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Objective:To investigate the effects of biologics on psychological status and quality of life in patients with inflammatory bowel disease (IBD).Methods:A cross-sectional survey was conducted in 42 hospitals in 22 provinces (autonomous regions and municipalities directly under the central government) from September 2021 to May 2022. General clinical information and the use of biologics were obtained from adult patients diagnosed with IBD who voluntarily participated in the study. Psychological status was evaluated using the Generalized Anxiety Disorder (GAD-7), Patient Health Questionnaire-9 (PHQ-9), Pittsburgh Sleep Quality Index (PSQI), and Inflammatory Bowel Disease Questionnaire (IBDQ) assessment tools. Counts were analyzed via the Chi-square test, and datasets that were not normally distributed were analyzed via nonparametric tests. P<0.05 was considered statistically significant. Results:A total of 2 478 valid questionnaires were collected. The GAD-7 score of the biologics group was significantly lower than that of the non-use group [6 (2, 9) vs. 7 (3, 10), Z=-3.49, P<0.001]. IBDQ scores [183 (158, 204) vs. 178 (152, 198), Z=-4.11, P<0.001], intestinal symptom scores [61 (52, 67) vs. 58 (49, 65), Z=-5.41, P<0.001], systemic symptom scores [28 (24, 32) vs. 27 (23, 31), Z=-2.37, P=0.018], emotional ability scores [69 (58, 77) vs. 67 (56, 75), Z=-3.58, P<0.001] and social ability scores [26 (22, 29) vs. 25 (22, 29), Z=-2.52, P=0.012] in the biologics group were significantly higher than in the non-use group. GAD-7 scores [5 (2, 9) vs. 6 (3, 10), Z=-3.50, P<0.001] and PSQI scores [6 (4, 9) vs. 6 (4, 9), Z=-2.55, P=0.011] were significantly lower in the group using infliximab than in the group not using it. IBDQ scores were significantly higher in patients using vedolizumab than in those not using it [186 (159, 205) vs. 181 (155, 201), Z=-2.32, P=0.021] and were also significantly higher in the group treated with adalimumab than in the group not treated with adalimumab [187 (159, 209) vs. 181 (155, 201), Z=-2.16, P=0.030]. However, ustekinumab had no significant effect on any of the scores. Conclusion:The use of biologics is strongly associated with improvements in anxiety status and quality of life in IBD patients.
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Objective:To investigate the psychology status and quality of life in patients with inflammatory bowel disease(IBD) in China, and to analyze the influencing factors.Methods:From September 2021 to May 2022, 42 hospitals in 22 provinces(autonomous regions and municipalities directly under the central government) in China, the clinical data of 2 478 IBD patients were collected, which included age, gender, weight, first visit or not, disease activity, disease course, main clinical manifestations(diarrhea, abdominal pain, hematochezia, extraintestinal manifestations), complications, treatment medication(5-aminosalicylic acid, glucocorticoids, immunosuppressive agents, and biological agents), and whether to have surgery. Anxiety, depression, sleep quality and quality of life of IBD patients were evaluated by generalized anxiety disorder-7 items, patient health questionnaire-9 items, Pittsburgh sleep quality index and inflammatory bowel disease questionnaire, and the related influencing factors were analyzed. Univariate analysis and multiple linear regression analysis were used for statistical analysis.Results:The average age of 2 478 IBD patients was 37.96 years old, and male counted for 62.43%(1 547/2 478). There were 61.82%(1 532/2 478) of the IBD patients in the active stage of disease, mostly mild or moderate(588 and 734 cases). There were 60.61%(1 502/2 478) of the IBD patients with different degrees of anxiety, 58.35%(1 446/2 478) of the IBD patients with different degrees of depression, and 48.87%(1 211/2 478) of the IBD patients had different degrees of sleep problems. The results of multiple linear regression analysis indicated that female, higher level of disease activity and longer disease course were independent risk factors of anxiety, depression and sleep quality in the IBD patients(unstandardized regression coefficient(95% confidence interval) 1.08(0.65 to 1.50), 0.45(0.23 to 0.68), 0.19(0.02 to 0.36), 0.83(0.33 to 1.32), 0.62(0.36 to 0.88), 0.28(0.08 to 0.47), 0.47(0.16 to 0.77), 0.39(0.23 to 0.55), 0.14(0.02 to 0.26); P<0.001, <0.001, =0.025 , =0.001, <0.001, =0.005, =0.003, <0.001, =0.027). The usage of biological agents was an independent protective factor of anxiety(unstandardized regression