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Objective@#To develop a method for rapid detection of influenza virus and its subtypes based on flow fluorescence technology, and then detect the clinical specimens by our established method .@*Methods@#We designed degenerate primers and specific probes, synthesized plasmids, used Luminex platform to establish detection method and later detected 430 clinical specimens, and compared the result with those of Fujian Center for Disease Control and prevention.@*Results@#A method for the simultaneous determination of influenza viruses A, B, C and its subtypes (H116, N19) was established. The time consumption was 3.5 hours, with good specificity, high sensitivity and feasible stability. The detection result of 430 clinical specimens showed high consistency with the result of Fujian Center for Disease Control and Prevention.@*Conclusions@#We established a method for simultaneous determination of influenza viruses and its subtypes, high sensitivity, specificity and stability.
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Objective To analyze the clinical manifestations of pulmonary fungal disease in patients with different pathogen infection and different immune states. Methods A retrospective analysis was carried out with the clinical data of 78 patients with pulmonary fungal disease who were treated in Fujian Provincial Hospital from June 2012 to June 2016. Results The identified pathogens included Cryptococcus (78.2％, 61/78), Aspergillus(17.9％, 14/78), Talaromyces marneffei, Mucor, and Trichosporon asahii (one each). Cryptococcus was mostly found in healthy individuals without underlying disease and Aspergillus largely infected those with underlying disease. Diabetes mellitus was the most common underlying disease. Pulmonary fungal infection was confirmed by pneumonectomy (41.0％, 32/78), CT-guided percutaneous transthoracic biopsy (43.6％, 34/78), transbronchial lung biopsy (14.1％, 11/78), or blood culture (1.3％, 1/78). Pulmonary cryptococcosis was often asymptomatic (47.5％, 29/61). Hemoptysis was only found in the patients with underlying disease. The patients with pulmonary aspergillosis showed higher prevalence of hemoptysis (57.1％, 8/14) than the patients with other pulmonary fungal diseases. Bronchoscopy usually gave negative finding in case of pulmonary cryptococcosis (77.8％, 14/18). Inflammatory exudation was the primary finding of pulmonary aspergillosis (6/10). Intraluminal necrosis and neoplasia were only found in the aspergillosis patients with underlying disease. The primary imaging findings in pulmonary cryptococcosis was nodule or mass (78.7％, 48/61). Halo sign and crescent sign were rarely found in pulmonary aspergillosis. Of the 78 patients, 45 (57.7％) patients received pharmacological therapy alone, 15 (19.2％) surgical treatment alone, and 18 (23.1％) received drugs in combination with surgery. Six patients died, 25 lost to follow-up, and 47 with stable disease. Conclusions The clinical characteristics of pulmonary fungal disease vary with the pathogen and the immune states of patients. Clinical symptoms and immune status of patients should be taken into account when making diagnosis of pulmonary fungal disease for the purpose to speculate the probable fungal pathogen and choose the most appropriate diagnostic tool.
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AIM: To investigate the role of autophagy in the apoptosis of human pulmonary artery endothelial cells (HPAECs) induced by cigarette smoke extract (CSE).METHODS: HPAECs were cultured routinely.HPAECs were treated with CSE at different concentrations, and the cell viability was detected by MTT assay.HPAECs were divided into control group, CSE group, 3-methyladenine (3-MA) group and 3-MA+CSE group.The autophagy was observed under fluorescence microscope with monodansylcadaverine (MDC) staining.Annexin V/propidium iodide staining and Hoechst 33342 staining were employed to detect apoptosis.In addition, the protein levels of LC3, beclin-1 and cleaved caspase-3 were determined by Western blot.RESULTS: MDC staining showed the increased production of autophagic vacuoles was observed in CSE group.The results of Western blot showed that the expression levels of autophagy-related proteins LC3 and beclin-1 were increased, while 3-MA pretreatment inhibited the expression of these proteins and the production of autophagic vacuoles.Observation with Annexin V/propidium iodide staining and Hoechst 33342 staining showed that the apoptotic rate in CSE group was significantly higher than that in control group, and pretreatment with 3-MA induced further increase in the cell apoptosis.The protein level of cleaved caspase-3 in CSE group was significantly higher than that in control group (P<0.05), and 3-MA+CSE treatment induced the further increase in the protein level of cleaved caspase-3.CONCLUSION: CSE induces autophagy and apoptosis in the HPAECs.Inhibition of autophagy promotes the apoptosis induced by CSE in HPAECs, which can be achieved through activation of caspase-3.
