[Study on the Protective effect and mechanism of Nicotinamide Riboside on lung injury in paraquat intoxicated mice].

Objective: To investigate the protective effect and mechanism of Nicotinamide Riboside (NR) on lung injury caused by Paraquat intoxicated mice. Methods: Eighty clean male BALB/C mice were selected and averagely divided forty mice into 4 groups with 10 mice in each group, PQ group was given 25% PQ solution (60 mg/kg) by one-time gavage. PQ+NR group were intraperitoneally injected with NR solution (300 mg/kg) 1 hour before given the same amount of PQ solution (60 mg/kg) by one-time gavage, The Control group were given the same amount of saline by one-time gavage, The same amount of NR was intraperitoneally injected before NR group were given saline by one-time gavage. Observed and recorded general condition of PQ intoxicated mice. Observed and recorded the death of mice every half an hour and counted the mortality and drew survival curve of each group after 72 hours exposure. another forty mice were averagely divided and treated by the same way. After 24 hours of modelling, mice were anaesthetized and killed. Then blood was extracted after eyeball was removed. The changes of TNF-a、IL-6 and MPO in serum of mice were detected by ELISA.Two lung tissues were removed from the chest and used to measure the D/W ratio of the lung. The pathological changes of lung were observed and scored under light microscope.The levels of SOD, MDA and Caspase-3 in lung tissues were determined by chemical colorimetry. The expression of Sirt1 and Nrf2 in lung tissues was detected by Western-blot. Results: Compared with the Control group and the NR group, the mice in the PQ group had a poor general condition, such as depression, crouching, skin disorder and reduced activity, food, urine and feces. The symptoms in the PQ+NR group were reduced compared with the PQ group. The survival rate at 72 hours after exposure: 80% in the PQ+NR group and 40% higher than that in the PQ group (P=0.029) . Compared with Control group and NR group, the D/W ratio (0.09±0.07) , lung pathology score under light microscope (11.80±0.37) , TNF-a (39.89±1.48) pg/ml、IL-6 (77.29±2.38) pg/ml、MPO (0.31±0.01) µg/ml、SOD (6.62±0.30) U/mgprot、MDA level (1.21±0.14) mmol/mgprot, Caspase-3 activity (356.00± 27.16) %, Sirt1 and Nrf2 protein expression (1.02±0.14、0.82±0.06) were significantly decreased in PQ group (P=0.004、0.023) ; Compared with PQ group, PQ+NR group significantly increased the D/W ratio (0.10±0.10) , decreased the pulmonary pathology score under light microscope (7.400.51) , decreased TNF-a (33.00± 0.65) pg/ml、IL-6 (52.23±4.23) pg/ml、MPO leve (0.23±0.01) µg/mll, increased SOD leve (9.28±0.45) U/mgprotl, decreased MDA level (0.78±0.02) mmol/mgprot, decreased Caspase-3 activity (222.80±7.59) %, and increased the protein expressions of Sirt1 and Nrf2 (1.62±0.16、1.06±0.04) (P=0.048、0.035) . Conclusion: NR can prolong the survival time of PQ poisoned mice; NR intervention can effectively inhibit the inflammatory response, peroxidation injury and apoptosis of PQ poisoned mice; NR intervention can upregulate the expression of Sirt1 and Nrf2 protein and effectively reduce the lung injury of PQ poisoning.

[Value of ATP synthase C subunit in predicting cardiac function and outcomes of sepsis].