coefficient(95% confidence interval) -0.67(-1.17 to -0.17), P=0.008), and older age was an independent risk factor of sleep quality(unstandardized regression coefficient(95% confidence interval) 0.35(0.09 to 0.61), P=0.008). Higher level of disease activity, symptoms of diarrhea, abdominal pain, presence of extraintestinal manifestations, usage of 5-aminosalicylic acid and glucocorticoid, and with surgical treatment were independent risk factors of quality of life(unstandardized regression coefficient(95% confidence interval) -11.00(-12.24 to -9.76), -2.90(-5.26 to -0.55), -3.93(-6.25 to -1.61), -5.79(-9.87 to -1.71), -4.78(-7.79 to -1.76), -7.71(-11.07 to -4.35), -4.37(-8.00 to -0.73); P<0.001, =0.016, =0.001, =0.005 , =0.002, <0.001, =0.019), while the usage of biological agents was an independent protective factor of quality of life (unstandardized regression coefficient(95% confidence interval) 4.72(1.97 to 7.48), P=0.001). Conclusion:IBD patients generally have different degrees of anxiety, depression and sleep problems, which affect the quality of life of patients. Gender, disease activity and disease course are the influencing factors of mental disorders in IBD patients.
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Objective To investigate the status of medical institution sewage treatment in Qingpu District of Shanghai, and to make corresponding measures in improving sewage management in medical institutions. Methods Questionnaire survey was conducted on selected 95 medical institutions in Qingpu District to investigate sewage treatment status and sewage samples were collected from 30 medical institutions with 20 beds or above to test their sanitary quality. Results Operating rate of sewage handling equipment was 98.9%, the proportion of medical institutions using chlorine disinfectant was 87.4%, the total qualified rate of residual chlorine was 66.7%, the total qualified rate of fecal coliform number was 70.0%, and overall qualified rate was 60.0%. Conclusion The qualified rate of medical institutions sewage in Qingpu District is not optimistic enough.It is necessary to strengthen sewage personnel training and health supervision to ensure the safety of medical institution sewage.
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Objective To explore the drug resistance mechanisms of carbapenem-resistant Citrobacter freundii (C.freundii) and its treatment strategies.Methods Clinical data of 17 strains of carbapenem-resistant C.freundii from this hospital were collected.Carbapenemase resistant genes were amplifies by polymerase chain reaction (PCR).The agar dilution method and the broth dilution method were used to determine the minimal inhibition concentration (MIC) of single antimicrobial drug and drug combination,the partial inhibitory concentration index (Σ FICI) was calculated.Results Eight strains were found to produce blaNDM-1 and 9 strains produced blaIMP,blaKPC,blaSPM and blaoxA-48 were not detected in the study.Furthermore,the synergistic effect and addictive effect of fosfomycin combined imipenem accounted for 75.00％,in which,the synergistic effect was up to 56.25 ％.the synergistic effect and addictive effect of fosfomycin and cefoperazone/sulbactam accounted for 50.00％.Conclusion Fosfomycin combined with imipenem or cefoperazone/sulbactam has good antibacterial activity in vitro,but imipenem combined with fosfomycin may have better effect.
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Objective To explore the drug resistance mechanisms of carbapenem-resistant Citrobacter freundii (C.freundii) and its treatment strategies.Methods Clinical data of 17 strains of carbapenem-resistant C.freundii from this hospital were collected.Carbapenemase resistant genes were amplifies by polymerase chain reaction (PCR).The agar dilution method and the broth dilution method were used to determine the minimal inhibition concentration (MIC) of single antimicrobial drug and drug combination,the partial inhibitory concentration index (Σ FICI) was calculated.Results Eight strains were found to produce blaNDM-1 and 9 strains produced blaIMP,blaKPC,blaSPM and blaoxA-48 were not detected in the study.Furthermore,the synergistic effect and addictive effect of fosfomycin combined imipenem accounted for 75.00％,in which,the synergistic effect was up to 56.25 ％.the synergistic effect and addictive effect of fosfomycin and cefoperazone/sulbactam accounted for 50.00％.Conclusion Fosfomycin combined with imipenem or cefoperazone/sulbactam has good antibacterial activity in vitro,but imipenem combined with fosfomycin may have better effect.
