Lipids of Thamnidium elegans,Mortierella ramanninace,Rhizopus arrhizus,Pythium irregulare and Rhodotorulla aurantiaca were extracted by Soxhlet extraction,supercritical-CO 2 fluid extraction,acid-heating extraction and organic solvent extraction,respectively.Four extraction methods were evaluated on sample treatment,minimum sample quantity,requirements of apparatus,ability of treating sample and content of lipid.The components of fatty acids were analysed by gas chromatography.Soxhlet extraction can acquired maximum lipid content,but it took the most time.Supercritical-CO 2 fluid extraction and acid-heating extraction has a same lipid content which was lower than that of Soxhlet extraction.Acid-heating extraction was the most handy,and its ability to treat sample in a hour was the most powerful.Organic solvent extraction was less efficient.Acid-heating extraction was a simple and efficient method of fungi lipid extraction fitting to breed mutant strains that highly producting lipid and polyunsaturated fatty acids.

OBJECTIVETo investigate the effects of pregnancy malnutrition on the occurrence of insulin resistance (IR) in rat offspring during adult stage and to find out the relationship between TNF-alpha and IR; and to find out a reasonable early nutritional intervention measure for the prevention of IR, through giving different diets to offspring.

METHODSAn IUGR model was built by maternal nutrition restriction. 80 newborn IUGR female pups were randomly divided into 4 groups, the mother rats were given the following diet respectively for 3 weeks after delivery, pups were fed by mother milk: (1) The IUGR (intrauterine growth retardation) rat model was used and the animals were divided into: IUGR control group (group S/N) fed with normal diet, (2) IUGR high-caloric diet group (group A), (3) IUGR high-protein and high-caloric diet group (group B) and (4) IUGR high-protein isocaloric diet group (group C). Each group had 20 pups and another 20 normal female pups were fed with normal diet as the normal control group (group C/N). All pups were weaned at the 4th week of age and fed with normal diet till the end of the experiment. At the 12th week (adulthood) and 48th week (senility) of life, body weight and length, the fasting blood glucose, insulin concentration, TNF-alpha of adipose tissue and body weight were measured. Body mass index (BMI), ISI (insulin sensitive index), IRI (insulin resistant index) and HBCI (beta cell insulin excretion index) and their correlation to TNF-alpha were calculated.

RESULTSAt 12th week and 48th week of life, the insulin sensitivity of IUGR model group was significantly lower than group C/N, although there was no significant difference of body weight between these two groups. TNF-alpha was negatively correlated with ISI, positively correlated with IRI and no relation to HBCI. Group A and B was fatter and developed more severe IR. There were no significant differences in ISI, IRI, HBCI and TNF-alpha between group C and group C/N.

CONCLUSIONSIUGR offspring of pregnancy malnutrition mother rats showed IR at the age of 12th week. TNF-alpha was closely related to the occurrence of IR in IUGR pups. IUGR pups fed with high caloric diet or high protein and caloric diet at the early postnatal period amplified the metabolic abnormality. The high protein isocaloric diet is effective early nutritional intervention measure for the prevention of occurrence of IR at adulthood.

Animals ; Animals, Newborn ; growth & development ; Body Weight ; drug effects ; Dietary Proteins ; pharmacology ; Female ; Fetal Growth Retardation ; blood ; etiology ; Insulin Resistance ; physiology ; Malnutrition ; physiopathology ; Pregnancy ; Pregnancy Complications ; Prenatal Exposure Delayed Effects ; Rats ; Rats, Sprague-Dawley ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo study the change in intestinal barrier and organ functions of burned dog after enteral administration of hypertonic electrolyte glucose solution (HEGS) in shock stage.