Objective: To investigate the value of serum mitochondrial ATP synthase C subunit level in the evaluation of cardiac functional status and prognosis in patients with sepsis. Methods: A total of 165 sepsis patients admitted to the Emergency Intensive Care Unit (EICU) of the First Affiliated Hospital of Wenzhou Medical University from January 1, 2017 to December 31, 2018 were included, there were 103 males (62.4%) and 62 females (37.6%) with an age of (63±14) years. Human ATP synthase lipid binding protein (ATP5G1) ELISA kit was used to detect the level of serum ATP synthase C subunit within 24 h after admission to EICU, and compared with that in 45 healthy subjects. Clinical data of patients were collected and divided into groups according to different left ventricular ejection fraction (LVEF) and clinical outcomes. The differences in clinical indicators among each group were compared to evaluate the value of serum ATP synthase C subunit level in the evaluation of cardiac functional state and prognosis of patients with sepsis, and the independent risk factors for cardiac functional state and prognosis of patients with sepsis were analyzed. Results: Compared with the control group, the level of serum ATP synthase C subunit in the sepsis group was higher ((116±62) µg/L vs (77±34) µg/L, P<0.001). Compared with normal cardiac function group, the level of serum ATP synthase C subunit in septic cardiac dysfunction group was higher (P<0.001). Compared with the survival group, the level of serum ATP synthase C subunit in the death group was higher (P<0.05). The receiver operating curve (ROC) was drawn to analyze the value of ATP synthase C subunit, creatine kinase isoenzyme (CK-MB), B-type natriuretic peptide (BNP), troponin I (cTnI), left atrial end diastolic diameter, left ventricular end systolic diameter, left ventricular end diastolic volume, left ventricular end systolic volume in evaluating the cardiac function in patients with sepsis, and the area under the curve (AUC) was 0.928, 0.661, 0.837, 0.814, 0.703, 0.831, 0.794 and 0.765, respectively. The cut-off value, sensitivity and specificity of ATP synthase C subunit in it was 139.44 ng/L, 100% and 75.2%, respectively. ROC was drawn to analyze the prognostic value of age, urea nitrogen (BUN), ATP synthase C subunit, APACHEⅡ score and SAPSⅡ score in patients with sepsis, and the AUC was 0.719, 0.772, 0.656, 0.868 and 0.884, respectively. The cut-off value, sensitivity and specificity of ATP synthase C subunit in it was 131.24 ng/L, 61.9% and 68.7%, respectively. Logistic regression analysis showed that age, BUN, ATP synthase C subunit, cardiac dysfunction, APACHEⅡ score and SAPS Ⅱ score were independent risk factors for the prognosis of patients with sepsis. Conclusion: The level of serum ATP synthase C subunit is closely related to cardiac dysfunction in patients with sepsis, and can effectively predict the prognosis of patients with sepsis.

[Role of Orai 1-mediated store-operated calcium entry in the immune function of CD4+ T cells in septic mice].

Objective: To investigate the role of Orai1-mediated store-operated calcium entry in the immune damage of CD4+ T cells in septic mice. Methods: Sepsis mouse model was established by cecal ligation and puncture(CLP). Balb/c mice of clean grade were sacrificed 1, 3, and 5 days after operation. Spleen samples were harvested at given intervals. Splenic CD4+ T cells were selected by immunomagnetic beads and the expression of Orai1 protein was detected by western blotting, the storage operated calcium entry (SOCE) was detected by flow cytometry, the apoptosis of CD4+ T cells was detected by flow cytometry, the proliferation of CD4+ T cells was detected by CCK-8, and the IFN-γ and IL-4 were detected by enzyme-linked immunosorbent assay (ELISA). Then the expression of Orai1 protein was regulated to further detect the SOCE and immune function of splenic CD4+ T cells in mice. The experiment was divided into 4 groups, sham group, CLP3 group, Orai1 down group (Orai1-down group) and Orai1 up regulation group (Orai1-up group). Results: The relative expression of Orai1 protein in splenic CD4+ T cells in sham group was 1.03±0.16. Compared with sham group, Orai1 protein levels in CLP Group were all significantly lower (F=19.64, P=0.000 5). The increased value of splenic CD4+ T cells fluorescence intensity in sham group was 494±41. Compared with sham group, the levels of SOCE in CLP Group were all lower (F=30.01, P=0.001). The ratio of early and late apoptosis of CD4+ T cells in sham group was 8.7%±1.5%. Compared with sham group, the early and late apoptosis rates of CLP Group were significantly higher (F=32.29, P=0.000 1). The OD of sham group was 0.81±0.10 at 450 nm. Compared with sham group, the proliferation ability of splenic CD4+ T cells in CLP Group were significantly decreased (F=7.26, P=0.001 8). Compared with sham group, the secretion of IFN-γ and IL-4 by CD4+ T cells and the ratio of IFN-γ/IL-4 in CLP Group were all significantly decreased (F=19.690, 6.183, 11.230, all P<0.05). Compared with CLP3 group, the increased value of fluorescence intensity of CD4+ T cells was significantly decreased, the early and late apoptosis ratio of CD4+ T cells was significantly increased, the OD450 nm value of CD4+ T cells was decreased, the multiplication capacity of splenic CD4+ T cells were decreased, the level of IFN-γ and IL-4 secreted by T cells were decreased, and the value of IFN-γ/IL-4 in orai1-down group was decreased (t=4.819, 7.952, 2.988, 28.760, 3.140, 7.670, all P<0.05). However, Orail-up group showed the opposite trend. Conclusion: Orai1-mediated store-operated calcium entry can alleviate the immune dysfunction of CD4+ T cells in septic mice.

