Objective:To investigate and compare the regulatory effects of umbilical cord mesenchymal stem cells (MSC) and their conditioned medium (MSC-CM) on gut microbiota of septic mice.Methods:Twenty-eight six-to-eight-week-old female C57BL/6J mice were randomly divided into sham operation group (Sham group), sepsis model group (CLP group), sepsis+MSC treatment group (CLP+MSC group) and sepsis+MSC-CM treatment group (CLP+MSC-CM group), with seven mice in each group. The septic mouse model was established by cecal ligation and puncture (CLP). In Sham group, CLP were not performed, and other operations were the same as CLP group. Mice in the CLP+MSC group and CLP+MSC-CM group received 0.2 mL 1×10 6 MSC or 0.2 mL concentrated MSC-CM via intraperitoneal injection 6 hours after CLP, respectively. Sham group and CLP group were given 0.2 mL sterile phosphate buffer saline (PBS) via intraperitoneal injection. Histopathological changes were evaluated by hematoxylin-eosin (HE) staining and colon length. Levels of inflammatory factors in serum were detected by enzyme-linked immunosorbent assay (ELISA). Phenotype of peritoneal macrophages was analyzed by flow cytometry, and the gut microbiota was analyzed via 16S rRNA sequencing. Results:Compared with Sham group, significant inflammatory injury in lung and colon was observed, and shorter colon was detected in CLP group (cm: 6.00±0.26 vs. 7.11±0.09), the level of inflammatory cytokine interleukin-1β (IL-1β) in serum was significantly increased (ng/L: 432.70±17.68 vs. 353.70±17.01), the proportion of F4/80 + peritoneal macrophages was increased [(68.25±3.41)% vs. (50.84±4.98)%], while the ratio of F4/80 +CD206 + anti-inflammatory peritoneal macrophages was decreased [(45.25±6.75)% vs. (66.66±3.36)%]. The α diversity sobs index of gut microbiota was downregulated significantly (118.50±23.25 vs. 255.70±6.87), the structure of species composition was altered, and the relative abundance of functional gut microbiota related to transcription, secondary metabolites biosynthesis, transport and catabolism, carbohydrate transport and metabolism, and signal transduction were decreased significantly in CLP group (all P < 0.05). Compared with CLP group, upon MSC or MSC-CM treatment, the pathological injury in lung and colon was alleviated to varying extent, the length of colon was increased (cm: 6.53±0.27, 6.87±0.18 vs. 6.00±0.26), the level of IL-1β in serum was downregulated (ng/L: 382.10±16.93, 343.20±23.61 vs. 432.70±17.68), the ratio of F4/80 + peritoneal macrophages was decreased [(47.65±3.93)%, (48.68±2.51)% vs. (68.25±3.41)%], the ratio of F4/80 +CD206 + anti-inflammatory peritoneal macrophages was increased [(52.73±5.02)%, (66.38±4.73)% vs. (45.25±6.75)%], and the α diversity sobs index of gut microbiota was increased (182.50±16.35, 214.00±31.18 vs. 118.50±23.25), and the effects of MSC-CM were more significant (all P < 0.05). At the same time, species composition of gut microbiota was rebuilt, and a tendency of increase in relative abundance of functional gut microbiota was observed upon MSC and MSC-CM treatment. Conclusion:Both MSC and MSC-CM could alleviate inflammatory injury in tissues, and showed regulatory effects on gut microbiota in septic mouse model, moreover, MSC-CM exhibited superior advantages over MSC.