METHODSTwenty-four Beagle dogs inflicted with 35% TBSA full-thickness burn were divided into no-fluid group (NF), intravenous infusion with isotonic electrolyte glucose solution (IEGS) group (II group), enteral infusion with IEGS group (EI), and enteral infusion with HEGS group (EH) according to the random number table, with 6 dogs in each group. Saline, containing 50 g/L glucose, was intravenously or enterally infused into dogs in II group and EI group respectively 0.5 hour post injury (PIH) for resuscitation. Total infusion volume within PIH 24 was 4 mL x kg(-1) x %TBSA(-1) (half of the total volume was infused in the first 8 hours in a constant speed, the other half volume was infused in the rest 16 hours in a constant speed). HEGS, containing 18 g/L NaCl and 50 g/L glucose, was enterally infused into dogs in EH group. Total infusion volume within PIH 24 was 2 mL x kg(-1) x %TBSA(-1), with the same infusion speed as that in II and EI groups. Liver and kidney function indexes [activity of ALT and CK-MB, expression levels of creatinine and blood urea nitrogen (BUN) in serum], activity of diamine oxidase (DAO), and activity of Na(+)-K(+)-ATPase in intestinal mucosa at PIH 24 were determined.

RESULTSALT activity in each group was close to one another. Serum levels of creatinine and BUN in II, EI, and EH groups were significantly lower than those in NF group. CK-MB activity obviously increased at PIH 2 in every group. CK-MB activity in EH group at PIH 2 to 8 was respectively lower than that in NF and II groups. DAO activity in serum in II, EI, and EH groups decreased since PIH 4 or PIH 6, respectively from (3.9 + or - 0.6) U/L to (3.6 + or - 0.5) U/L, (4.8 + or - 0.4) U/L to (2.8 + or - 0.8) U/L, (6.4 + or - 1.8) U/L to (3.5 + or - 0.8) U/L, all were significantly lower than those in NF group [from (12.5 + or - 0.4) U/L to (9.7 + or - 1.1) U/L, comparison between EH group and NF group, t value at PIH 4, 6, 8, 24 was respectively 10.25, 12.44, 17.99, 16.21, P values all below 0.05]. The order of Na(+)-k(+)-ATPase activity in intestinal mucosa at PIH 24 in each group from high to low was II group, EH group, EI group, and NF group (comparison between former 3 groups and NF group, t value was respectively 10.09, 4.96, 8.32, F value was 26.79, P values all below 0.05).

CONCLUSIONSHEGS does not cause significant harm to the barrier function of intestinal mucosa of shock dog after burn. Compared with NF, HEGS can significantly improve functions of heart, liver, and kidney, and it can achieve the same resuscitation effect as enteral or intravenous infusion of IEGS with only half of the solution volume.

Animals ; Burns ; drug therapy ; metabolism ; physiopathology ; Disease Models, Animal ; Dogs ; Fluid Therapy ; Glucose Solution, Hypertonic ; administration & dosage ; therapeutic use ; Heart ; physiopathology ; Intestinal Mucosa ; metabolism ; Intestine, Small ; physiopathology ; Kidney ; physiopathology ; Liver ; physiopathology

OBJECTIVETo investigate the effects of different anesthesia methods on immune surveillance and tumor metastasis in tumor-bearing rats.

METHODSSeventy-two Fischer 344 rats were randomly assigned into 3 equal groups and anesthetized for 1 h with ketamine (group K), propofol (group P), or neuraxial block (group B). All the rats were subjected to laparotomy followed by intravenous injection of MADB106 tumor cells, and 24 h after the injection, the number and activity of circulating CD3(+), CD4(+), CD8(+), and D4(+)/CD8(+) lymphocyte subsets and NK cellèCD161a(+)éwere assessed. Three weeks later, the lung metastases were counted.

RESULTSCompared with those in group B, the numbers of CD3(+), CD4(+), CD8(+), and CD161a(+) lymphocytes and the activity of circulating NK cells were significantly reduced, and the lung metastases of MADB106 increased significantly in groups K and P (P<0.05 or 0.01 ). The activity of immune surveillance in group K was significantly lower than that in group P except for CD8(+) cells, and the tumor metastases in group K increased significantly in comparison with those in group P (P<0.05 or 0.01).

CONCLUSIONNeuraxial block provides protection of the activity of immune surveillance and reduces tumor metastases in tumor-bearing rats compared with general anesthesia.