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Objective To do screening acute lymphoblastic leukemia patients scFv antibody single chain variable region to cre-ate conditions for the expression and obtain further specificity of antibody fragments.Methods In this study,patients with newly diagnosed acute lymphoblastic leukemia serum as coating antigen using phage display technology,screening phage an-tibody specificity from the semi-synthetic human phage antibody libraries,the first to target the immune antigen-coated tab-let,phage library was added,so that with the target antigen-specific binding phage antibody was immobilized on plates immu-nization,could not be specifically bound phages were rinsed.The eluted specific binding phage,E.coli infection.Could get the specific antibody gene containing phagemid.Results After three “adsorption-elution-amplification”screening process,got stronger leukemia patient antigen-specific phage antibody variable region fragment and identification.Conclusion Got better strain affinity antibody fragments,to create the conditions for the next fragment expression,identification and clinical appli-cation.
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Objective To investigate the protective effect of continuous intravenous infusion of Isoproterenol (ISO)on myocardial mitochondria of early septic rats and the corresponding mechanism.Methods Thirty Sprague Dawley (SD)rats were randomly divided into 5 groups (6 cases per group):control group,endotoxin group,ISO small-dose group,ISO medium-dose group and ISO large-dose group.Endotoxin group and ISO intervene group received same management apart from drug intervention:receiving intravenous injection of lipopolysaccharide (LPS)10 mg/kg followed by an continuous intravenous infusion of 9 g/L saline 1 mL/h or ISO 0.06 μg/(kg · min),0.30 μg/(kg · min)and 0.60 μg/(kg · min).Control group received intraperitoneal injection and continuous intravenous infusion with the same amount of 9 g/L saline.The primary endpoint of the study was 24 hours after injection of 9 g/L saline or LPS.Serum creatine kinase (CK) and creatine kinase isoenzyme (CK-MB),oxidative and nitrosative stress levels and swelling of isolated heart mitochondrion were detected.The pathological changes of the myocardium and morphologic changes of the heart mitochondria were observed through light microscope and scanning electron microscope,respectively.Results The levels of CK,CK-MB,nitric oxide (NO) content,inducible nitric oxide synthase (iNOS) activity and malondialdehyde (MDA)content in endotoxin group were increased compared with control group (all P ＜ 0.05),while the superoxide dismutase (SOD) activity decreased [(11.543 ± 1.080) U/mg prot vs (9.892 ±0.815) U/mg prot,P ＜0.05].The morphology of the heart mitochondria significantly changed (such as swelling,disordered arrangement,crest fracture,fusion and cavitations,and so on).ISO intervention significantly decreased the levels of CK,CK-MB and mitochondrial swelling (all P ＜ 0.05) and increased the SOD activity (all P ＜ 0.05).The levels of NO content,iNOS activity and MDA content were significantly decreased in small-dose group [(10.823 ± 2.240) μmol/g prot vs (7.917 ± 2.203) μmol/g prot,(0.045 ± 0.008) U/mg prot vs (0.033 ± 0.003) U/mg prot,(1.663 ± 0.618) mmol/mg prot vs (0.768 ± 0.312) mmol/mg prot,all P ＜ 0.05],while the levels of iNOS activity and MDA content were significantly increased in medium-and large-dose group (all P ＜ 0.05) ; compared with medium-dose group,the degree of mitochondrial swelling in large-dose group increased (1.160 ± 0.186 vs 1.393 ± 0.128,P ＜ 0.05).The pathological changes of the myocardium mitochondria significantly improved.Conclusions The myocardium and myocardial mitochondria of early septic rats were damaged,continuous intravenous infusion of low-dose ISO revealed protective effect on these damages,and the corresponding mechanism may relate to the decrease of the oxidative and nitrosative stress.
