate and multivariate logistic regression analysis were used to analyze the independent risk factors of early enteral nutrition intolerance of the patients. Results: There were no statistically significant differences in age, sex, BMI, and percentage of underlying disease of patients between the two groups (P>0.05). The total burn area, full-thickness burn area, ABSI score, mechanical ventilation percentage on the day of admission, percentage of unstable shock period, percentage of vomiting before feeding of patients in intolerance group were significantly higher than those in tolerance group (Z=-4.559, -3.378, -4.067, χ2=18.375, 23.319, 8.339, P<0.01). In intolerance group, the onset time of intolerance was (9±4) d after injury, and the duration length was 4 (2, 6) d, with a total of 46 times occurred. Compared with tolerance group, the percentage of sepsis and mortality of patients in intolerance group were significantly higher within 2 weeks after injury (χ2=16.571, 12.665, P<0.01). The number of operation and length of hospitalization of patients in the two groups were similar (P>0.05); however the length of hospitalization of patients in the intolerance group was significantly more than that in tolerance group after excluding the death cases (Z=-2.266, P<0.05). On PBD 1, the level of fasting blood glucose and AST of patients in intolerance group were significantly higher than those in tolerance group (t=3.070, Z=-3.070, P<0.01). On PBD 5, the levels of hs-CRP, albumin, fasting blood-glucose, ALT, AST, and γ-GT of patients in the two groups were similar (P>0.05). On PBD 9, the level of hs-CRP of patients in intolerance group was significantly higher than that in tolerance group (t=2.836, P<0.01), and the levels of ALT and γ-GT of patients in intolerance group were significantly lower than those in tolerance group (Z=-3.932, -2.052, P<0.05 or P<0.01). On PBD 13, the level of hs-CRP of patients in intolerance group was significantly higher than that in tolerance group (t=3.794, P<0.01), and the levels of fasting blood glucose, ALT, and γ-GT of patients in intolerance group were significantly lower than those in tolerance group (t=-2.176, Z=-2.945, -2.250, P<0.05 or P<0.01). Binary univariate logistic regression analysis showed that total burn area, full-thickness burn area, ABSI score, implementation of mechanical ventilation on the day of admission, unstable shock period, vomiting before feeding, and fasting blood-glucose on PBD 1 of patients were related to early enteral nutrition intolerance (odds ratio=1.086, 1.052, 1.775, 9.167, 12.797, 10.125, 1.249, 95% confidence interval=1.045-1.129, 1.019-1.085, 1.320-2.387, 3.132-26.829, 4.199-39.000, 2.003-51.172, 1.066-1.464, P<0.01). Multivariate logistic regression analysis showed that the large total burn area, unstable shock period, vomiting before feeding, and high fasting blood-glucose on PBD 1 of patients were the independent risk factors of early enteral nutrition intolerance in patients (odds ratio=1.073, 6.390, 9.004, 1.246, 95% confidence interval=1.021-1.128, 1.527-26.734, 1.134-71.496, 1.007-1.540, P<0.05 or P<0.01). Conclusions: The percentage of early enteral nutrition intolerance is very high in extremely severe burn patients, which is closely related to poor prognosis. Large total burn area, vomiting before feeding, unstable shock phase, high fasting glucose on PBD 1 of patients are the independent risk factors for early enteral nutrition intolerance in extremely severe burn patients. The benefits and risks should be carefully evaluated before starting enteral nutrition in such patients, and early enteral nutrition should not be blindly pursued.

[Analysis of the dynamic changes in gut microbiota in patients with extremely severe burns by 16S ribosomal RNA high-throughput sequencing technology].