Objective:To study the crosstalk between the activating transcription factor 6 (ATF6) and inositol-requiring enzyme 1 (IRE1) - X-box binding protein 1 (XBP1) pathway in oxygen-glucose deprivation/reoxygenation (OGD/R)-injured mouse hippocampal neuronal cell line HT22.Methods:The OGD/R-injured HT22 cell model was used to observe the changes of the indicators of endoplasmic reticulum stress (ERS), cell viability, and apoptosis at different OGD/R time points (0, 3, 6, 12, and 24 hours). HT22 cells in the logarithmic growth phase were randomized into blank control group, control+ATF6 activator (AA147) group, control+IRE1 inhibitor (4μ8c) group, OGD/R model group, OGD/R+AA147 group and OGD/R+4μ8c group (10 μmol/L AA147 or 16 μmol/L 4μ8c was given during the whole process in the AA147 group and 4μ8c group). Western blotting was used to detect the expression of ERS-related proteins [glucose-regulated protein 78 (GRP78), phosphorylated-inositol-requiring enzyme 1 (p-IRE1), and phosphorylated-eukaryotic translation initiation factor-2α (p-eIF2α)], and apoptosis-related proteins (Bcl-2, Bax, caspase-3, and cleaved caspase-3). The mRNA of ERS-related genes, and ATF6 [homocysteine-inducible, endoplasmic reticulum stress-inducible, ubiquitin-like domain member 1 (Herpud1), protein disulfide isomerase associated 4 (Pdia4) and Sel-1 suppressor of lin-12-like (Sel1L)] and spliced XBP1 [XBP1s, include DnaJ heat shock protein family member B9 (Erdj4), Sec24 related gene family, member D (Sec24d) and signal sequence receptor, gamma (Ssr3)] induced transcriptional response-related genes were measured by real-time quantitative polymerase chain reaction (RT-qPCR). A cell counting kit-8 (CCK-8) assay was used to detect the viability of HT22 cells. Immunofluorescence was utilized to test the expression of cleaved caspase-3.Results:Compared with the blank control group, the expression of ERS-related proteins p-IRE1 and p-eIF2α were significantly increased at 12 hours and 3 hours following OGD/R, respectively (p-IRE1/β-actin: 2.09±0.10 vs. 1.00±0.00, p-eIF2α/β-actin: 1.39±0.11 vs. 1.00±0.00, both P < 0.01). The mRNA expressions of ERS-related genes [ATF6, XBP1s, unspliced XBP1 (XBP1u), activating transcription factor 4 (ATF4), CCAAT/EBP homologous protein (CHOP)] were also upregulated in different OGD/R timepoint in HT22 cells, which indicated ERS was activated in OGD/R-stimulated HT22 cells. Compared with the OGD/R model group, the expression of protein p-IRE1 was not changed, but the mRNA of XBP1s and XBP1u were obviously downregulated in the OGD/R+AA147 group [XBP1s (2 -ΔΔCt): 0.76 (0.71, 0.92) vs. 1.13 (1.03, 1.29), XBP1u (2 -ΔΔCt): 0.29±0.05 vs. 0.52±0.04, both P < 0.01], whereas the expressions of XBP1s-induced transcriptional response downstream genes did not change significantly. Compared with the OGD/R model group, the protein of short-form ATF6 (sATF6) and GRP78 were not changed after administration of 4μ8c, neither was the mRNA expression of ATF6-induced transcriptional response-related genes. These results showed that the mRNA expression of XBP1s and XBP1u were inhibited by AA147-induced activation of ATF6, but no crosstalk was observed between the transcriptional response induced by ATF6 and XBP1s. Compared with the blank control group, the cell viability decreased significantly at OGD/R 3 hours [(44.64±5.12) % vs. (99.13±5.76) %, P < 0.01], the ratios of apoptosis-related proteins Bax/Bcl-2 and cleaved caspase-3/caspase-3 were significantly increased at OGD/R 3 hours and OGD 0 hour, respectively (Bax/Bcl-2: 6.15±1.65 vs. 1.00±0.00, cleaved caspase-3/caspase-3: 17.48±2.75 vs. 1.00±0.00, both P < 0.01), which indicated that apoptosis was activated in OGD/R-treated HT22 cells. Compared with the OGD/R model group, the cell viability decreased significantly [(36.52±17.78)% vs. (69.90±9.43)%, P < 0.01], and the ratios of Bax/Bcl-2 and cleaved caspase-3/caspase-3 were significantly upregulated in the OGD/R+AA147 group in HT22 cells (Bax/Bcl-2: 2.06±0.31 vs. 1.10±0.25, cleaved caspase-3/caspase-3: 3.35±0.59 vs. 0.55±0.09, both P < 0.01). Conclusion:Under our experimental conditions, no obvious crosstalk between the transcriptional response induced by ATF6 and XBP1s was observed, while ATF6 activation induced by AA147 suppressed mRNA expression of XBP1s and XBP1u and promoted cell death in OGD/R-treated HT22 cells.

