Objective:To evaluate the role of spinal cord microglia in chronic pain after chronic heart failure (CHF) in mice.Methods:Eighteen SPF healthy male C57BL/6 mice, aged 8-12 weeks, weighing 20-25 g, were divided into 3 groups ( n=6 each) using a random number table method: sham operation group (Sham group), CHF group, and CHF+ microglia inhibitor PLX3397 group (CHF+ PLX group). The model of chronic heart failure was prepared by ligating the anterior descending branch of the left coronary artery in anesthetized mice. Mice were continuously fed a PLX3397-containing diet from 7 days before preparing the model to 28 days after preparing the model in CHF + PLX group. The cardiac function was evaluated using echocardiography at 28 days after surgery. The mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) were measured on 1 day before developing the model (T 0) and 1, 3, 7, 14 and 28 days after developing the model (T 1-5). The mice were sacrificed under deep anesthesia after the pain threshold was measured at T 5, and the spinal cord tissues of the lumbar segment (T 2-6 and L 4-6) were removed for determination of the expression of microglial marker Iba1 in the spinal dorsal horn (by immunofluorescence method). Results:Compared with Sham group, the left ventricular ejection fraction and left ventricular short axis shortening rate were significantly decreased, and the left ventricular internal diameter at end-diastole and the left ventricular internal diameter at end-systole were increased, the TWL was shortened and MWT was decreased at T 4-5, and the expression of Iba1 in the spinal dorsal horn was up-regulated in CHF group ( P<0.05). Compared with CHF group, the left ventricular ejection fraction and left ventricular short axis shortening rate were significantly increased, and the left ventricular internal diameter at end-diastole and the left ventricular internal diameter at end-systole were decreased, the TWL was prolong and MWT was increased at T 2-5, and the expression of Iba1 in the spinal dorsal horn was down-regulated in CHF+ PLX group ( P<0.05). Conclusions:The chronic pain after CHF may be related to microglial activation in the spinal cord of mice.

With the number of cases of coronavirus disease-2019 (COVID-19) increasing rapidly, the World Health Organization (WHO) has recommended that patients with mild or moderate symptoms could be released from quarantine without nucleic acid retesting, and self-isolate in the community. This may pose a potential virus transmission risk. We aimed to develop a nomogram to predict the duration of viral shedding for individual COVID-19 patients. This retrospective multicentric study enrolled 135 patients as a training cohort and 102 patients as a validation cohort. Significant factors associated with the duration of viral shedding were identified by multivariate Cox modeling in the training cohort and combined to develop a nomogram to predict the probability of viral shedding at 9, 13, 17, and 21 d after admission. The nomogram was validated in the validation cohort and evaluated by concordance index (C-index), area under the curve (AUC), and calibration curve. A higher absolute lymphocyte count (
Aged ; Aged, 80 and over ; Antibodies, Viral/blood* ; Area Under Curve ; COVID-19/virology* ; Female ; Humans ; Lymphocyte Count ; Male ; Middle Aged ; Nomograms ; Proportional Hazards Models ; Retrospective Studies ; Viral Load ; Virus Shedding

Objective:To investigate the value of programmed death-1（PD-1） expression on the T lymphocytes for the prognosis of septic patients.Methods:From September 2017 to May 2019, septic patients were included in Department of Intensive Care Unit at 6 hospitals. The PD-1 expression on T cells were measured by flow cytometry. Logistic regression was conducted to analyze independent risk factors related to death within 28 days，and receiver operating characteristic curve(ROC) was conducted to evaluate the prognostic value of PD-1 expression on T cells in septic patients.Results:A total of 64 septic patients were enrolled to this study，including 32 survivors and 32 deaths. The PD-1 expression on T cells in the death group was significantly higher than that in the surviving group ( P<0.05). Correlation analysis showed that the percentages of PD-1 +/CD3 +T cells and PD-1 +/CD8 +T cells were positively correlated with procalciton in ( r=0.313, P =0.015； r=0.375, P=0.003), logistic regression analysis showed that the percentages of PD-1 +/CD3 +，PD-1 +/CD4 +，PD-1 +/CD8 +T cells were independent risk factors for the death of sepsis patients. The percentage of PD-1 +/CD3 +T cell was 3.63%, with AUC 0.842, sensitivity to predict the mortality 96.43% and specificity 59.38%, ( P<0.000 1). The percentage of PD-1 +/CD4 +T cell was 4.65%, with AUC 0.847, sensitivity 96.43%, specificity 62.50%，( P<0.000 1). The percentage of PD-1 +/CD8 +T cell was 3.91%, with AUC 0.771, sensitivity 64.29%, specificity 81.25%，( P=0.000 3). Conclusions:The T cell PD-1 expression is an independent risk factor to predict the 28-day mortality in septic patients. Combining the proportions of PD-1 +/CD3 +, PD-1 +/CD4 +and PD-1 +/CD8 +T cells may further enhance the predictive value for death.

