Between April 2012 and May 2014, 100 patients with AOPP admitted to the Affiliated Hospital of Taishan Medical University (Tai'an, China) were selected for the study. The diagnosis of AOPP was based on the following criteria: i) History of exposure to toxins; ii) typical clinical manifestations and symptoms (5); iii) symptom improvement following treatment with reactivating agent; and iv) reduction in activity of plasma cholinesterase. All patients had accidentally ingested organophosphorus pesticides (50–450 ml). According to the AOPP diagnosis and classification standards (13), 54 patients presented mild AOPP, 32 presented moderate AOPP and 14 presented severe AOPP. The basic clinical characteristics of all patients are listed in Table I. In addition, 100 healthy subjects, including 45 men and 55 women with an average age of 39.6±8.17 years (age range, 30–50 years), were included as the control group. The gender ratio and average age of the control group was not significantly different, as compared with the AOPP group. The present study was approved by the Ethics Committee of Taishan Medical University. Written informed consent was obtained from all patients or their families.

All AOPP patients received routine gastric lavage with water, oxygen intake, electrocardiogram monitoring and intravenous injection with 1–1.5 g pralidoxime chloride (Sihuan Pharmaceutical Holdings Group, Co., Ltd., Beijing, China) and 1–2 mg atropine (China Resources Double-Crane Pharmaceutical, Co., Ltd., Beijing, China). In cases of respiratory failure, the patients were subjected to intubation and ventilation, assisted by the S9 VPAP™ Auto double level automatic ventilator (ResMed, San Diego, CA, USA). In cases of asystole, the patients were treated with chest compression and cardiopulmonary resuscitation, in addition to water and electrolyte imbalance rectification, nutritional improvement and appropriate dehydration. Patients with severe symptoms underwent plasma purification therapy.

APACHE II scoring was conducted in order to measure the severity of AOPP in patients (14). APACHE II scores are determined from a patient's age and physiological indicators, including the alveolar-arterial oxygen gradient or partial pressure of oxygen in arterial blood, body temperature (rectal), mean arterial pressure, arterial pH, heart rate, respiratory rate, serum levels of sodium and potassium, levels of creatinine, the hematocrit, white blood cell count, and the Glasgow Coma Scale (14). The worst value for each indicator within 1 day after hospitalization was recorded into a electronic table produced by Microsoft Excel 2010 (Microsoft Corporation, Redmond, WA, USA). Patients with one missing/unrecorded indicator received an APACHE II score of zero, while those with more than two missing/unrecorded indicators were not scored. Factors associated with a high-risk of disease progression and mortality, including blood lactate level, acetylcholine esterase level, twitch disappearance time and an inability to raise the head, as determined by our hospital, were measured via routine blood and urine tests upon admission of the patients to the hospital. If these factors were exacerbated following active treatment, the patient was considered to have toxification progression. A higher APACHE II score corresponded to a more severe disease and indicated an elevated risk of mortality (14)

On days 1, 3 and 7, 5 ml plasma was obtained from fasting peripheral blood. The concentrations of plasma CRP and copeptin were measured using the CRP ELISA kit (DRG Instruments GmbH, Marburg, Germany) and the Copeptin ELISA kit (48T/96T; MLBIO, Shanghai, China), respectively, according to the manufacturer's protocol. The protein expression levels were quantified using the SpectraMax® M5 Microplate Reader (Molecular Devices, LLC, Sunnyvale, CA, USA) and analyzed using the SoftMax® Pro Data Acquisition and Analysis software, version 5.4.1 (Molecular Devices, LLC).

All the results were analyzed using SPSS software, version 19.0 (IBM SPSS, Armonk, NY, USA). The data are expressed as the mean ± standard deviation. Differences between groups were compared using the Student's t-test, and multivariate analysis was performed using logistic regression analysis. The Kaplan-Meier method was used to analyze the prognostic asymptomatic survival curves in relation to the CRP and copeptin levels in the patient plasma. Analysis of prognostic determination was performed using the area under the receiver operating characteristic (ROC) curve. P<0.05 was considered to indicate a statistically significant difference.

To measure the changes of CRP and copeptin contents in the plasma after treatment, ELISA was conducted. The plasma CRP and copeptin levels were increased with the increase of AOPP severity on days 1, 3 and 7 after AOPP, and were significantly higher compared with those in the control group (P<0.05). Patients with severe AOPP exhibited increasing plasma CRP and copeptin contents over time (P<0.01), while patients with mild and moderate AOPP exhibited reduced plasma CRP and copeptin contents over time. On day 7 after AOPP, plasma CRP and copeptin contents in all severity groups of AOPP patients remained significantly increased compared with the levels in the control group (P<0.05; Table II). These results suggest that the plasma levels of CRP and copeptin in patients with severe AOPP increased over time; however, this is did not occur in patients with moderate or mild AOPP.

To evaluate the effect of relative factors on the prognosis of AOPP, logistic regression analysis was used. The data showed that higher APACHE II scores, low blood lactate and acetylcholine esterase levels, an increased twitch disappearance time, a higher reactivating agent dose and an inability to raise the head were high risk factors (P<0.05; Table III). Therefore, the amelioration of these parameters may improve the prognosis of AOPP.

To test the effect of plasma CRP and copeptin on prognosis, Kaplan-Meier analysis was performed on the prognostic asymptomatic survival curves. According to the plasma CRP and copeptin levels at 1 week after AOPP, the number of patients with CRP levels that were higher and lower than the median value (11.87 mg/l) was 15 and 85, respectively. In addition, the number of patients with copeptin levels that were higher and lower than the median value (6.90 ng/ml) was 19 and 81, respectively. Kaplan-Meier analysis of prognostic asymptomatic curves showed that patients with plasma CRP and copeptin levels higher than the median values had a worse prognosis (P<0.05; Fig. 1). The results suggest that higher CRP and copeptin levels in the plasma indicate a poorer prognosis.

To evaluate the predictive role of CRP levels, copeptin levels and APACHE II scores on the survival of AOPP patients, ROC curves were plotted using SPSS software. The areas under the ROC curves were 0.89, 0.75 and 0.72 for CRP, copeptin and APACHE II scores, respectively (P<0.01). In addition, the optimum diagnostic cut-off points for CRP, copeptin and APACHE II scores were 10.2 mg/l, 6.3 ng/ml and 10.5, respectively. Notably, all the CRP levels, copeptin levels and APACHE II scores had a sensitivity and specificity of >90% (Fig. 2). The results indicate that CRP levels, copeptin levels and APACHE II scores may be used for the prediction of survival in AOPP patients.