Anesthesia, Epidural ; adverse effects ; Anesthesia, General ; adverse effects ; Animals ; Breast Neoplasms ; immunology ; surgery ; Female ; Immunologic Surveillance ; immunology ; Ketamine ; pharmacology ; Lung Neoplasms ; immunology ; secondary ; Male ; Neoplasm Metastasis ; Neoplasm Transplantation ; Nerve Block ; Propofol ; pharmacology ; Random Allocation ; Rats ; Rats, Inbred F344

OBJECTIVE: To evaluate the relationship between serum levels of soluble Fas(sFas), soluble Fas li gand(sFasL), soluble IL-2 receptor(sIL-2R) and coronary heart disease (CHD). METHODS: With enzyme-linked immunosorbent assay (ELISA) tests, sFas, sFasL and sIL-2R were measured in the sera from 30 patients with CHD and 26 subjects without CHD as controls. RESTULTS: Mean level of sFas was significantly higher in patients with CHD than in controls [(1 583.41+/-174.46)ng/L compared with (1 374.55+/-142.42)ng/L, P<0.01]. Compared with the controls, the mean level of sIL-2R was significantly higher in patients with CHD [(944.50+/-395.59)ng/L compared with (652.45+/-163.36)ng/L P<0.01]. Moreover, in patients with CHD sFas and sIL-2R were positively correlated (r=0.418 P<0.05). Whereas no such difference was found between both groups in sFasL (P<0.05). CONCLUSION: High levels of serum sFas and sIL-2R were associated with CHD, and elevation of sFas may inhibit apoptosis in activated T cells, leading to coronary events.

OBJECTIVETo study the resuscitative effect of hypertonic electrolyte glucose solution (HEGS) in enteral resuscitation of burn shock.

METHODSEighteen Beagle dogs with 35% TBSA full-thickness flame injury were used in this study. They were randomized to a control group (no-fluid resuscitation, N group), a HEGS resuscitation group (H group) or an isotonic electrolyte glucose solution (IEGS) resuscitation group (I group). The solution enterally was given for resuscitation from half an hour after burn. The volumes and rates of fluid infusion in the H group were basically in accordance with 2 ml/(kg x 1%TBSA), those in the I group were basically in accordance with parkland formula [4 ml/(kg x 1%TBSA)]. The haemodynamic parameters, global end-diastolic volume index, plasma volume, osmotic pressure of plasma, intestinal absorptive rates of water and Na(+), and intestine mucosa blood flow were continuously assessed.

RESULTSThe cardiac output index, global end-diastolic volume index, plasma volume and intestine blood mucosa flow reduced markedly after burn in the three groups, and then gradually returned from 2 h after burn in two resuscitation groups, which were higher than that in the N group (P < 0.05). The activities of diamine oxidase in plasma in the two resuscitation groups were higher than that in N group (P < 0.05). The intestinal absorption rates of water and Na(+) reduced markedly after burn in two resuscitation groups with the lowest levels, and then returned from 6 h after burn. The rates of water in H group were lower than that in I group (P < 0.05); the rates of Na(+) in H group were higher than in I group (P < 0.05).

CONCLUSIONThe results indicated that 35%TBSA III degrees burn-injury dogs be resuscitated effectively with 1.8% hypertonic electrolyte-glucose solution by enteral, which 1/2 volume of an isotonic electrolyte glucose solution.

Animals ; Burns ; therapy ; Disease Models, Animal ; Dogs ; Enteral Nutrition ; Fluid Therapy ; methods ; Glucose Solution, Hypertonic ; administration & dosage ; therapeutic use ; Random Allocation ; Resuscitation ; methods ; Saline Solution, Hypertonic ; administration & dosage ; therapeutic use

OBJECTIVETo investigate epidermal growth factor receptor (EGFR) gene mutations in exons 19 and 21 of patients with non-small cell lung cancer (NSCLC) and to analyze the relationship of EGFR mutations with clinicopathological features and prognosis.

METHODSThe EGFR gene exons 19 and 21 of paraffin-embedded tumor tissue were amplified by PCR, followed by direct sequencing in 282 surgically-removed specimens of NSCLC. The relationship of EGFR gene mutations in NSCLC with clinicopathological features and prognosis were analyzed.