[Treatment report of three cases of multiple wasp stings].

From September to November 2019, three patients with Hornet stings were admitted to hospital with acute anaphylaxis and various degrees of organ dysfunction. After hemoperfusion combined with continuous hemodialysis and hormone pulse therapy, 1 patient was successfully treated and 2 patients were not cured. Bee Stings can cause systemic allergic reactions and toxic reactions. In severe cases, acute anaphylaxis, rhabdomyolysis, intravascular hemolysis, abnormal coagulation, and acute liver and kidney injury may occur, myocardial damage and other multi-organ function damage or even failure, is a clinical acute and critical illness, mortality is higher. Through the analysis of the diagnosis and treatment process of these 3 cases, the experience is summarized: the Bee sting should be treated as soon as possible, the early prevention and treatment of multiple organ dysfunction, the choice of accurate blood purification and comprehensive treatment, can effectively improve the prognosis, increase the success rate of treatment.

[Construction of prognostic model and identification of prognostic biomarkers based on the expression of long non-coding RNA in bladder cancer via bioinformatics].

OBJECTIVE: To construct the prognostic model and identify the prognostic biomarkers based on long non-coding RNA (lncRNA) in bladder cancer. METHODS: The lncRNA expression data and corresponding clinical data of bladder cancer were collected from The Cancer Genome Atlas (TCGA) database. The software Perl and R, and R packages were used for data integration, extraction, analysis and visualization. Detailly, R package "edgeR" was utilized to screen differentially expressed lncRNA in bladder cancer tissues compared with the normal bladder samples. The univariate Cox regression and the least absolute shrinkage and selection operator (Lasso) regression were performed to identify key lncRNA that were utilized to construct the prognostic model by the multivariate Cox regression. According to the median value of the risk score, all patients were divided into the high-risk group and low-risk group to perform the Kaplan-Meier (K-M) survival curves, receiver operating characteristic (ROC) curve and C-index, estimating the prognostic power of the prognostic model. In addition, the hazard ratio (HR) and 95% confidence interval (CI) of each key lncRNA were also calculated by the multivariate Cox regression. Moreover, we performed the K-M survival analysis for each significant key lncRNA from the result of the multivariate Cox regression. RESULTS: A total of 691 lncRNA were identified as differentially expressed lncRNA, and 35 lncRNA signatures were initially considered associated with the prognosis of bladder cancer, where in 23 lncRNA were identified as key lncRNA associated with the prognosis. The overall survival time in years of the low-risk group was obviously longer than that of the high-risk group [(2.85±2.72) years vs. (1.58±1.51) years, P<0.001]. The area under the ROC curve (AUC) was 0.813 (3-year survival) and 0.778 (5-year survival) respectively, and the C-index was 0.73. In addition, HR and 95%CI of each key lncRNA were calculated by the multivariate Cox regression and 11 lncRNA were significant. Furthermore, K-M survival analysis revealed the independent prognostic value of 3 lncRNA, including AL589765.1 (P=0.004), AC023824.1 (P=0.022)and PKN2-AS1 (P=0.016). CONCLUSION: The present study successfully constructed the prognostic model based on the expression level of 23 lncRNA and finally identified one protective prognostic biomarker AL589765.1, and two adverse prognostic biomarkers including AC023824.1 and PKN2-AS1 in bladder cancer.

[Baicalin regulates STIM1-mediated calcium overload and reduces apoptosis of cardiomyocytes induced by lipopolysaccharide].