The present study developed an ultra-fast liquid chromatography coupled with triple quadrupole-linear ion trap composite mass spectrometry(UHPLC-QTRAP-MS) to simultaneously determine the content of potential active components in Scutellariae Barbatae Herba and also to provide a reference approach for screening out the differential quality control components among different batches of Scutellariae Barbatae Herba. Chromatographic separations were conducted on a Thermo Acclaim~(TM) RSLC 120 C_(18) column(3.0 mm×100 mm, 2.2 μm) in a gradient program. The mobile phase consisted of 0.1% aqueous formic acid and acetonitrile, and the column temperature was maintained at 40 ℃. The flow rate was 0.4 mL·min~(-1) and the injection volume was 2 μL. The targeted compounds were monitored in the multiple reaction monitoring(MRM) mode. The acquired data were processed by hierarchical cluster analysis(HCA) and partial least square discriminant analysis(PLS-DA). Sixteen compounds all showed good linear relationship within the corresponding linear ranges and the R~2 values were all higher than 0.993 2. The RSDs of precision, repeatability, and stability were less than or equal to 3.7%. Mean recovery rates were in the range of 95.67% and 104.8% with RSDs≤3.2%. According to HCA and PLS-DA, all samples were clustered into four categories. Scutellarin, acteoside, scutellarein, and scutebarbatine X(VIP>1) were considered as differential chemical markers in the four categories. In conclusion, the developed method can be used for the simulta-neous determination of the multiple components and quality control of Scutellariae Barbatae Herba.
Chemometrics ; Chromatography, High Pressure Liquid/methods* ; Chromatography, Liquid ; Scutellaria ; Tandem Mass Spectrometry/methods*

Ischemia/reperfusion (I/R) caused by cardiac arrest (CA) and subsequent cardiopulmonary resuscitation (CPR) was the primary cause of post-cardiac arrest syndrome (PCAS), including post-cardiac arrest myocardial dysfunction and post-cardiac arrest brain injury. Disturbance of endoplasmic reticulum proteostasis, so-called endoplasmic reticulum stress (ERS) was one of the pathological changes induced by I/R injury. The unfolded protein response (UPR) was an adaptive response triggered by ERS in cells. Modulating the UPR arms to alleviate ERS to promote cell survival was promising for attenuating I/R injury. Activating the activating transcription factor6 (ATF6) signaling pathway, one of the arms of the UPR, confers protection against I/R injury in multiple tissues by restoring endoplasmic reticulum proteostasis and reducing oxygen free radicals. This article reviewed the structural characteristics and biological function of ATF6 and focused on its essential role in cardiac and cerebral I/R injury as well as potential therapeutic targets, hoping to provide new ideas for the effective treatment of PCAS.

BACKGROUND: Various prediction tools have been developed to predict biochemical recurrence (BCR) after radical prostatectomy (RP); however, few of the previous prediction tools used serum prostate-specific antigen (PSA) nadir after RP and maximum tumor diameter (MTD) at the same time. In this study, a nomogram incorporating MTD and PSA nadir was developed to predict BCR-free survival (BCRFS). METHODS: A total of 337 patients who underwent RP between January 2010 and March 2017 were retrospectively enrolled in this study. The maximum diameter of the index lesion was measured on magnetic resonance imaging (MRI). Cox regression analysis was performed to evaluate independent predictors of BCR. A nomogram was subsequently developed for the prediction of BCRFS at 3 and 5 years after RP. Time-dependent receiver operating characteristic (ROC) curve and decision curve analyses were performed to identify the advantage of the new nomogram in comparison with the cancer of the prostate risk assessment post-surgical (CAPRA-S) score. RESULTS: A novel nomogram was developed to predict BCR by including PSA nadir, MTD, Gleason score, surgical margin (SM), and seminal vesicle invasion (SVI), considering these variables were significantly associated with BCR in both univariate and multivariate analyses (P < 0.05). In addition, a basic model including Gleason score, SM, and SVI was developed and used as a control to assess the incremental predictive power of the new model. The concordance index of our model was slightly higher than CAPRA-S model (0.76 vs. 0.70, P = 0.02) and it was significantly higher than that of the basic model (0.76 vs. 0.66, P = 0.001). Time-dependent ROC curve and decision curve analyses also demonstrated the advantages of the new nomogram. CONCLUSIONS PSA nadir after RP and MTD based on MRI before surgery are independent predictors of BCR. By incorporating PSA nadir and MTD into the conventional predictive model, our newly developed nomogram significantly improved the accuracy in predicting BCRFS after RP.
Humans ; Male ; Neoplasm Grading ; Neoplasm Recurrence, Local/surgery* ; Nomograms ; Prognosis ; Prostate-Specific Antigen ; Prostatectomy ; Prostatic Neoplasms/surgery* ; Retrospective Studies ; Seminal Vesicles