Objective To investigate the effect of levosimendan on cardiac function and prognosis in elderly patients with septic myocardial contractility impairment.Methods A prospective,randomized,controlled study was conducted.The elderly patients with septic myocardial contractility impairment who were admitted to Intensive Care Unit in Zhejiang Hospital were consecutively enrolled from January 2017 to September 2017.The key inclusive criterion was left ventricular ejection fraction (LVEF) ≤ 50％ after fluid resuscitation.A total of 30 patients were randomly assigned to levosimendan group (n=15) and dobutamine group (n=15).Based onconventional treatment,intravenous dobutamine (5 μg per kilogram of body weight per minute) or levosimendan (0.2 μg per kilogram of body weight per minute)were continuously administrated for 24 hours in two groups.At 0 h,24 h,48 h,72 h after injection,the following parameters or values were recorded including serum lactic acid (Lac),and echocardiographic parameters such as LVEF,stroke volume (SV).The time of mechanical ventilation,length of stay in ICU and 28-day mortality were compared in two groups.Results Compared with dobutamine group,blood Lac at 24 h [(1.97±1.10)mmol/L vs.(2.73 ± 2.06) mmol/L,P=0.002] decreased significantlyin levosimendan group.LVEF and SV were significantly higher in levosimendan group at 24 h [LVEF:(47.93±5.01)％ vs.(45.60±5.47)％,P=0.004;SV:(47.73 ± 14.01) ml vs.(44.80±16.89) ml,P=0.035;respectively],48 h [LVEF:(51.07 ± 5.05)％ vs.(46.73 ± 6.34)％,P=0.004;SV:(49.87 ± 14.15) ml vs.(45.07± 16.94) ml,P=0.005;respectively] and 72 h [LVEF:(53.20±5.92)％ vs.(47.70±6.71)％,P=0.002;SV:(51.27±14.98) ml vs.(45.73±17.34) ml,P=0.010].The time of mechanical ventilation,length of stay in ICU and 28-day mortality were comparable between two groups (P＞0.05).Conclusions Levosimendan improves cardiac systolic function and tissue perfusion in elderly patients with septic myocardial contractility impairment.However,cardiac diastolic function,liver and kidney function are not further improved by levosimendan compare with dubutamine.Time of mechanical ventilation,length of stay in ICU and 28-day mortality in two groups are similar.

To analyze the correlation between transcutaneous oxygen pressure (PtcO2) and blood lactate in patients with septic shock. Fifty-sixpatients with septic shock were prospectively investigated. PtcO2 was monitored continuously for 6 hours, and arterial blood gas was measured at baseline (T0) and 6 hours (T6). Records of PtcO2, were analyzed for the correlation with lactate level and lactate clearance rate. PtcO2 valuesin the high lactate clearance group and the low one were compared.The lowest value of PtcO2 at T6 and duration of PtcO2<40 mmHg (1 mmHg=0.133 kPa) were both correlated with lactate level and lactate clearance rateat T6. The low predictive value of PtcO2 was 29 mmHg of lactate clearance under 20％with a sensitivity 85.2％and a specificity 65.5％. The low predictive value of PtcO2 in high lactate clearance group was significantly higher than that in low lactate clearance group, while the duration of PtcO2<40 mmHg was shorter than the latter. During 6 h continuous monitoring, patients with a significant low PtcO2 or prolonged duration of low PtcO2 have relatively high lactate or low lactate clearance after resuscitation.