Objective: To investigate the protective effect of Baicalin on apoptosis induced by lipopolysaccharide in H9C2 cardiomyocytes and its possible mechanism. Methods: In order to establish apoptosis model of H9C2 cardiomyocytes, H9C2 cardiomyocytes were cultured and divided into four groups: the control group; the baicalin group was treated with baicalin at the final concentration of 10µmol/L for 12 hours; the LPS group was stimulated with LPS at the final concentration of 1 µg/ml for 6 hours; The LPS+baicalin group was stimulated with LPS at the final concentration of 1 µg/ml for 6 hours within treated with baicalin at the final concentration of 10µmol/L for 12 hours. Collecting cell samples, CCK-8 (The Cell Counting Kit-8) was used to detect cell activity, and Terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling (TUNEL) was used to detect the expression levels of apoptosis. Laser Scanning Confocal Microscopy was used to detect the expression levels of store-operated calcium entry in H9C2 cardiomyocytes. Western blot was used to detect the protein expression levels of STIM1, cleaved-caspase3, Bax and Bcl-2. Fluorogenic quantitative PCR was used to detect the mRNA expression level of STIM1. Results: Compared with the control group, LPS-induced H9C2 cardiomyocyte survival rate decreased (P<0.05), the expression level of apoptosis increased (P<0.05), the internal flow of calcium increased (P<0.05), the expression levels of cleaved-caspase3, Bax protein levels increased (P<0.05), Bcl-2 protein level decreased (P<0.05), the expression of STIM1 mRNA and protein level increased (P<0.05). Compared with LPS group, the survival rate of H9C2 cardiomyocytes in baicalin intervention group increased (P<0.05), the expression level of apoptosis decreased (P<0.05), the internal flow of calcium decreased (P<0.05), the expression levels of cleaved-caspase3, Bax protein decreased (P<0.05), and the level of Bcl-2 protein increased (P<0.05), the expression of STIM1 mRNA and protein level decreased (P<0.05). Conclusion: Baicalin may alleviate LPS-induced cardiomyocyte apoptosis by alleviating calcium overload, and improve cell survival.

[Comparison of perioperative conditions between total and partial colpocleisis for elderly patients suffered from severe pelvic organ prolapse].

Objective: To study perioperative period conditions of total and partial colpocleisis with severe pelvic organ prolapse (POP) in elderly patients. Methods: From Jan. 2014 to Oct. 2017 286 severe POP elderly patients underwent total or partial colpocleisis. The mean age was (76.0±5.1) years (60-90 years) . Of which, 220 patients (76.9%, 220/286) presented more than one kind of medical disease. The operative time, intraoperative blood loss, hospital stay after surgery, postoperative complications, preoperative urinary dysuria symptoms of total and partial colpocleisis in treatment of 286 patients were analyzed. Results: In 286 patients, 172 patients (60.1%) underwent total colpocleisis and 114 patients (39.9%) underwent partial colpocleisis. Totally 48 patients (16.8%) in 286 patients underwent anti-urinary incontinence procedure. The mean operating time of 286 patients was (67±22) minutes,the mean blood loss was (52± 57) ml, the mean hospital stay after surgery was (5.9±1.9) days (2-16 days) . The rate of postoperative complications was 8.7% (25/286) . Those complications including 1 case of bladder injury during operation, 3 cases of postoperative pelvic hematoma, 1 case of intestinal obstruction, 5 cases of atrial fibrillation, 10 cases of postoperative morbidity, 3 cases of pulmonary embolism, 1 case of lower limb thrombosis, 1 case of urinary retention underwent electrical stimulation treatment because of ineffective application of urinary retention catheter. The mean operating time was (67±22) minutes in total colpocleisis and (68±20) minutes in partial colpocleisis (P>0.05) , the mean blood loss was (58±62) ml in total colpocleisis and (41±45) ml in partial colpocleisis (P>0.05) , the mean hospital stay after surgery was (5.9±1.9) days in total colpocleisis and (6.0±1.8) days in partial colpocleisis (P>0.05) , the rate of post operative complications was 8.1%(14/172) in total colpocleisis and 9.6% (11/114) in partial colpocleisis (P>0.05) , respectively; those four of comparisons showed no significant difference. There were 129 patients (45.1%, 129/286) with voiding difficulty before surgery; the mean postvoid residual volumes of the above two operation types of people after operation were (35±43) and (34±41) ml, which showed no significant difference (P>0.05) . Conclusions: Colpocleisis is a safe and effective management in selected elderly patients with severe POP, who no longer desire to maintain vaginal coital function. There is no significant difference in the perioperative period conditions between total and partial colpocleisis. The choice of surgical procedure is based on factors such as the presence or absence of malignancy in the uterus, age and willingness of the patient, and general condition.

[Clinical analysis of 22 cases of syndrome caused by acute poisoning].