Objective: To analyze the dynamic change rule of gut microbiota in patients with extremely severe burns using 16S ribosomal RNA (rRNA) high-throughput sequencing technology. Methods: Five patients with extremely severe burns who were admitted to Hwa Mei Hospital of University of Chinese Academy of Sciences from February to June 2017 and conformed to the inclusion criteria were included in the prospective observational study. All patients were males with age of 32-48 years. Fecal samples were collected in the shock stage (within 3 days after injury), early stage of acute infection (4-14 d after injury), middle stage of acute infection (15-28 d after injury), late stage of acute infection (from 29 d after injury to 1 week before discharge) and within 1 week before discharge. The number of samples was 5 in each stage. The fecal pH value was measured using a pH meter. High-throughput sequencing technology was applied for sequencing of 16S rRNA V3 and V4 regions of fecal samples. QIIME software was used to analyze the number of operational taxonomic units (OTUs), α diversity (Chao1 index and Shannon index), and the relative abundance of gut microbiota at the phylum and family levels. Unweighted pair group method with arithmetic mean clustering method was used to analyze the ß diversity of gut microbiota, and Tax4Fun was used to predict functional changes of gut microbiota. Data were statistically analyzed with one-way analysis of variance for repeated measurement, Bonferroni method, Wilcoxon rank sum test for paired samples, and Bonferroni correction. Results: (1) The pH value of feces in the early and middle stages of acute infection in patients with extremely severe burns in this group was 7.40±0.45 and 7.56±0.45 respectively, which were significantly higher than 6.68±0.36 in the shock stage (P<0.05 or P<0.01). (2) A total of 2 333 584 efficient and high-quality sequences were obtained, and the length of the sequences was about 415 bp. A total of 1 209 OTUs were obtained. The sequencing coverage of all samples was over 99.0%. The number of OTUs and Chao1 index in the early, middle, and late stages of acute infection in patients with extremely severe burns in this group were significantly lower than those in the shock stage (Z=2.023, P<0.05). The number of OTUs and Chao1 index within 1 week before discharge were significantly higher than those in the early, middle, and late stages of acute infection, and Shannon index within 1 week before discharge was significantly higher than that in the early and middle stages of acute infection (Z=2.023, P<0.05). (3) The structure of gut microbiota in the shock stage in patients with extremely severe burns in this group was highly similar to that within 1 week before discharge, and lowly similar to that in the early, middle, and late stages of acute infection. The analysis of individual sample showed that the clustering rule of most of the samples was in accordance with that of the staged samples. The weighted Unifrac distance of gut microbiota in the shock stage was significantly shorter than that in the early, middle, and late stages of acute infection (Z=3.326, 2.570, 2.690, P<0.05 or P<0.01), while the weighted Unifrac distance of gut microbiota in the other stages was similar. (4) At the phylum level, compared with that in the shock stage, the relative abundance of Firmicutes was decreased in the early, middle, and late stages of acute infection, while the relative abundance of Bacteroidetes and Proteobacteria increased. However, the relative abundance of the above three phyla within 1 week before discharge was similar to that in the shock stage. At the family level, the top five dominant bacteria in relative abundance in different stages after injury were quite different. The relative abundance of dominant five family bacteria in the shock stage was decreased in the early, middle, and late stages of acute infection. The relative abundance of non-dominant bacteria such as Enterobacteriaceae, Streptococcaceae, and Bacteroidaceae in the shock stage increased significantly in the early, middle, and late stages of acute infection, which became new dominant families in these stages. The relative abundance of some acid-producing bacteria within 1 week before discharge resumed to the similar level in the shock stage. (5) Functions such as some amino acid metabolism, glycolysis and gluconeogenesis, and pyruvate metabolism of gut microbiota were obviously weaker in the early and middle stages of acute infection than those in the shock stage. Functions such as some amino acid metabolism and carbohydrate metabolism of gut microbiota were significantly enhanced in the late stage of acute infection compared with that in the shock stage. The distributions of functional genes in gut microbiota were similar between the shock stage and within 1 week before discharge. Conclusions: The internal environment and gut microbial compositions in extremely severe burned patients change significantly in the early and middle stages of acute infection. The pH value increases, the bacterial species and diversity decrease, especially the relative abundance of acid-produced bacteria is significantly reduced, which gradually recover with the improvement of the patient's condition. The pH value and the changes of Proteobacteria and acid-producing bacteria could be considered as suitable parameters for reflecting the disorder level of gut microbiota in patients with extremely severe burns.

Lichen planus pemphigoides associated with Chinese herbs.

We report a 62-year-old Chinese woman with a 2-year history of lichen planus presenting with extensive violaceous maculopapules and plaques 1 week after taking an oral preparation of Chinese herbs. The patient developed vesiculobullous skin lesions 7 weeks later. Histopathological examination showed subepidermal blisters and adjacent bandlike lymphocytic infiltration. Direct immunofluorescence revealed linear deposits of IgG and C3 along the basement membrane zone. Indirect immunofluorescence showed IgG antibody deposition along the epidermal side of salt-split human skin. Circulating anti-bullous pemphigoid 180 antibodies were detected by ELISA. Lichen planus pemphigoides (LPP) was diagnosed. To our knowledge, this is the first report of LPP associated with oral Chinese herbs.

Correlation of radiobiological assays of hypoxic fraction with phosphorus-31 magnetic resonance spectroscopy across multiple tumor lines.