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Objective To observe the ultrastructure of blood-brain barrier in the nude mouse model of brain me-tastases from lung cancer by transmission electron microscopy using lanthanum nitrate tracing.Methods PC-9 cells (1 × 106/0.1 mL) in logarithmic phase were respectively injected into six nude mice ( model group) selected from eight nude mice randomly via the left ventricle, the other two mice without any treatment as the control group.The general status of the mice was observed after implantation.In the fourth week all the mice were sacrificed and brain tissue samples were taken and prepared for transmission electron microscopic observation using lanthanum nitrate tracing.besides, the lung and brain were removed and stained with HE to detect the presence of tumor metastasis.Results Mice in the model group began to lose weight almost simultaneously in the third week and became moribund slowly, and were all sacrificed at the fourth week when showing clear signs of cachexia.At autopsy, the thoraxes were clear, with normal lungs.Histology showed evidence of brain metastasis in all the six mice.The electron microscopy showed that lathanum nitrate tracer was escaped from the capillaries and diffusely or sparsely distributed in the brain tissues of the model group mice, however lathanum nitrate tracer was still confined in the capillary lumen in the mice of control group.Conclusions The diffuse lathanum nitrate tracer in the brain parenchymal tissue indicates the impairment of blood-brain barrier in the nude mouse model of lung cancer brain metastasis and the formation of these metastases is accompanied with the destruction of blood brain barrier.
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Objective To establish an appropriate animal model of brain metastases from lung cancer in nude mice by thoracic orthotopic implantation in the chest or left ventricular injection , and to serve further studies on the mechanisms of lung cancer brain metastasis .Methods PC-9 cells (1 ×106/0.1 mL) in logarithmic phase were respectively injected into 18 nude mice by orthotopic implantation in the chest or left ventricular injection ( n=9 each group ) .The statuses of nude mice were observed after implantation .Animals showing clear signs of dyscrasia were killed .At autopsy, the lung, brain, liver and kidney were removed and histological sections were stained with H /E to detect the presence of tumor cells . Results In the thoracic orthotopic implantation group , three weeks after implantation , the number 4, 6, 9 mice showed tumor nodules in the chest wall , they began to lose weight in the fourth to sixth week differently , showing signs of dyscrasia gradually , and were sacrificed at the fifth to seventh week .The thoracotomy revealed that the whole thorax was occupied by many large lung cancer masses , spreading into bilateral ribs , pleura and spinal vertebra , with scarce eroded , compressed , pale and distorted lung tissues left .Histological examination with HE staining showed the presence of neoplasms in their lung tissues but only the number 6 mouse showed metastatic lesions in the brain tissue .In the left ventricular injection group, the mice almost began to lose weight in the third week simultaneously and became moribund slowly , which were all sacrificed at the fourth week .After thoracotomy , the thoraxes were clear except the number 11 and 18 mice which appeared 2-3 tiny tumor foci in the chest wall , with normal lung tissues .Histological examination with HE staining showed the pres-ence of brain metastases in all the nine mice .The rate of brain metastases from lung cancer in the left ventricular injection group was 100%, compared with 11.1% in the thoracic orthotopic implantation group .Conclusions The establishment method of mouse model by left ventricular injection shows significantly higher rate of lung cancer brain metastases than that by thoracic orthotopic implantation .
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Objective To explore the anti-hyperuricemia activity of bergenin in the model of hyperuricemic mice induced by potassium oxonate. Methods 60 Kunming male mice were divided randomly into six groups, which were normal control group;hyperuricemic model group;and hyperuricemic groups with 20 , 40 , 60 mg/kg berge-nin, and 5 mg/kg allopurinol. Mice were orally administered once daily with 250 mg/kg potassium oxonate for 7 continuous days to create the model, and then three doses of bergenin and allopurinol were orally initiated on the day 1 h after potassium oxonate was given, separately. Serum uric acid, creatinine and urea nitrogon levels, as well as urinary uric acid and creatinine levels were measured. mRNA and protein expression levels of mouse kidney u-rate transporter 1(URAT1), and glucose transporter 9(GLUT9) were also determined. Results Compared with hyperuricemic model group, bergenin significantly reduced serum uric acid, creatinine and urea nitrogon levels, in-creased 24 h uric acid and creatinine excretion, and fractional excretion of uric acid in hyperuricemic mice;mRNA and protein levels of mURAT1 and mGLUT9 were also markedly down-regulated. Conclusion Anti-hyperuricemia effect of bergenin is attributed to the enhancement of uric acid excretion and reversion of mouse urate transporters o-ver-expression.