Objective: To analyze the clinical characteristics, treatment and prognosis of rhabdomyolysis (RM) caused by acute poisoning.Summarize the clinical characteristics and treatment experience, pay attention to the complications and improve the quality of rescue. Methods: We collecte and summarize the clinical data, treatment and prognosis of 22 cases of RM caused by acute poisoning. Results: We found that 21 patients (95.5%) had muscle damage, 13(59.1%) with coma, 8(36.4%) with brown, tea or even soy sauce urine, 6(27.3%) had acute renal injury (AKI), and 4(18.2%) had multiple organ dysfunction syndrome (MODS). After the treatment, 21 cases (95.5%) got better, and one case were discharged. All the patients with AKI were survived, three of them were treated by hemodialysis, and the other recovered gradually after massive fluid replacement. Conclusion: Acute poisoning combined with RM is not uncommon in clinic. We should pay attention to examination of serum enzymes and other indicators, observe the clinical symptoms and make early diagnosis. The key to diagnosis and treatment is early fluid resuscitation, comprehensive treatment, blood purification and maintain the stability of water and electrolyte.

[Comparison of different scoring systems in prognosis evaluation of acute poisoning].

Objective: To compare the predictive value of PSS, APACHEII, SAPSII and SOFA in the prognosis evaluation of acute poisoning. Methods: Clinical data (including PSS score, APACHEII score, SAPSII score and SOFA score, within 24 hours after admission) of 231 acute poisoning patients admitted to the emergency intensive care unit EICU of our hospital from January 2015 to October 2016 was retrospectively analyzed. The patients were divided into the survival group and the dead group according to the 28-day clinical outcomes, comparing the differences of clinical data in each group. To analyze the correlation between PSS score, APACHEII score, SAPSII score and SOFA score in each group, comparing the value and the area under the ROC curve of four scoring systems and evaluate the predictive value of the four scoring systems. Results: Comparing with the survival group and the dead group, PSS score, APACHEII score, SAPSII score and SOFA score were significantly different (P<0.01) . PSS score, APACHEII score, SAPSII score and SOFA score were significantly positive correlation (P<0.01) , the area under the ROC curve (AUC) of the four scoring systems were 0.833, 0.887, 0.843 and 0.843 respectively. The area under the ROC curve (AUC) of APACHEII score was higher than PSS score, SAPSII score and SOFA score, the difference was statistically significant (z=2.351, 2.317, 2.217; P=0.019, 0.021, 0.027) , there was no significant difference in the area (AUC) between the three scoring curves (P>0.05) . The cutoff value (cut-off) , sensitivity, specificity and accuracy rates of PSS score, APACHEII score, SAPSII score and SOFA score were (2.5, 93.1%, 50.9%, 61.5%) , (14.5, 82.8%, 75.7%, 77.48%) , (31.5, 77.6%, 76.90%, 77.08%) , (5.5, 77.60%, 74.60%, 75.35%) . Conclusion: PSS score, APACHEII score, SAPSII score and SOFA score can evaluate the prognosis of patients with acute poisoning, but the APACHEII score is better than the other three scoring systems in evaluating the prognosis for its evaluation ability and accuracy rate.

[Analyses on relevant factors of the prognosis of elderly patients with acute poisoning].

Objective: To explore the risk factors influencing the prognosis of elderly patients with acute poisoning. Methods: We retrospected 177 elderly patients with Acute Poisoning who were treated in the emergency department of the first affiliated hospital of wenzhou medical university from July 2009 to May 2015. According to the outcome of patients, we distributed the patients to death group (31 cases) and survival group (146 cases) , compared the clinic data and using multivariate analysis with Logistic regression to prognosis factors. Results: There were 177 cases in total, with 146 survivors (82.5%) and 31 deaths (17.5%) . In which 102 cases (57.6%) had chronic underlying diseases. There were 28 cases of pesticide poisoning in the death group, and the fatality rate of pesticide poisoning was 23.5%. The mortality rate was 12.8% in the 60-69 years-old group (11/86) , 20% (13/65) in the 70-79 years-old group, 26.9% (7/26) in the 80-89 years-old group. The most common reason of poisoning was intentional ingestion, with 100 cases (56.5%) . The tract of the poisoning was mainly in digestive system, including 148 cases (83.6%) . The PSS score and APACHE-II score were 2.97±0.18 and 19.8±2.8 in the death group, 2.27±0.81 and 12.8±5.3 in the survival group. Compared with the survival group, poison (pesticides or non) 、poisoning route、cause of poisoning、PSS score、APACHEⅡ score have significant difference in death group (P<0.05) . Poison (pesticides or non) 、PSS score、APACHEⅡ, were the independent risk factors of poor prognosis. Conclusion: Most of the elderly patients with acute poisoning have one or more chronic underlying diseases, the digestive tract ingestion and pesticide poisoning are more common. The fatality rate of the old patients is significantly higher than that of non elderly poisoning. Type of toxications, PSS score and APACHE-II score are the prognostic factors in elderly patients.