The aim of this study is to establish a multiplex polymerase chain raction (PCR) to identify of four kinds of laboratory animal pathogens: Pasteurella multocida, Bordetella bronchiseptica, Mycoplasma pneumoniae and Klebsiella pneumoniae.Methods Specific primers were designed based on GenBank data.The multiplex PCR system was established through optimization of multiple PCR and detection of its specificity and sensitivity.This technique was used to test artificially infected samples and tracheal secretions of experimental animals (rat, mouse, guinea pig, rabbit, hamster), and comparing the detection results by this method and traditional detection test.Results Target bands of Pasteurella multocida (356 bp), Bordetella bronchiseptica (237 bp), Mycoplasma pneumoniae (266 bp), and Klebsiella pneumoniae (142 bp) were obtained, with a detection sensitivity of Klebsiella pneumoniae of 10 pg, and that of Pasteurella multocida, Bordetella bronchiseptica and Mycoplasma pneumoniae of 1 pg by this newly developed multiplex PCR assay.No target bands were observed from the non-specific pathogens of artificially infected samples.The tracheal secretions taken from 45 experimental animals (mice and rabbits) were tested with this new PCR assay, among which 15 cases of Klebsiella pneumonia and 9 cases of Pasteurella multocida were detected as positive, while all the results of traditional method and serological test were negative.Conclusions A simple, rapid, specific and highly sensitive multiplex PCR system has been successfully established.It is valuable for detection of Pasteurella multocida, Bordetella bronchiseptica, Mycoplasma pneumoniae, and Klebsiella pneumoniae in laboratory animals.

Objective To investigate the effect of intrathecal morphine preconditioning on the expression of nerve growth factor (NGF) in the dorsal root ganglia (DRG) in a rat model of myocardial ischemia-reperfusion (I/R).Methods Thirty healthy adult male Sprague-Dawley rats in which intrathecal catheters were successfully placed without complications,weighing 250-350 g,were randomly divided into 5 groups (n =6 each) using a random number table:sham operation group (S group),I/R group,intrathecal morphine preconditioning group (ITMP group),μ receptor antagonist CTOP + intrathecal morphine preconditioning group (CTOP + ITMP group),and CTOP control group (CTOP group).Myocardial ischemia was induced by 30 min of occlusion of the anterior descending branch of the left coronary artery followed by 120 min of reperfusion in all the groups except S group.Intrathecal morphine preconditioning was produced by 3 cycles of 5 min intrathecal injection of morphine 3 μg/kg (10 μl) at 5 min intervals within 30 min before ischemia in ITMP group.In CTOP+ITMP and CTOP groups,1 μg/μ1 CTOP 10 μl was injected intrathecally at 10 min before morphine preconditioning and 40 min before ischemia,respectively.At 120 min of reperfusion,the rats were sacrificed,and myocardial specimens were obtained for determination of myocardial infarct size,and DRGs were removed for determination of the expression of NGF by using immunohistochemistry and Western blot.Results Compared with S group,the myocardial infarct size was significantly increased,and the expression of NGF in DRGs was significantly up-regulated in I/R group (P＜0.05).Compared with I/R group,the myocardial infarct size was significantly decreased,and the expression of NGF in DRGs was significantly down-regulated in ITMP group (P＜ 0.05),and no significant change was found in the parameters mentioned above in CTOP group (P＞0.05).Compared with ITMP group,the myocardial infarct size was significantly increased,and the expression of NGF in DRGs was significantly up-regulated in CTOP+ITMP and CTOP groups (P＜0.05).Conclusion The mechanism by which intrathecal morphine preconditioning reduces myocardial I/R injury is related to activation of spinal μ receptors,inhibition of NGF expression in DRGs,and reduction of responses to noxious stimulation in the rats.