Hypoxic fractions were measured in multiple tumor models and compared to 31P magnetic resonance spectroscopy (MRS) parameters. Hypoxic fractions were measured by excision assays in intramuscular and subcutaneous RIF-1 tumors, large and small SCCVII tumors, and KHT tumors. The hypoxic fractions ranged from 3% (small SCCVII) to 21% (KHT). Measurements of 31P MRS parameters (indices of tumor metabolism) were made under the same conditions used in the hypoxic fraction assays. The PCR/Pi ratios in the five tumor models were strongly correlated (r2 = 0.98, tau = 1.00, P < 0.02) with their hypoxic fractions, and the NTP/Pi ratio was weakly correlated (r2 = 0.67, tau = 0.60, P < 0.17) with hypoxic fraction. Smaller values of the PCr/Pi and NTP/Pi ratios were found in the tumors with larger hypoxic fractions. These experiments indicate that 31P MRS may provide a relative measurement of hypoxic fractions across different tumor lines that can be applied non-invasively to individual tumors.

Similar potency of the enantiomers of huperzine A in inhibition of [(3)H]dizocilpine (MK-801) binding in rat cerebral cortex.

The inhibition of huperzine A, a potential therapeutic agent to treat Alzheimer's disease, on rat cortical acetylcholinesterase was found to be highly stereospecific. In the present study the effect of the enantiomers of huperzine A on [(3)H]dizocilpine (MK-801) binding to synaptic membrane of rat cerebral cortex was compared. The natural (-)-huperzine A and the synthetic (+)-huperzine A inhibited the specific binding of [(3)H]MK-801 with a similar potency. The IC(50) values were 65+/-7 and 82+/-12 microM (n=5 for each enantiomer, P=0.248), respectively. The result indicates that huperzine A inhibits N-methyl-D-aspartate (NMDA) receptor in rat cerebral cortex without stereoselectivity.

Dynamic measurements of hexafluoromisonidazole (CCI-103F) retention in mouse tumours by 1H/19F magnetic resonance spectroscopy.

Selective retention of hexafluoromisonidazole, CCI-103F, in RIF-1 and SCCVII tumours of C3H/Km mice has been measured by 1H/19F magnetic resonance spectroscopy (MRS) on a Bruker AM-400 multinuclear spectrometer. CCI-103F concentrations in tumours and in normal tissues were measured using an MRS technique in which the water component in the tissues serves as an internal concentration reference. The biodistribution and elimination half-life of the drug in the tissues after i.p. injections were determined. The plasma half-life of the drug (41 min) was measured by high-pressure liquid chromatography. The two tumour lines and liver have longer retention times with half-lives of 47, 129 and 81 min, respectively, while normal tissues, muscle and brain have little retention of CCI-103F and clear the drug very quickly. Dynamic measurements of CCI-103F retention in tumours by MRS may provide a non-invasive probe for assessing tumour hypoxia.

Posterior mediastinal paraganglioma with bilateral adrenal pheochromocytoma.

Paraganglioma and adrenal pheochromocytoma are tumors of common origin arising from chromaffin cells. However, it is extremely rare to find mediastinal paraganglioma simultaneously with bilateral adrenal pheochromocytoma. We report a 53-year-old man who was diagnosed with posterior mediastinal paraganglioma and bilateral adrenal pheochromocytoma and who underwent successful excision of the posterior mediastinal mass and bilateral total adrenalectomy.

Predictors of functional outcome after emergency carotid artery stenting and intra-arterial thrombolysis for treatment of acute stroke associated with obstruction of the proximal internal carotid artery and tandem downstream occlusion.

BACKGROUND AND PURPOSE: Patients who develop severe stroke symptoms due to acute internal carotid artery occlusion eventually in combination with a thromboembolic obstruction of the middle cerebral artery incur a major risk of developing extensive MCA infarction with a poor outcome. The purpose of this study was to evaluate the outcome for patients with tandem occlusions in the MCA and/or distal ICA, retrospectively, who had undergone stent implantation in the proximal segment of the ICA in addition to intra-arterial thrombolysis. MATERIALS AND METHODS: Thirty-five patients with tandem occlusions of the MCA and/or distal ICA and acute occlusion of the proximal ICA underwent stent implantation for the proximal ICA occlusion and IAT for the tandem occlusion. Clinical outcome measures were assessed on admission and at discharge by using the National Institutes of Health Stroke Scale as well as 3 months after treatment by using the modified Rankin Scale. RESULTS: The median NIHSS score on admission was 12 (range, 6-22). All patients had patent flow into the M1 and ICA after carotid artery stent placement and IAT. After the procedure, 19 patients (54.3%) were TICI grade III; 7 (20.0%), TICI grade IIb; and 9 (25.7%), TICI grade IIa. Symptomatic intracerebral hemorrhage occurred in 1 patient (2.9%). The overall mortality rate was 11.4% (4/35). At 3-month follow-up, the median NIHSS score was 4 (range, 1-17). NIHSS score at admission and TICI grade were all found to be independently associated with an unfavorable outcome at 3 months. CONCLUSIONS: Initial stroke severity, degree of successful revascularization, and the side of ischemia were found to independently predict the functional outcome at 3 months after treatment.