Analgesia ; methods ; Betacoronavirus ; Conscious Sedation ; methods ; Consensus ; Coronavirus Infections ; complications ; Humans ; Pandemics ; Pneumonia, Viral ; complications

Objective:To compare the severity of brain injury between asphyxia and electrical stimulation induced cardiac arrest in rats.Methods:Forty-two healthy male Sprague-Dawley (SD) rats were randomized into sham group ( n = 6), asphyxia group ( n = 18) and electrical stimulation group ( n = 18). Rats in each group were given invasive mechanical ventilation and femoral blood vessels catheterization for monitoring blood pressure and fluid infusion. In the asphyxia group, the tracheal tube was clamped to induce cardiac arrest, and in the electrical stimulation group, the esophageal electrical stimulation was used to induce cardiac arrest, and cardiopulmonary resuscitation (CPR) was performed 4 minutes after cardiac arrest. In the sham group, only tracheal intubation and femoral artery intubation were performed after anesthesia, but cardiac arrest was not induced. Animals were allowed to survive until 72 hours after resuscitation, and survival analysis was performed using Kaplan-Meier curves. At 24 hours and 72 hours after resuscitation, the neurological deficit score (NDS) was measured. The vena cava blood was collected, and the brain injury associated serum biomarkers, neuron-specific enolase (NSE) and S100B, were detected by enzyme-linked immunosorbent assay (ELISA). The brain tissues were then harvested to perform hematoxylin-eosin (HE) staining for observing pathological changes in the hippocampal CA1 area with light microscopy. Results:Cardiac arrest was successfully induced in both the asphyxia group and the electrical stimulation group, 94.4% (17/18) and 88.9% (16/18) animals were resuscitated successfully in the two groups respectively. Kaplan-Meier curves analysis showed that 72-hour cumulative survival rate was similar in the asphyxia group and the electrical stimulation group (Log-Rank test: χ2 = 0.040, P = 0.841). Both asphyxia group and electrical stimulation group had higher NDS score than sham group at 24 hours after resuscitation (37.50±4.26, 32.17±4.02 vs. 8.33±2.33, both P < 0.01). NDS score showed a downwards trend at 72 hours after resuscitation in both model groups, and the decline was more significant in the electrical stimulation group, which was significantly different as compared with asphyxia group (14.00±2.89 vs. 26.33±4.84, P < 0.05). ELISA results showed that the levels of serum NSE at 24 hours after resuscitation in the asphyxia and electrical stimulation groups were significantly higher than those in the sham group (μg/L: 1.02±0.07, 1.02±0.02 vs. 0.87±0.02, both P < 0.05). NSE kept increasing at 72 hours after resuscitation in the asphyxia group, which showed significant difference as compared with sham group (μg/L: 1.03±0.05 vs. 0.87±0.02, P < 0.01). But it had almost recovered to the normal level in the electrical stimulation group without significant difference as compared with sham group (μg/L: 0.96±0.04 vs. 0.87±0.02, P > 0.05). There was no significant difference in S100B level at different time points after resuscitation among three groups. It was displayed under light microscope that there was no significant neuronal damage in the hippocampal CA1 area in the two model groups at 24 hours after resuscitation as compared with the sham group. At 72 hours, there were certain damages in the hippocampal CA1 area in both model groups, which were more obvious in the asphyxia group. Conclusions:Both cardiac arrest models induced by asphyxia and electrical stimulation show a certain degree of brain injuries after resuscitation. Brain injuries are more severe in asphyxia-induced cardiac arrest compared with trans-esophageal electrical stimulation method.