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<p><b>BACKGROUND</b>The extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae has increasingly become a major contributor to nosocomial infections and can exhibit multiple antibiotic resistance. Previous studies have focused on the resistance genes in ESBL-producing strains, and the resistance-associated genetic environment of non-ESBL-producing strains has been ignored until now. Here, we investigated the occurrence and characteristics of non-ESBL-producing K. pneumoniae, which potentially carries unexpressed resistance genes.</p><p><b>METHODS</b>K. pneumoniae strains were collected from five medical institutions in China from February 2010 to August 2013. The VITEK-2 ESBL detection system was used as a primary screen to identify the ESBL-producing phenotype, and the three primary types of ESBL-associated genes (CTX, SHV, and TEM) were detected by polymerase chain reaction (PCR) to confirm the strains presenting with a non-ESBL-producing phenotype. mRNA expression in the non-ESBL-producing strains was further screened by reverse-transcription PCR (RT-PCR) to validate their transcriptional efficiency.</p><p><b>RESULTS</b>Out of 224 clinically isolated antibiotic-sensitive K. pneumoniae strains with a non-ESBL-producing phenotype, 5 (2.2%) were identified to carry inactivated ESBL blaSHV genes with intact upstream promoter regions and resistance gene sequences. Interestingly, three of the five antibiotic-sensitive K. pneumoniae strains containing ESBL blaSHV genes still exhibited mRNA transcription of blaSHV, while the other two exhibited no mRNA transcription.</p><p><b>CONCLUSION</b>These findings suggest that inactivated ESBL genes exist in non-ESBL-producing antibiotic-sensitive K. pneumoniae strains, which have the potential to transform the strain into an ESBL phenotype if an inappropriate application or overdose of antibiotics is implemented during clinical management.</p>
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Humans , Anti-Bacterial Agents , Pharmacology , China , Drug Resistance, Multiple, Bacterial , Genetics , Klebsiella pneumoniae , Genetics , Microbial Sensitivity Tests , beta-Lactamases , GeneticsABSTRACT
Objective:To observe the effect of ketogenic diet on the growth of human colon cancer cells in nude mice and to de-termine its possible mechanisms. Methods:A total of 24 male BALB/C nude mice were injected subcutaneously with the tumor cells of the colon cancer cell line HCT116. These animals were randomized into two feeding groups. One group was fed with a ketogenic diet (KD group;n=12), and the other group was given a standard diet (SD group;n=12) ad libitum. Experiments were completed upon at-taining a target tumor volume of 600 mm3 to 700 mm3. The two diets were compared based on body weight, serum glucose, ketone body, insulin, tumor growth, and survival time, which is the interval between tumor cell injection and attainment of target tumor vol-ume. Results:The tumor growth was significantly more delayed in the KD group than in the SD group. Tumors in the KD and SD groups reached the target tumor volume at 33.8 ± 6.7 days and 24.8 ± 3.1 days, respectively. The ketone body in the KD group was ele-vated with a slight reduction in serum insulin, and the difference in serum glucose in the two groups was insignificant. Importantly, the KD group had significantly larger necrotic areas and less vessel density than the SD group. Conclusion:The application of an unre-stricted ketogenic diet delayed tumor growth in a mouse xenograft model. Further studies are needed to address the mechanism of this diet intervention and its effect on other tumor-relevant functions, such as invasive growth and metastasis.
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Objective To establish an optimal animal model of pulmonary metastasis of human lung adenocarcino-ma, to serve further investigation of mechanism of lung adenocarcinoma metastasis.Methods Eleven nude mice aged 4-6 weeks were used in this study.Suspension of human lung adnocarcinoma A549 cells (0.1 mL, 107 cells/mL) was injec-ted into the tail vein in nude mice.From four weeks after inoculation, two nude mice were killed each time at 4, 5, 6 weeks after the tumor cell injection at random for examination.The remaining 3 mice were killed at the end of the experi-ment.At autopsy, the lung, brain, liver, kidney and other organs were removed, fixed in neutral buffered formalin and embedded in paraffin.Sections were cut and stained with hematoxylin-eosin, and examined by histopathology.The number of metastatic foci was counted.Results No mouse died after tumor cell inoculation.Serially euthanized mice revealed evi-dence of gradually increasing pulmonary metastases in the mice:No metastasis was found before 4 weeks after tumor cell in-oculation, the first histological metastases appeared at 5 weeks, gross metastatic foci were observed at 6 weeks, widely spread metastatic foci were observed at 7 weeks, and the remain 3 mice developed cachesia at 11, 13, and 14 weeks after tumor cell inoculation.Mediastinal lymph node metastases were found in the nude mice by 11 weeks after tumor cell inocu-lation.Conclusions We have successfully established a nude mouse model of pulmonary metastasis by injecting human lung adenocarcinoma A549 cells into the tail vein.