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<p><b>OBJECTIVE</b>To preliminarily investigate the long-term structural and functional injuries of mitochondria in rat brain caused by sepsis.</p><p><b>METHODS</b>Wistar rats were randomly assigned into sepsis and control groups. A rat model of sepsis was prepared by an intraperitoneal injection of 10 mg/kg lipopolysaccharide (LPS) of gram-negative bacteria, and the survival assay was performed. Eight rats in the sepsis group were sacrificed at 12, 24, 48, or 72 hours after LPS injection, while rats in the control group were sacrificed after an intraperitoneal injection of an equal volume of normal saline. Mitochondria were extracted from rat brain tissue. Mitochondrial membrane potential (MMP) and mitochondrial swelling level were determined by flow cytometry, and the activities of electron transport chain complexes (I-V) were measured using enzyme assay kits. Hematoxylin-eosin (HE) staining and electron microscopy were used to observe morphological changes in brain tissue and mitochondria.</p><p><b>RESULTS</b>The sepsis group had a significantly lower survival rate than the control group (P<0.01). The MMP and activities of electron transport chain complexes (I-V) in the sepsis group, which were significantly lower than those in the control group (P<0.05), were reduced to the lowest levels at 48 hours and partially recovered at 72 hours. The mitochondrial swelling level in the sepsis group, which was significantly higher than that in the control group (P<0.05), increased to the peak level at 48 hours and partially recovered at 72 hours. Hematoxylin and Eosin staining revealed substantial damages in the structure of brain tissue, and electron microscopy showed mitochondrial swelling, and vacuolization in a few mitochondria.</p><p><b>CONCLUSIONS</b>In the rat model of LPS-induced sepsis, both structural and functional injuries are found in cerebral mitochondria, and achieve the peak levels probably at around 48 hours.</p>
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Animals , Male , Rats , Brain , Pathology , Lipopolysaccharides , Toxicity , Membrane Potential, Mitochondrial , Mitochondria , Physiology , Rats, Wistar , Sepsis , MortalityABSTRACT
Objective To investigate the effect of concurrent exercise intervention in metabolic endotoxemia induced by high-fat diet in rats,and further understand the damage of liver mitochondrial ultramicrostructure.Methods Eighteen SD rats with the weight of 100g were randomly divided into 3 groups:group A (standard diet group),group B(high-fat diet group) and group C (treadmill-trained group with high-fat diet).Training (1 hour/d) initiated at the same time as the HF diet was fed.After being raised for 6 weeks,the rats was euthanized and weighed up.Blood samples were taken and the levels of serum lipid were detected.The levels of serum endotoxin were detected by enzyme-linked immunoadsorbent assay.The membrane potentials of isolated mitochondrion were detected by flow cytometry instrument and the morphologic changes in mitochondria in liver were observed by electronic microscopy.Results In group B,the levels of endotoxin increased significantly(2.916 ± 0.761 rs 5.454 ± 1.254,t =-4.236,P ＜ 0.05),and the liver mitochondrial density and membrane potential also increased significantly compared with group A after 6 weeks (4.330 ±0.501 vs 3.507 ±0.532,t =2.759,P ＜0.05;l.660 ±0.202 vs 0.473 ±0.064,t =13.712,P ＜0.05).But there was no markedly different in serum endotoxin between group B and group C (4.972 ± 1.757 vs 5.454 ± 1.254,t =-0.547,P ＞ 0.05).Compared with group B,the liver mitochondrial density of group C decreased significantly (4.330±0.501 vs 3.581 ±0.188,t =3.426,P ＜ 0.05).The mitochondrial ultrastructurctural changes in each group were not obvious.Conclusions The rats fed with high-fat diet for 6 weeks can reach the state of metabolic endotoxemia.The increasing levels of the liver mitochondrial membrane potential caused by metabolic endotoxin may affect the happening and development of other diseases in the future.Concurrent exercise can not decrease the level of endotoxin.It also shows that metabolic disease caused by high-fat diet should be prevented by moderation in eating and drinking.