RESULTSEGFR mutations were detected in 120 of 282 (42.6%) patients with NSCLC. There were 61 cases of the mutations in exon 19 and 66 cases of the mutations in exon 21, including 7 cases of the mutations both in exons 19 and 21. Mutations were more frequently observed in women (55.2%, 53/96) than in men (36.0%, 67/186), in 51 to 60-years-old (51.3%, 39/76) than ≤50-years-old (30.4%, 21/69) and >60-years-old (43.8%, 60/137), in non-smokers (54.3%, 69/127) than smokers (32.9%, 51/155), there was negative correlation of EGFR mutations with smoking status (P=0.000, rs=-0.216). EGFR mutations were more frequently observed in adenocarcinomas (47.8%, 64/134), bronchiolo-alveolar carcinomas (73.0%, 27/37), adenosquamous carcinomas (7/9) than squamous cell carcinomas (23.6%, 17/72) and other types (16.7%, 5/30). The EGFR mutation rate in the well differentiated, the middle differentiated, the poorly differentiated and the undifferentiated was 55.7% (68/122), 50.8% (30/59), 22.7% (17/75), 19.2% (5/26) respectively, the incidences of EGFR mutations decreased with the degrading of differentiation, there was positive correlation of EGFR mutations with differentiation of lung cancer (P=0.000, rs=0.296). The patients with EGFR mutations had better prognosis than those with wild-type EGFR (P=0.027). There was no association of EGFR mutations with clinical TNM stage.

CONCLUSIONSEGFR mutations occur frequently in females, non-smokers and adenocarcinomas, bronchioloalveolar carcinomas, and adenosquamous carcinomas. The patients with EGFR mutations have better prognosis. The results may offer a practical approach to select the patients who may benefit from anti-EGFR target therapy.

Adenocarcinoma ; genetics ; Adenocarcinoma, Bronchiolo-Alveolar ; genetics ; Adult ; Age Factors ; Aged ; Aged, 80 and over ; Carcinoma, Adenosquamous ; genetics ; Carcinoma, Non-Small-Cell Lung ; genetics ; pathology ; Exons ; Female ; Gene Amplification ; Genes, erbB-1 ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Lung Neoplasms ; genetics ; pathology ; Male ; Middle Aged ; Mutation ; Mutation Rate ; Polymerase Chain Reaction ; methods ; Prognosis ; Receptor, Epidermal Growth Factor ; genetics ; Sequence Analysis, DNA ; methods ; Sex Factors ; Smoking ; Survival Rate

OBJECTIVETo investigate the expression of T-bet mRNA in peripheral blood mononuclear cells (PBMC) as well as its relations with total IgE (TIgE), eosinophil cationic protein (ECP) in serum and nasal allergic symptoms in patients with allergic rhinitis (AR).

METHODSThe allergen, TIgE and ECP in serum of patients with AR were detected by Unicap CAP system. Blood sample was taken from 8 healthy individuals and 22 patients with allergic rhinitis. PBMC was isolated by density gradient centrifugation and one part of them was cultured with 50 microg/ml mite allergen. PBMC was subjected to analysis of T-bet mRNA expression using semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR).

RESULTSThe ratio of T-bet to beta-actin mRNA levels was 0.381 +/- 0.099 in patients and 0.750 +/- 0.067 in normal individuals, the difference was significantly (P <0.01). The expression intensity of T-bet mRNA had no relation to varying severity of allergic symptoms and concentration of ECP and the correlation coefficient was 0.187 and -0.165 (all P > 0.05). However, there was an inverse correlation between expression intensity of T-bet mRNA and TIgE concentration (r = -0.525, P < 0.05). Mean mRNA level (x +/- s) of T-bet expression before and after being stimulated by allergen was 0.381 +/- 0.099 and 0.365 +/- 0.104 respectively, which indicated no significant differences (P > 0.05).

CONCLUSIONSAmong allergic patients whose allergen was mite, there was a down-regulated expression of T-bet mRNA, which had no relation to ECP concentration and allergic symptoms, but was one of important links in mechanisms of imbalance of Th1/Th2 in AR. There was no effect of specific allergen on T-bet mRNA in patients with AR T-bet was one of indirect factors that affected the level of IgE.