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Objective To investigate and compare the change of extravascular lung water (EVLW) and levels of cytokines in septic patients without clinical acute lung injury (ALI) /acute respiratory distress syndrome (ARDS) with those in spetic patients with sepsis-induced ALI/ARDS in order to determine the role of EVLW involved in the pathogenesis of lung injury in the patients by quantifying the relationship between EVLW and biomarkers of lung injury in patients with sepsis.Methods A total of 40 septic patients complicated either with or without clinical ALI/ARDS after sepsis. In each patient,transpulmonary thermodilution (PiCCO) was used to measure cardiovascular hemodynamics and EVLWI for 7 days via an arterial cannula indwelled within 72 hours after diagnosis of severe sepsis was made, and serial bronchoalveolar lavages (BAL) were carried out.Other examinations including blood gas analysis,ventilator parameters,chest X-ray and the levels of pro-inflammatory cytokines such as tumor necrosis factor (TNF-oα),interleukin-1 in the BAL were recorded.In-hospital and ICU mortalities were also observed.Results Of total 40 patients,29 were complicated with clinically defined septic ALI/ARDS ( ARDS n =15,and ALI n =14).The septic patients complicated with ALL/ARDS had significantly higher amount of EVLWI and higher levels of TNF-α and interleukin-1 in the BAL than patients without ALI/ARDS ( P ＜ 0.05).The arterial oxygen tension/fractional inspired oxygen ratio,lung injury score,and the levels of TNF-α,IL-1 in the BAL correlated with EVLWI.Moreover,in-hospital mortality,ICU mortality and the length of ICU stay of the patients with high amount of EVLWI were markedly increased than those of patients with low amount of EVLWI. Conclusions In septic patients complicated with ALI/ARDS, the extravascular lung water index correlates with oxygenation,lung injury severity and inflammatory cytokines in lung.Determination of EVLWI may be useful for evaluation of severity of lung injury and prognosis of septic patients.
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Objective To analyze the alterative characteristics of the extravascular lung water (EVLW ) in the patients with septic shock and clarify its value on the prognosis of these patients.Methods By the methods of retrospective analysis,according to the ultimate survival,21 patients with septic shock were divided into survivor group (10 cases) and non-survivor group (11 cases).The clinical features of the patients were observed and hemodynamic monitoring was made with PiCCO monitor.The EVLW was measured and the relationship between the EVLW and the prognosis of patients was analyzed.Results On the first,second and third day,EVLW was (12.7 ±1.8),(11.3 ±1.3),(10.1 ±1.3) ml/kg in survivor group,and (14.4 ± 1.0),(14.6 ± 1.4),(14.6 ±1.3) ml/kg in non-survivor group respectively,and there were statistical differences between two groups (P 0.05).Conclusions The EVLW in the patients with septic shock increases significantly.The dynamic changes of the EVLW may be one of the factors for predicting the prognosis of patients with septic shock.
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Objective To explore the effect of mild hypothermia on inflammation status,lung function protection and clinical prognosis in patients with acute respiratory distress syndrome (ARDS).Methods All of 56 patients with ARDS were randomly divided into two groups: trial group (29 patients,treatment with mild hypothermia) and control group (27 patients, treatment with common practice). The following parameters including tumor necrosis factor (TNF)-α,interleukin (IL)-6 and C reactive protein (CRP), oxygenation index, SOFA evaluation and injury of lungs evaluation were detemined before treatment and at the 3rd, 7th day after treatment, and survival rates and adverse reaction in 28 days also were observed.Results After treatment, the levels of TNF-α ,IL-6 and CRP were decreased significantly, and oxygenation index, the scores of SOFA evaluation and injury of lungs evaluation were improved significantly in trial group than those in control group (P<0.05 ). The survival rate in trial group was higher than that in control group after treatment of 28 days [65.5%(19/29) vs 51.9%(14/27)]. The courses of mechanical ventilation and staying in ICU in trial group were shorter than those in control group [(11.9±3.6)d vs (17.0±5.1)d,(14.1±4.2)d vs (21.5±7.7)d](P<0.05). Conclusion Mild hypothermia can effectively attenuate inflammation disorder, improve damaged lung function and prognosis in patients with ARDS.