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The objective of this study was to evaluate the value of morphologic diagnosis for acute leukemia (AL), to explore the relation of morphologic diagnosis with immunology, cytogenetics and molecular biology diagnosis of AL and to analyze the onset characteristics of AL in 10 years. The samples of bone marrow and peripheral blood from 233 newly diagnosed cases of AL were collected during 2001-2011 years; the morphologic examination and immunologic, cytogenetic and molecular biologic examination (ICM) were carried out, the consistency of morphologic diagnosis with ICM diagnosis was compared, the onset characteristics of AL was analyzed. The results showed that: (1) the consistent rate of immunology, cytogenetics, molecular biology diagnosis with morphologic diagnosis was 84.3%. The order of consistent rat was AUL, M0 < M1 < HAL < M4 < M2 < M3 < M5 < ALL < M6, M7, AP; (2) Misdiagnosis always occurred among AUL, M0, M1, ALL and HAL or among M2a, M3v, M4 and M5. (3) In 233 cases, the highest ratio of blast was observed in M1 (92.5%), while the lowest ratio of blast was observed in M2 (49.5%). (4) AL occurred more frequently in males than that in female (147:86). (5) AL occurred in patients aged from 1 to 88 years. The median age was 41.5 for AUL, 40.8 for M0, 43.4 for M1, 46.3 for M2, 33.8 for M3, 42.6 for M4, 48.8 for M5, 77.3 for M6, 2.5 for M7, 65.0 for AP, 29.1 for ALL and 40.3 for HAL. (6) The number of patients in the later five years (139 cases) was significantly greater than that in the first five years (94 cases), especially the patients with M1, M2, M3, M4, and M5. It is concluded that morphologic diagnosis has important clinical value in the MICM diagnosis of AL. Attaching importance to the confusing cell morphology and onset characteristics of AL can improve the diagnostic accuracy.
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Adolescent , Adult , Aged , Aged, 80 and over , Child , Child, Preschool , Female , Humans , Infant , Male , Middle Aged , Young Adult , Acute Disease , Cytogenetic Analysis , Leukemia , Diagnosis , Pathology , Molecular Biology , Retrospective StudiesABSTRACT
Objective To explore the effect of TNF-α in intracellular Ca~(2+)-overload associated myocardial injury. Methods Thirty Wistar rats were randomly divided into three groups:control group,Ca~(2+)+-paradox group,and pentoxifylline treatment group. The intracellular Ca~(2+)-overload rat model was established by pre-filling the rats with Langendorff for 20 minutes,isolated rat hearts subjected to Ca~(2+)-depletion for 5 minutes and Ca~(2+)-repletion for 30 minites(Ca~(2+)-paradox). Changes in hemodynamics indexes were monitored continuously. TNF-α in cardiac tissues was tested by ELISA method,and nuclear factor-κB(NF-κB)in cardiac tissues was detected with Western blot. Results Dramatic depression in left ventricle contraction function was found in the Ca~(2+)-paradox hearts:significant decrease in left ventricular diastolic pressure(LVDP),markedly elevated left ventricular end diastolic pressure(LVEDP),decreased dP/dt ratio,increased TNF-α content,decreased cytosolic/homogenate NF-κB ratio. All these changes in Ca~(2+)-paradox group were significantly attenuated upon the treatment with 100 μmol/L pentoxifylline. Conclusions Activation of NF-κB and increased production of TNF-α may play important roles in cardiac injury associated with intracellular Ca~(2+)-overload.