Adolescent ; Adult ; Case-Control Studies ; Child ; Child, Preschool ; Eosinophil Cationic Protein ; blood ; Female ; Humans ; Immunoglobulin E ; blood ; Male ; Rhinitis, Allergic, Seasonal ; blood ; T-Box Domain Proteins ; blood ; Young Adult

OBJECTIVETo observe the effects of cryopreserved olfactory ensheathing cells (OECs) transplantation on axonal regeneration and functional recovery following spinal cord injury in adult rats.

METHODSTwenty-four rats were divided into experimental and control groups, each group having 12 rats. The spinal cord injury was established by transecting the spinal cord at T10 level with microsurgery scissors. OECs were purified from SD rat olfactory bulb and cultured in DMEM (Dulbecco's minimum essential medium) and cryopreserved (-120 degree) for two weeks. OECs suspension [(1-1.4)x10(5)/ul] was transplanted into transected spinal cord, while the DMEM solution was injected instead in the control group. At 6 and 12 weeks after transplantation, the rats were evaluated with climbing test and MEP (moter evoked potentials) monitoring. The samples of spinal cord were procured and studied with histological and immunohistochemical stainings.

RESULTSAt 6 weeks after transplantation, all of the rats in both transplanted and control groups were paraplegic, and MEPs could not be recorded. Morphology of transplanted OECs was normal, and OECs were interfused with host well. Axons could regrow into gap tissue between the spinal cords. Both OECs and regrown axons were immunoreactive for MBP. No regrown axons were found in the control group. At 12 weeks after transplantation, 2 rats (2/7) had lower extremities muscle contraction, 2 rats (2/7) had hip and/or knee active movement, and MEP of 5 rats (5/7) could be recorded in the calf in the transplantation group. None of the rats (7/7) in the control group had functional improvement, and none had MEPs recorded. In the transplanted group, histological and immunohistochemical methods showed the number of transplanted OECs reduced and some regrown axons had reached the end of transected spinal cord. However, no regrown axons could be seen except scar formation in the control group.

CONCLUSIONSCryopreserved OECs could integrated with the host and promote regrowing axons across the transected spinal cord ends.

Animals ; Axons ; pathology ; physiology ; Cell Transplantation ; Cryopreservation ; Immunohistochemistry ; Male ; Nerve Regeneration ; Olfactory Nerve ; cytology ; Rats ; Rats, Sprague-Dawley ; Spinal Cord Injuries ; physiopathology ; surgery

Osteosarcoma is the most common primary malignant bone cancer in children and adolescents. Emerging evidence has suggested that the capability of a tumor to grow is driven by a small subset of cells within a tumor, termed cancer stem cells (CSCs). Although several methods have been explored to identify or enrich CSCs in osteosarcoma, these methods sometimes seem impractical, and chemotherapy enrichment for CSCs in osteosarcoma is rarely investigated. In the present study, we found that short exposure to chemotherapy could change the morphology of osteosarcoma cells and increase sarcosphere formation in vitro, as well as increase tumor formation in vivo. Furthermore, methotrexate (MTX)-resistant U2OS/MTX300 osteosarcoma cells were larger in size and grew much more tightly than parental U2OS cells. More importantly, U2OS/MTX300 cells possessed a higher potential to generate sarcospheres in serum-free conditions compared to parental U2OS cells. Also, U2OS/MTX300 cells exhibited the side population (SP) phenotype and expressed CSC surface markers CD117 and Stro-1. Notably, U2OS/MTX300 cells showed a substantially higher tumorigenicity in nude mice relative to U2OS cells. Therefore, we conclude that chemotherapy enrichment is a feasible and practical way to enrich osteosarcoma stem cells.
Animals ; Antigens, Surface ; metabolism ; Antimetabolites, Antineoplastic ; pharmacology ; Bone Neoplasms ; metabolism ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Drug Resistance, Neoplasm ; Humans ; Methotrexate ; pharmacology ; Mice ; Mice, Nude ; Neoplasm Transplantation ; Neoplastic Stem Cells ; drug effects ; pathology ; Osteosarcoma ; metabolism ; pathology ; Phenotype ; Proto-Oncogene Proteins c-kit ; metabolism