[Value of base excess in predicting the prognosis of patients with paraquat poisoning].

Objective: To explore prognostic factors of acute paraquat poisoning (APP) , analyze the correlation between base excess (BE) and plasma concentration of paraquat (C-PQ) and discuss BE level in evalua-tion of prognosis of acute paraquat poisoning patients. Methods: We retrospectively selected 84 APP patients who admitted to Emergency Intensive Care Unit (EICU) of our hospital from 2009.9.1 to 2015.8.31.Clinical data from 84 APP patients were analyzed. BE、C-PQ、time of gastric lavagesince ingestion、time of hemoperfusion since ingestion、severity index of paraquat poisoning (SIPP) 、white blood cell (WBC) 、percentage of neutrophils (N%) 、hemoglobin (HB) 、creatinine (Cr) 、alanine aminotransferase (ALT) 、aspartate aminotransferase (AST) 、partial pressure of oxygen (PaO(2)) 、partial pressure of carbon dioxide (PaCO(2)) and other laboratory parameters were measured. A total of 41 patients in non-survivors died during the 30 days after admission and 43 patients in survivors survived during the 30 days. The factors of prognosis in paraquat poisoining and the role of BE in evalu-ating prognosis was analyzed, as well as the correlation between BE and C-PQ. Results: 1.Logistic regression analyses showed BE、C-PQ、ALT、AST、Cr was of prognostic significance[odds ratio (OR) of BE: 0.511, 95%CI 0.267, 0.978; C-PQ:-=0.999, 95%CI 0.999, 1.000; both P<0.05] ; 2.The area under the receiver operating characteristic curve (ROC curve) of BE、C-PQ and prognosis were 0.775、0.927 respectively, BE≤-1.7 mmol/L was the best cut-off value, the sensitivity、specificity for predicting were 82.9%、62.8%, the evaluation value was lower to C-PQ>3 273.935 ng/ml (AUC 0.927, 78.0%、95.3%) ; 3.BE negative correlated with C-PQ[-1.100 (-4.100, -0.200) , -5.900(-8.650, -2.500) , both P<0.05]. (r=-0.4, P<0.01). Conclusion: These results suggest that BE may be useful for the prediction of prognosis in PQ poisoning and BE negative correlated with C-PQ.

[Mono-carbonyl analogues of curcumin prevents paraquat-induced apoptosis in HK-2 cell line by inhibiting oxidative damage and inflammation].

Objective: To investigate the effects of mono-carbonyl analogues of curcumin (L6H21) on paraquat (PQ) -induced injury in HK-2 cell line and explore its underlying mechanisms. Methods: Cultured HK-2 cells were challenged by PQ with or without L6H21 treatment. Cell viability and apoptosis were determined by CCK-8 assay and flow cytometry, respectively. Gene expressions and protein levels of apoptotic and inflammatory factors were assessed by RT-PCR, ELISA, and western blot. Intracellular ROS production was detected by DCFH-DA staining. Superoxide dismutase (SOD) and malondialdehyde (MDA) were examined by chemical colorimetry. Results: 1) PQ challenge significantly inhibited HK-2 cells proliferation, which was prevented by L6H21 administration. PQ dramatically induced HK-2 apoptosis evidenced by increasing expressions of caspase-9, caspase-3 and Bax, while decreasing Bcl-2 level. However, PQ induced these apoptotic effects in HK-2 cells were reversed by L6H21. Similarly, PQ exposure obviously enhanced activity of NF-κB and levels of cytokines (TNF-α、IL-6) in HK-2 cells, which was inhibited by L6H21. Furthermore, administration of L6H21 inhibited PQ induced ROS and MDA production, and promoted SOD level in HK-2 cells. Conclusion: L6H21 administration inhibits PQ-induced apoptosis in HK-2 cells possibly by reducing inflammation and oxidative damage.

Amino acid composition of royal jelly harvested at different times after larval transfer.