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Objective To screen permethrin human single-chain variable region (scFv) antibody for aims of developing rapid detection kit. Methods Phage display technology was used in this study. Permethrin was solid phase coated on Nunc plate as antigen. Semi-synthetic single-chain variable region of human antibody library technology was applied, and single chain variable region was screened from phage antibody library after 3 rounds "adsorption - elution - amplification" of the selection process. 100 clones were random selected as resistance to permethrin clones , enzyme-linked immunosorbent assay (ELISA), crossreactivity and competitive inhibition experiments were used to validate permethrin binding activity with strong scFv clones from the selected phage antibody clones plasmid. The plasmid was digested with restriction enzyme Sfi Ⅰ / Not Ⅰ and subcloned into pCANTAB5E vector. After transformed into E. coli XL1BIue, the plasmid was identified by restriction enzyme analysis. Results After screening in 100 clones, 18 clones had high ELISA absorbance values ( A value) at 490nm wavelength ( A490nm), then bovine serum albumin (BSA) cross-reactions identified five weak cross-reaction. Combined with the triplicate ELISA and competitive inhibition experiment results, one positive clone was acquired at last. And this clone was subcloned into pCANTAB5E vector and transformed into competent cells XL1-Blue. Conclusion Plasmid fragment was consistent with the purpose, which provided the foundation for further study of its specific affinity.
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This study was aimed to investigate the influence of cryopreservation on biological properties and function of leukemic dendritic cells (L-DCs) derived from patients with acute or chronic leukemia. Some fresh leukemic cells were detected immediately; some were cultured immediately; some were cryopreserved in -80 degrees C with 5% DMSO-6% HES as cryopreservor. After being thawed, they were cultured. The combination of rhGM-CSF, rhIL-4, rhTNF-alpha and other cytokines were added into the culture system. 12 days later, L-DCs were assayed for morphology, immunophenotype, mixed lymphocytic reaction (MLR) and CTL cytotoxicity on autologous leukemic cells. The results showed that both fresh and cryopreserved leukemic cells obtained from patients with acute or chronic leukemia revealed typical DC morphologically by means of using combinations of cytokines in culture, but there was no significant difference between pre-or post cryopreservations. L-DCs also upregulated the expression of CD80, CD54, HLA-DR, CD1a, CD83 and CD86, and downregulated the expression of CD14, but there was also no difference as compared with L-DCs befor cryopreservation. L-DCs derived from leukemic cells were also capable of stimulating MLR and inducing CTL which could kill autologous leukemic cells obviously. It is concluded that leukemic cells, regardless of fresh or frozen, can induce L-DCs after culture with cytokine combination. The L-DCs can induce CTL targeting autologous leukemic cells, and may be used to treat MRD as immunotherapy. The induction and biological properties of L-DCs are not influenced by cryopreservation.
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Humans , Bone Marrow Cells , Cell Biology , CD8 Antigens , Metabolism , Cryopreservation , Dendritic Cells , Cell Biology , Allergy and Immunology , Granulocyte-Macrophage Colony-Stimulating Factor , Pharmacology , Interleukin-4 , Pharmacology , Leukemia, Myeloid , Allergy and Immunology , Pathology , Lipopolysaccharide Receptors , Metabolism , Lymphocyte Activation , Recombinant Proteins , T-Lymphocytes, Cytotoxic , Allergy and Immunology , Tumor Cells, Cultured , Tumor Necrosis Factor-alpha , PharmacologyABSTRACT
<p><b>OBJECTIVE</b>To establish a BALB/c mice model system of cytomegalovirus-induced myocarditis.</p><p><b>METHODS</b>Twenty five specific pathogen-free inbred female BALB/c mice (5 weeks old, 16 - 18 g, seronegative for MCMV) were infected with 1 x 10(4) PFU MCMV by the intraperitoneal (i.p.) route. All experimental mice were sacrificed at 3, 5, 7, 10, 14 days i.p. (n = 5 per time point). Hearts were removed under aseptic conditions, and were transected along the midline. One part of each heart was processed with Bouin's fixative for histological examination. The other part of each heart was immediately frozen in liquid nitrogen and stored at -80 degrees C until MCMV titre was determined by plaque assay. Serum cTnI level was assayed by ELISA.</p><p><b>RESULTS</b>MCMV was detected in the hearts at extremely low levels on 3 days i.p. and could not be detected on 10 days i.p. A mixed cellular infiltrate composed of polymorphonuclear neutrophils and mononuclear lymphocytes was observed on 3 days, which reached a peak at 7 to 10 days after MCMV infection and was maintained for at least 3 - 4 months postinfection. Serum cTnI levels were elevated on 3 days i.p., reaching a peak at 7 to 10 days i.p..</p><p><b>CONCLUSIONS</b>These data highlight the possible therapeutic uses of antiviral drugs in viral myocarditis as well as further elucidating the pathogenic nature of the disease.</p>