Objective To investigate the role of p38 mitogen-activated protein kinase (p38MAPK) signaling pathway in reduction of myocardial ischemia-reperfusion (I/R) injury by morphine preconditioning in the rats with chronic heart failure in vitro.Methods Adult male Sprague-Dawley rats,weighing 200-230 g,aged 6-7 weeks,in which doxorubicin 2 mg/kg was injected via the tail vein once a week for 6 consecutive weeks to induce chronic heart failure,were studied.At the end of 8th week,30 rats with chronic heart failure were randomly divided into 5 groups (n =6 each) using a random number table:sham operation group (group Sham),I/R group,morphine preconditioning group (group MPC),SB203580 (p38MAPK inhibitor) + morphine preconditioning group (group SBM),and SB203580 group (group SB).The hearts were quickly excised and passively perfused in a Langendorff apparatus and subjected to 30 min of occlusion of the anterior descending branch of the left coronary artery followed by 120 min of reperfusion to establish the model of myocardial I/R injury.After equilibration,the hearts were subjected to 3 cycles of 5 min perfusion with K-H solution containing morphine 1 μmol/L at 5-min intervals before ischemia in group MPC.In group SBM,the hearts were perfused with K-H solution containing SB203580 (5 μmol/L) for 45 min starting from l0 min before morphine preconditioning until 5 min of ischemia.In group SB,morphine preconditioning was not performed,and the hearts were only perfused with K-H solution containing SB203580 (5 μmol/L) starting from 40 min before ischemia until 5 min of ischemia.At 15 min of equilibration (baseline),5 and 10 min of reperfusion,the coronary effluent was collected to detect the activity of lactate dehydrogenase (LDH) using the chemical colorimetry.At 10 min of reperfusion,the expression of phosphor-p38MAPK (p-p38MAPK) in the myocardium was determined by Western blot in Sham,I/R and MPC groups.At 120 min of reperfusion,the area at risk (AAR),total areas of right and left ventricles (LV+RV),and infarct size (IS) were measured,and the IS/AAR ratio was calculated.Results Compared with group Sham,the LDH activity in coronary effluent during reperfusion and IS/AAR ratio were significantly increased in the other groups,and the expression of p-p38MAPK was significantly up-regulated in I/R and MPC groups (P＜0.05).Compared with group I/R,the LDH activity in coronary effluent during reperfusion was significantly decreased,the expression of p-p38MAPK was significantly up-regulated,and the IS and IS/AAR ratio were significantly decreased in group MPC (P＜0.05),and no significant change was found in the LDH activity in coronary effluent,IS and IS/AAR ratio in SBM and SB groups (P＞0.05).Compared with group MPC,the LDH activity in coronary effluent during reperfusion was significantly increased,and the IS and IS/AAR ratio were significantly increased in group SBM (P＜0.05).Conclusion The mechanism by which morphine preconditioning reduces myocardial I/R injury is related to activation of p38MAPK signaling pathway in the rats with chronic heart failure in vitro.

Objective To investigate the effect of intrathecal morphine preconditioning (ITMP) on the excitability of substantia gelatinosa (SG) neurons in the dorsal horn of the spinal cord in a rat model of myocardial ischemia-reperfusion (I/R).Methods Thirty-six adult male Sprague-Dawley rats,weighing 200-300 g,in which intrathecal catheters were successfully placed without complications,were randomly divided into 3 groups (n =12 each) using a random number table:sham operation group (group S),group I/R,and group ITMP.Myocardial I/R injury was produced by occlusion of the left anterior descending branch of the coronary artery for 30 min followed by 120 min reperfusion.In group ITMP,the rats received intrathecal morphine 3 μg/kg (10 μl) by three cycles of 5 min infusions interspersed with 5 min infusion-free periods starting from 30 min before ischemia,and the equal volume of normal saline was given instead of morphine in group I/R.At 10 min of reperfusion,6 rats randomly selected in each group were sacrificed,and the T2-6 segments of the spinal cords were acutely isolated to prepare spinal cord slices.The resting potential,threshold of action potential (APT),peak of action potential (APP) and action potential duration in SG neurons in the dorsal horn of spinal cord slices were determined using the whole-cell patch-clamp technique,and the number of action potentials evoked by currents of 40,60,80 and 100 pA was recorded.At 120 min of reperfusion,6 rats randomly selected in each group were sacrificed,and myocardial specimens were obtained for determination of myocardial infarct size (IS) and area at risk (AAR),and IS/AAR ratio was calculated.The expression of c-fos in the T2-5 dorsal horns of the spinal cords was detected by Western blot.Results Compared with group S,the IS/AAR ratio was significantly increased,the expression of c-fos was up-regulated,the number of action potentials in SG neurons in dorsal horns of spinal cord was increased,APT was decreased,and APP was increased in group I/R (P＜0.05).Compared with group I/R,the IS/AAR ratio was significantly decreased,the expression of c-fos was down-regulated,the number of action potentials in SG neurons in dorsal horns of spinal cord was decreased,APT was increased,and APP was decreased in group ITMP (P＜0.05).Conclusion The mechanism by which ITMP attenuates myocardial I/R injury is related to decrease in the excitability of SG neurons in the dorsal horn of the spinal cord and reduction of responses to nociceptive stimuli in rats.

Secretory diarrhea provides a major health challenge worldwide, which is one of the most important reasons for children morbidity and death. The activation of Cl- channels in intestinal epithelial cells resulting in the excessive fluid secretion in the intestine is the main reason of diarrhea caused by enterotoxins. In diarrhea caused by cholera and the other bacterial enterotoxins, cystic fibrosis transmembrane conductance regulator ( CFTR) is the main cAMP-control Cl- channel to promote the fluid secretion in epithelial cells. Therefore, CFTR inhibitors are the new choices for secretory diarrhea. CFTR inhibitors include thiazolidinone, glycine hydrazide and quinoxalinedione chemical classes, and some components from natural plants also exhibit CFTR inhibition activity, however, further studies should be done.