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Objective To study the effect of continuous blood purification on inflammation and nutritional status in patients with sepsis, and to study relationship between inflammatory, malnutrition and illness. Method Forty-eight patients with severe sepsis were randomly divided into two groups: continuous renal replacement treat-ment (CRRT) group (n=27) and control group (n=21). The flowing biomarkers including albumin, preallbu-min, transfenin, insulin-like growth factor-1 (IGF-1), tumor necrosis factor (TNF-α) and C reactive protein (CRP) were determined before,and 1 week and 2 weeks after treatment. Results Compared with control group, levels of albumin, prealbumin and IGF-1 in treatment group increased significantly at 14 days after CRRT (P<0.05), and levels d CRP and TNF-α decreased significantly (P<0.05), resulted in increase in survival rate, shortened me-chanical ventilation time and decreased SOFA scores (P<0.05). The IGF-1 and prealbumin had a negative cor-relations with CRP and TNF-a in both groups (P<0.05). The survivors in both groups had significantly higher levels of IGF-1,prealbumin and TNF-α than the deads before trearment(P<0.05). Conclusions Continuous blood purification can effectively attenuate inflammation and improve nutritional status in patients with severe sep-sis.Maybe IGF-1 and prealbumin act as prognostic markers more sensitive in severe sepsis.
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<p><b>BACKGROUND</b>Livin is a novel inhibitor of apoptosis protein(IAP),recent studies showed that it overexpressed in many carcinomas including lung cancer and contributed much to the cancerous development.The objective of this study is to explore the expression of the two isoforms of livin in lung cancer tissues and their relationship with histological types and chemotherapy,and to explore their relationship to the expression of caspase-3 as well.</p><p><b>METHODS</b>Expression of livin α,livin β and caspase-3 mRNAs were detected by reverse transcription polymerase chain reaction(RT-PCR) assay in lung cancer tissues as well as in controls.</p><p><b>RESULTS</b>Livin α and livin β were expressed in 12 of 27 and 19 of 27 lung cancer tissues respectively,much higher than those in lung para-cancerous(0/6,0/6) or benign disease lung tissues(0/12,1/12)(both P < 0.01).Moreover,the positive rate was 7/14 and 9/14 in lung adenocarcinoma and 4/12 and 9/12 in squamous and large cell carcinoma respectively,and both of them were detected in one small cell carcinoma.The levels of these two isoforms in lung cancer were significantly higher than those in controls by Gel Imaging System(both P < 0.05),the level of livin α was remarkably higher in adenocarcinoma than that in squamous cell carcinoma(P < 0.05),while the level of livin β was similar in both carcinomas(P > 0.05).Meanwhile,the level of caspase-3 in lung cancer was significantly lower than that in controls,the levels of either each of two isoforms or their sum were negatively associated with that of caspase-3(P < 0.05,P < 0.01,P < 0.01).Two isoforms of livin mRNA expression seemed to increase after chemotherapy but not related to clinical stages(P > 0.05).</p><p><b>CONCLUSIONS</b>Two isoforms of livin are differently expressed in different histological types of lung cancer and may contribute to corresponding cancerous development;the levels of livin are negatively associated with those of caspase-3,this may due to the fact that livin could resist against apoptosis;high expression of livin seems to be related to chemotherapy but not clinical stage.</p>
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Objective To explore the clinical features and the cause of secondary phosgene poisoning after rescuing the patients with acute phosgene poisoning.Methods According to the diagnostic criteria of occupational acute phosgene poisoning,the differences of clinical manifestation,laboratory results and chest X-ray between secondary poisoning patients and primary patients were compared.Results Among the 25 patients with secondary phosgene poisoning,14(56.0%) had cough,13(52.0%)had throat stimulus,10(40.0%)had chest stuffiness,2(8.0%)had polypnea,1(4.0%) had pain in the eye.There was no significant difference in clinical manifestation between the secondary and primary patients. No positive sign was found after the examination of pulmonary function in the 25 patients,but all of them had abnormal chest X-ray,and typical bronchitis could be found.According to the diagnostic criteria,the 25 patients had slight acute phosgene poisoning,and recovered after treatment for 7 to 10 days.Conclusion To prevent the secondary phosgene poisoning after treating the patients with acute phosgene intoxication,medical workers should enhance protection awareness and take some necessary measures.