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Animals , Female , Mice , Disease Models, Animal , Herpesviridae Infections , Pathology , Mice, Inbred BALB C , Muromegalovirus , Myocarditis , Virology , Troponin I , MetabolismABSTRACT
<p><b>OBJECTIVE</b>Human rhinovirus (HRV) is the most common respiratory pathogen, which causes not only acute respiratory infection and community acquired pneumonitis in children, but also asthma episode and deterioration. However, the detection of respiratory pathogen, which mainly focuses on respiratory syncytial virus, influenzaviruses A and B, parainfluenza viruses 1-3 and adenoviruses, does not include HRV yet by now in China. The absence of detection method limits the clinical understanding of HRV pathogenicity, and causes unreasonable use of antibiotics. This study aimed to establish a one-step reverse transcription (RT) PCR system for detecting specific fragment of HRV RNA, and to analyze the sequences of amplicons.</p><p><b>METHODS</b>A pair of degenerate primers based on the HRV highly conserved 5'' noncoding region (NCR) were used to develop a one-step RT-PCR system for detecting HRV RNA in nasopharyngeal aspirates from 78 children with acute respiratory tract infections in the spring of 2004. All the positive PCR products were sequenced, and the sequences of the nucleotides were analyzed by using biological software and compared with those in the GeneBank.</p><p><b>RESULTS</b>Eleven (14.1%) of 78 samples were positive on RT-PCR, these patients were clinically diagnozed as upper respiratory tract infection (n = 7), bronchitis (n = 3) and bronchopneumonia (n = 1), respectively. Compared with the sequences of clinical and standard HRV viruses in the GeneBank, the nucleotide sequences of these 11 amplicons shared high homology of 89%-95.5%. Within the 11 amplicons, nucleotide identity varied from 75.2% to 91.8%, and the ratio of genetic variation was from 8.8% to 31.0%, which occurred in highly conserved regions and usually showed single nucleotide mutation in some special locations. These 11 amplicons attribute to the two branches of HRV cladogram, respectively. Most of mutations in highly conservative domain occurred on single ribonucleotide, mainly as transversion (C/G, A/G) and transition (T/C, A/G), some were mutations among 3 bases (A/C/G, A/T/G, A/C/T). And a few mutations involved two nearby ribonucleotide which were also found in highly conservative domain. However, ribonucleotide deletion and insertion were usually found in highly variable domain.</p><p><b>CONCLUSION</b>The findings showed that this one-step RT-PCR system was highly specific, rapid and convenient for the detection of HRV RNA in nasopharyngeal secretions of patients with acute respiratory tract infections and that the genome of HRV viruses was highly variable.</p>
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Child , Child, Preschool , Female , Humans , Male , Base Sequence , Genes, Viral , Molecular Sequence Data , Picornaviridae Infections , Diagnosis , Virology , RNA, Viral , Respiratory Tract Infections , Virology , Reverse Transcriptase Polymerase Chain Reaction , Methods , Rhinovirus , Genetics , Sensitivity and Specificity , Sequence Analysis, DNAABSTRACT
The objective was to observe the influence of previously activated psoralens on the proliferation of K562 cells, and to provide laboratory data for its clinical usage. K562 cells were treated separately with previously and late activated psoralens, then their trypan blue exclusion inhibited rates (TBIR), cell proliferation inhibited rates (CPIR) and colony forming inhibited rates (CFIR) after culture were compared. The results showed that previously activated psoralens displayed an inhibiting effect on the proliferation of K562 cells with a dose-effect relationship. There was no obvious difference between previously and late activated psoralens on TBIR, CPIR and CFIR. In order to exert the inhibitive effect of previously activated psoralens, the time of ultraviolet ray exposure should be 10 minutes at least, and longer than 12 hours for inhibiting K562. The inhibitive effect of previously activated psoratens decreased as the time interval from activation to its use was prolonged. The inhibiting effect of previously activated psoralens was strongest within 6 hours after activation. In conclusion, both previously and late activated psoralens show inhibiting effects on the proliferation of K562, which may be able to use an antineoplastic drug in clinic.