The amino acids in royal jelly (RJ) have a wide range of pharmacological and health-promoting functions in humans. Multiple studies on the amino acid quality and composition in RJ have investigated RJ harvested at 72 h after larval transfer. In contrast, the concentration of amino acids in RJ harvested before 72 h remains unknown. In this study, the concentration of free amino acids (FAAs) and total amino acids (TAAs) in RJ harvested at 13 time points between 24 and 72 h after transfer of ten Apis mellifera colonies were measured. Our results indicated that the most abundant FAAs were Pro, Phe, Lys, Glu, and Tyr, whereas the most abundant TAAs were Asp, Glu, Leu, Lys, and Val. The total FAA concentration in RJ increased with increasing harvest time, from 4.30 mg/g at 24 h to 9.48 mg/g at 72 h. In contrast, the variation in concentration of TAAs observed was a decrease-increase-decrease trend with 40 h (149.53 mg/g) and 52 h (169.62 mg/g) as inflection points. The highest and lowest concentrations of TAA were 197.96 and 121.32 mg/g at 24 and 72 h, respectively. To our knowledge, this is the first study to investigate the concentration changes of FAAs and TAAs prior to 72 h after transfer. Our results will provide theoretical support to guide production practices of beekeeping, as well as elucidate the relationship between the harvest time point and RJ content.

[The effects of P - glycoprotein expression induced by ulinastatin on HK - 2 cells damage induced by paraquat].

Objective: To investigate the protective effect of P-glycoprotein up-regulated by ulinastatin (UTI) on HK-2 cells during paraquat (PQ) -induced injury and its underlying mechanisms. Methods: The re- search was divided into two parts. The first part of the research was divided into normal control group, PQ group, UTI+PQ group, UTI control group. The second part of the research was divided into negative virus group (including control group, PQ group, PQU+TI group, UTI group) and P-gp siRNA group (including control group, PQ group, PQU+TI group, UTI group) . Negative virus group: the cells were transfected into the blank virus; siRNA P-gp group: the cells were transfected with P-gp siRNA virus. HK-2 cells were routinely cultured. After 800 µmol/L PQ treatment, the changes of P-gp protein levels in the HK-2 cells were determined by West-ern-blot (WB) . Then, transfected lentivirus bringing P-gp silent gene, the cell viability was determined by CCK-8 assay, the expression of P-gp in the cells after transfection was detected by WB and the concentration of PQ in HK-2 cells were measured by high performance liquid chromatography (HPLC) . Results: Compared with the normal control group, the P-gp expression of PQ group had no significantly changes (P>0.05) . Compared with the PQ group, the P-gp expression of UTI+PQ group significantly increased (P>0.05) . Compared with the corre-sponding control siRNA group, the P-gp siRNA group had no significantly changes in cell viability (P>0.05) . and significantly decreased in P-gp expression. Compared with the corresponding control siRNA group, the P-gp siRNA group had no significantly changes in PQ concentration in HK-2 cell (P>0.05) , but compared with P-gp siRNA PQ group, the PQ concentration of P-gp siRNA PQ+UTI group significantly decrease (P<0.05) . Conclusion: UTI significantly reduced the accumulation of PQ in HK-2 cells and increased the viability of HK-2 cells in vitro may be not by increased P-gp activity. UTI could significantly reduce HK-2 cell injury induced by PQ in vitro and improve the survival rate of HK-2 cells. It may not be related to the up regulation of P-gp expres-sion.

[Mechanism research and effect of ulinastatin in the brain tissue injury of acute hydrogen sulfide intoxicated rats].