Objective:To compare the therapeutic effects and safety of dexmedetomidine and midazolam on patients with severe coronavirus disease 2019 (COVID-19) who received non-invasive ventilation.Methods:Patients with COVID-19 who needed non-invasive ventilation in one critical care medicine ward of Wuhan Jinyintan Hospital during the team support period from the department of critical care medicine of Renmin Hospital of Wuhan University from January 23rd to February 15th in 2020 were investigated retrospectively. Ramsay score, mean arterial pressure (MAP), heart rate (HR), respiratory rate (RR), arterial oxygen partial pressure (PaO 2) before sedation and at 1, 12, 24 hours after sedation, sleep time were collected, and the side effects such as excessive sedation, fall of tongue, abdominal distension, aspiration, bradycardia, escalation to invasive mechanical ventilation during 24 hours were also collected. According to different sedative drugs, patients were divided into the control group (without sedative drugs), dexmedetomidine group and midazolam group. The changes of indicators among the three groups were compared. Results:Fourteen patients were injected with dexmedetomidine (loading dose of 1 μg/kg for 10 minutes, maintained at 0.2-0.7 μg·kg -1·h -1); 9 patients were injected with midazolam (loading dose of 0.05 mg/kg for 2 minutes, maintained at 0.02-0.10 mg·kg -1·h -1); 12 patients didn't use sedative drugs due to limitations of previous hospital or patients' rejection. In dexmedetomidine group and midazolam group, the Ramsay score was maintained at 2-3 points after sedation, which were higher than those of control group at different time points after sedation, and there was no significant difference between dexmedetomidine group and midazolam group. MAP of dexmedetomidine group and midazolam group decreased gradually after sedation. MAP after 1-hour sedation was significantly lower than that before sedation, and MAP after 24 hours sedation was significantly lower than that in the control group [mmHg (1 mmHg = 0.133 kPa): 109.7±11.5, 107.1±12.3 vs. 121.1±13.3, both P < 0.05]. HR decreased gradually after sedation treatment, which was significantly lower after 12 hours of sedation than that before sedation, and HR in dexmedetomidine group was significantly lower than that in control group after 12 hours of sedation (bpm: 84.0±13.9 vs. 92.8±15.4 at 12 hours; 81.0±16.7 vs 92.6±12.7 at 24 hours, both P < 0.05). PaO 2 increased and RR decreased in all three groups after ventilation. PaO 2 in dexmedetomidine group and midazolam group were significantly higher than that in the control group after 12 hours of sedation [cmH 2O (1 cmH 2O = 0.098 kPa): 79.0±6.5, 79.0±8.9 vs. 70.0±7.8, both P < 0.05]; the decreases of RR in dexmedetomidine group and midazolam group were significant than that in control group after 1 hour of sedation (bpm: 34.0±3.9, 33.8±4.6 vs. 39.0±3.6, both P < 0.05). There were no differences of MAP, HR, PaO 2 and RR between dexmedetomidine group and midazolam group at different time points. The sleep duration in dexmedetomidine group and midazolam group were significantly longer than that in the control group (hours: 4.9±1.9, 5.8±2.4 vs. 3.0±1.8, both P < 0.05), but there was no difference between dexmedetomidine group and midazolam group ( P > 0.05). Adverse events occurred in all three groups. In midazolam group, there were 2 cases of excessive sedation with fall of tongue and abdominal distension, including 1 case of aspiration, 1 case receiving intubation due to refractory hypoxemia and 1 case due to unconsciousness. In dexmedetomidine group, there were 2 cases of bradycardia, 1 case of intubation due to refractory hypoxemia. In control group, 4 cases underwent intubation due to refractory hypoxemia. Conclusions:Non-invasive mechanical ventilation is an important respiratory support technology for patients with severe COVID-19. Appropriate sedation can increase the efficiency of non-invasive mechanical ventilation. Dexmedetomidine is more effective and safer than midazolam in these patients, but attention should be paid to HR and blood pressure monitoring.

COVID-19/therapy* ; Consensus ; Critical Illness ; Humans ; SARS-CoV-2

Betacoronavirus ; COVID-19 ; Coronavirus ; Coronavirus Infections/therapy* ; Humans ; Pandemics ; Pneumonia, Viral/therapy* ; SARS-CoV-2