Subject(s)
Humans , Cell Proliferation , Dose-Response Relationship, Drug , Furocoumarins , Pharmacology , K562 Cells , Time FactorsABSTRACT
<p><b>AIM</b>To observe the basical properties of adherent stromal cells in culture derived from cryopreserved bone marrow cells (BMCs), and to provide laboratory evidences for clinical application of cryopreserved stromal cells.</p><p><b>METHODS</b>Fresh BMCs and adherent stromal cells cultured for 14 days in Dexter long-term culture system (fresh stromal cells) plus 5% DMSO-6% HES cryopreservatives were frozen in -80 degrees C refrigerator, and cryopreserved in - 196 degrees C liquid nitrogen for 2 weeks (the former is called cryopreserved BMCs, the latter called cryopreserved stromal cells). These cells were cultured in Dexter long-term culture system after they were thawed. We have examined the growth features, constituents and stimulating functions of the culture of these cells.</p><p><b>RESULTS</b>Growth features: Cryopreserved BMCs produced adherent stromal cells, cell clusters and cell layer 1-2 days later than fresh BMCs. Cryopreserved stromal cells formed cell layer in 2nd day of culture, and were 12-18 h later than fresh stromal cells. Cryopreserved BMCs and stromal cells proliferated significantly less than fresh BMCs and stromal cells. Constituents: The ratio of fibrocytes and endothelial cells were lower, and the ratio of macrophages and fat cells were higher in culture of cryopreserved BMCs than that in fresh BMCs. The measurements in culture of cryopreserved stromal cells were much more significant compared with that in fresh stromal cells. Numbers of cells containing apoptic bodies in culture of cryopreserved BMCs and stromal cells were more than that in fresh BMCs and stromal cells. TBRR of cryopreserved BMCs and stromal cells were 92.5% and 89.5% respectively. The expression rates of CD14 and HLA-DR in culture of cryopreserved BMCs and stromal cells were higher than that in fresh BMCs and stromal cells, and just in contrast with the expression rates of CD45 and CD33. Stimulating functions: CAFA and LTC-IC on the stromal cell layer derived from cryopreserved BMCs, cryopreserved stromal cells, fresh BMCs and fresh stromal cells were all growing well, and there were no significant differences among these groups.</p><p><b>CONCLUSION</b>Biological properties of adherent stromal cells derived from BMCs and stromal cells were injured slightly and still maintained completely after cryopreservation with 5% DMSO-6% HES cryopreservatives.</p>
Subject(s)
Humans , Bone Marrow Cells , Cell Biology , Cell Survival , Cells, Cultured , Cryopreservation , Stromal Cells , Cell BiologyABSTRACT
<p><b>AIM</b>To provide experimental basis for clinical use of cryopreserved dendritic cells (DCs) by investigating the biological properties of cryopreserved DCs derived from cord blood.</p><p><b>METHODS</b>The biological discrepancies between unfrozen DCs (UFDC) derived from unfrozen cord blood (UFCB) and DCs from cryopreserved cord blood (CPCB) or cryopreserved DCs (CPDCs) were explored by morphological observation and immunophenotype analysis. Moreover, the stimulating index (SI) in mixed lymphocyte culture and cytotoxic eliminating rate (ER) were measured by MTT.</p><p><b>RESULTS</b>The TBR of CPCB and CPDCs were 95.8% and 88.7% respectively. Compared to UFCB, CPCB differentiated toward DCs in a delayed and decreased mode, displayed lower expression of CD1a, CD83 and HLA-DR, and had reduced SI and ER. Similarly, the CPDCs became adherent DCs later and grew less in number than UFDCs. After culture, the expression of CD1a, CD83 and HLA-DR as well as SI and ER were lower in CPDCs than those in UFDCs.</p><p><b>CONCLUSION</b>Though the biological properties of CPCB and CPDCs were injured after cryopreservation, they still had relatively complete basic function. Their recoveries rate were > 85%.</p>