Granulocytic sarcoma manifested as a parametrial mass mimicking a haemorrhagic abscess: a case report with CT and MR findings.

Granulocytic sarcoma is a neoplasm arising from myeloid precursor cells and frequently accompanies leukaemia and myeloproliferative disorders. Granulocytic sarcoma can arise anywhere, and it frequently involves bones, perineural tissues and lymph nodes. However, granulocytic sarcoma in the female genital organs is uncommon, and it is extremely rare that it presents as an adnexal or parametrial mass. We report here the CT and MR findings in a case of granulocytic sarcoma that manifested as a uterine cervical and parametrial mass mimicking a haemorrhagic abscess in a 50-year-old woman with chronic myelogenous leukaemia.

Prediction of tumor radiosensitivity by hexafluoromisonidazole retention monitored by [1H]/[19F] magnetic resonance spectroscopy.

Hexafluoromisonidazole (CCI-103F) is a hypoxic cell label that can be measured by in vivo [1H]/[19F] magnetic resonance spectroscopy (MRS). The retention of CCI-103F in SCCVII tumors was found to be correlated with tumor size, with larger tumors retaining more label. Such a correlation was anticipated, as larger SCCVII tumors have larger hypoxic fractions. A significant but less strong correlation was also found between CCI-103F retention and the fraction of tumor cells surviving after 10 Gy irradiation, with tumors that retained larger amounts of CCI-103F having higher surviving fractions. These results indicated that non-invasive MRS measurement of CCI-103F retention could predict radiosensitivity in SCCVII tumors. The lack of a strong correlation between CCI-103F retention and radiosensitivity, however, indicated that hypoxic fraction was not the only factor influencing radiosensitivity in these tumors.

Effects of local irradiation on spin-lattice relaxation time of phosphate metabolites in mouse tumors monitored by 31P magnetic resonance spectroscopy.

An inversion-recovery pulse sequence and solenoidal surface coil were employed to determine the spin-lattice relaxation time (T1) in murine tumors (RIF-1 and SCCVII). Reduction in T1s of inorganic phosphate (Pi) and nucleotide triphosphates (NTP) has been observed in irradiated tumors compared with unirradiated tumors. The reduction is accompanied by tumor regression and improved tumor energetic status in both tumor lines. The results of the T1 measurement of phosphate metabolites in tumors suggest that partial saturation effects may exist in spectra and could effect quantitative analysis when short repetition time is used. The reduction in T1s of Pi and NTP may be attributed to the active biochemical exchange and better tumor oxygenation in irradiated tumors.

X-ray structures of Torpedo californica acetylcholinesterase complexed with (+)-huperzine A and (-)-huperzine B: structural evidence for an active site rearrangement.

Kinetic and structural data are presented on the interaction with Torpedo californica acetylcholinesterase (TcAChE) of (+)-huperzine A, a synthetic enantiomer of the anti-Alzheimer drug, (-)-huperzine A, and of its natural homologue (-)-huperzine B. (+)-Huperzine A and (-)-huperzine B bind to the enzyme with dissociation constants of 4.30 and 0.33 microM, respectively, compared to 0.18 microM for (-)-huperzine A. The X-ray structures of the complexes of (+)-huperzine A and (-)-huperzine B with TcAChE were determined to 2.1 and 2.35 A resolution, respectively, and compared to the previously determined structure of the (-)-huperzine A complex. All three interact with the "anionic" subsite of the active site, primarily through pi-pi stacking and through van der Waals or C-H.pi interactions with Trp84 and Phe330. Since their alpha-pyridone moieties are responsible for their key interactions with the active site via hydrogen bonding, and possibly via C-H.pi interactions, all three maintain similar positions and orientations with respect to it. The carbonyl oxygens of all three appear to repel the carbonyl oxygen of Gly117, thus causing the peptide bond between Gly117 and Gly118 to undergo a peptide flip. As a consequence, the position of the main chain nitrogen of Gly118 in the "oxyanion" hole in the native enzyme becomes occupied by the carbonyl of Gly117. Furthermore, the flipped conformation is stabilized by hydrogen bonding of Gly117O to Gly119N and Ala201N, the other two functional elements of the three-pronged "oxyanion hole" characteristic of cholinesterases. All three inhibitors thus would be expected to abolish hydrolysis of all ester substrates, whether charged or neutral.