OBJECTIVE: To observe the effect of Ulinastatin (UTI) in the dynamic changes of aquaporin 4 (AQP4) and cyclooxygenase-2 (COX-2) in the brain tissue injury of acute hydrogen sulfide (H2S) intoxicated, to explore the Mechanism of brain tissue injury of acute H2S-intoxicated and the protection effect of UTI. METHODS: A total of 96 SD rats of clean grade were divided randomly into four groups: normal control group (NS group, n=8) , UTI control group (UTI group, n=8) , H2S-intoxicated model group (H2S group, n=40) , UTI treatment group (H2S+UTI group, n=40). The H2S group and H2S+UTI group were exposed to H2S (284 mg/m(3)) by inhalation for 1 h, then H2S+UTI group was intraperitoneally exposed to UTI at the dose of 10(5) U/kg for 2 h, H2S group and H2S+UTI group were sacrificed at 2, 6, 12, 24 and 48 h after exposure, respectively. Remove the brain tissue, observe the rats behavioral changes at each time points. The mRNA expression of AQP4、COX-2 and NSE in the brain tissue were measured by RT-PCR method, and the protein expression of AQP4、COX-2 and NSE in the brain tissue were detected by immunohistochemical Streptavidin-perosidase method. Pathological changes of brain tissue were observed by lightmicroscope. RESULTS: 1、Nerve cells in the H2S group rats had edema, degeneration, focal inflammatory cell infiltrate, capillary hyperplasia, expansion. Compared with NS group, the cerebral NSE mRNA and protein expression at each time point in H2S group after exposure were significantly increased (P<0.01). 2、Compared with NS group, the cerebral AQP4 and COX-2 mRNA and protein expression at each time point in H2S group after exposure were significantly increased (P<0.01). 3、The degree of brain damage was significantly decreased in H2S+UTI group than that in H2S group. Compared with H2S group, the cerebral NSE mRNA and protein expression at 6, 12, 24 and 48 h in H2S+UTI group after exposure were significantly decreased (P<0.01) , no significantly difference at 2h (P>0.05). 4、Compared with the H2S group, the cerebral AQP4 and COX-2 mRNA and protein expression at 6, 12, 24 and 48 h in H2S+UTI group after exposure were significantly decreased (P<0.01) , slightly decreased at 2 h. CONCLUSION: The mechanism of brain injury of acute hydrogen sulfide intoxicated associated with abnormal expression of the cerebral AQP4, COX-2 levels. Intervention of UTI can reduce the cerebral AQP4 and COX-2 levels after hydrogen sulfide intoxicated, reduce the degree of brain injury.

Molecular cloning and evolutionary analysis of captive forest musk deer bitter taste receptor gene T2R16.

Sensing bitter tastes is crucial for most animals because it can prevent them from ingesting harmful food. This process is mainly mediated by the bitter taste receptors (T2R) that are largely expressed in the taste buds. Previous studies have identified some T2R gene repertoires. Marked variation in repertoire size has been noted among species. However, research on T2Rs is still limited and the mechanisms underlying the evolution of vertebrate T2Rs remain poorly understood. In the present study, we analyzed the structure and features of the protein encoded by the forest musk deer (Moschus berezovskii) T2R16 and submitted the gene sequence to NCBI GenBank. The results showed that the full coding DNA sequence (CDS) of musk deer T2R16 (GenBank accession No. KP677279) was 906 bp, encoding 301 amino acids, which contained ATG start codon and TGA stop codon, with a calculated molecular weight of 35.03 kDa and an isoelectric point of 9.56. The T2R16 protein receptor had seven conserved transmembrane regions. Hydrophobicity analysis showed that most amino acid residues in T2R16 protein were hydrophobic, and the grand average of hydrophobicity (GRAVY) was 0.657. Phylogenetic analysis based on this gene revealed that forest musk deer had the closest association with sheep (Ovis aries), as compared to cow (Bos taurus), Tursiops truncatus, and other species, whereas it was genetically farthest from humans (Homo sapiens). We hope these results would complement the existing data on T2R16 and encourage further research in this respect.

Application and a proposed modification of the 2010 McDonald criteria for the diagnosis of multiple sclerosis in a Canadian cohort of patients with clinically isolated syndromes.

BACKGROUND: The 2005 and 2010 McDonald criteria utilize magnetic resonance imaging (MRI) to provide evidence of disease dissemination in space (DIS) and time (DIT) for the diagnosis of multiple sclerosis (MS) in patients who have clinically isolated syndromes (CIS). METHODS: Data from 109 CIS patients not satisfying the 2005 criteria at entry into a randomized controlled minocycline trial were analyzed to determine the proportion who would have been diagnosed with MS at screening based on 2010 criteria. The impact of including symptomatic, as well as asymptomatic, MRI lesions to confirm DIT was also explored. RESULTS: Thirty percent (33/109) of patients, retrospectively, met the 2010 criteria for a diagnosis of MS at baseline. When both symptomatic and asymptomatic lesions were used to confirm DIT, three additional patients met the 2010 criteria. There was a significant 10.1% increase in the proportion of patients who met the 2010 DIS criteria, compared with the 2005 DIS criteria; however, two patients satisfied the 2005 DIS but not 2010 DIS criteria. CONCLUSION: Using 2010 McDonald criteria, 30% of the CIS patients could be diagnosed with MS using a single MRI scan. Inclusion of symptomatic lesions in the DIT criteria further increases this proportion to 33%.