Objective To establish a new rat model of biliary atresia by pure ethanol injection into the common bile duct.Methods A catheter was inserted and fixed in the common bile duct in male SD rats .Saline (8 rats) or pure ethanol (16 rats) was injected through the catheter ,respectively, and the biochemical and pathological changes in the rats were examined .Results SD rats in the experimental group were divided into a persistent injury and a restoration of liver dysfunction groups according to pathological and biochemical detection .In the persistent injury group , biochemical impair-ments were significantly higher at 8 weeks after ethanol injection than those in the control group and restoration group .Dis-tinct pathological changes in the liver were observed using HE , SMA, and Masson staining .Conclusions It is a reliable animal model of biliary atresia induced by injection of pure ethanol into the common bile duct in the rat .It will provide a useful tool in future studies of biliary atresia .

To separate and identify chemical signals which induce Thesium chinense haustorium formation, the components of T. chinense roots secretion collected with XAD-4 resin were detected by GC-MS. The effect of DMBQ as exogenous signals to induce haustorium formation in T. chinense was studied. Fifty-three compounds of 9 types had been detected, including hydrocarbons, esters, organic acids, ketones, alcohols, nitrogen containing compounds, phenolic acids, aldehyde and quinine. It is worth noting that the 2, 5-di-tert-butyl-1,4-benzoquinone has the core structure of 1,4-benzoquinone, which may play an important role in the parasitic relationship of Prunella vulgaris and T. chinense: DMBQ worked effectively on inducing haustoria, but induction effects vary widely in different concentrations. DMBQ with the concentration of 1 μmol x L(-1) showed the best effect of the inducing ability with a ratio of 110.52 when treated to induce haustoria.
Gas Chromatography-Mass Spectrometry ; Host-Parasite Interactions ; Magnoliopsida ; chemistry ; physiology ; Plant Roots ; chemistry ; physiology ; Prunella ; chemistry ; physiology

S1 nuclease (from Aspergillus oryzae) is a specific enzyme to degrade single stranded DNA or RNA molecules. It has been reported to be able to convert superhelical circular DNA molecules into open circle or linear forms under certain conditions, but this function has not been well explored. In order to use the action of S1 nuclease to linearize circular DNA and develop a novel way of cloning microcircular DNAs, the pUC19 was used to investigate the relationship between the linearization efficiency of S1 nuclease and the amount of enzyme used. By this way the optimal conditions for linearization of circular DNAs by S1 nuclease would be determined. 0.3u to 17u S1 nuclease per 100ng pUC19 DNA was added into a 25 microL system, respectively, to perform the reaction. The effectiveness of enzyme digestion was realized by electrophoresis in a 1.2% agarose gel. The results showed that along with the increase in enzyme amount from 0.3u to 17u a gradual decrease in the superhelical form, a gradual increase in the linear form and then in the circular form was obvious. The conversion from superhelical form to linear and circular form was directly related to the enzyme amount used. A higher proportion of linear DNA molecules was achieved by using 5 to 17u S1 nuclease per 100ng DNA. Besides, electrophoretic mobility of the S1 nuclease-linearized pUC19 was the same as that of the linear form produced by restriction enzyme digestion. According to the result of phiX174 digested by S1 nuclease it has been proposed that the enzyme cleaves first randomly on one site of one strand, thus converting the superhelical molecules into open circle form, and then on the same site of the complementary strand to produce the linear form. Therefore, the S1 nuclease-linearized DNA molecules are intact in the sense of their length and can be used for cloning. The plasmid-like DNA pC3 from cucumber mitochondria is a double stranded circular DNA molecule with about 550bp and the smallest known plasmid-like DNA in eukaryotic mitochondria. Many attempts have been made to linearize the molecule by using restriction enzymes but failed. Therefore, S1 nuclease was used to linearize pC3 based on the results obtained with pUC19. The linearized pC3 DNA molecules formed a very sharp band in a 2.5% agarose gel after electrophoresis. They were then recovered from the gel, added an "A" tail and ligated with T-vector. After transformation into E. coli JM109 cells, the positive clones were, screened by the blue-white selection. The insert was then cut using restriction enzymes EcoRI and Pst I. The result of electrophoresis shows that the electrophoretic mobility of the insert is just the same as that predicted. A 32 P-labled probe was synthesized using pC3 as the template and Southern blot analysis was carried out. The result shows that the inserted DNA is hybridized to the probe, which indicates that the cloned DNA fragment is from pC3. The sequence information of the insert shows that the plasmid-like DNA pC3 was 537bp in length. The nucleotide sequence was deposited in the GenBank (the accession number is AF522195).
Blotting, Southern ; Cloning, Molecular ; methods ; DNA, Circular ; genetics ; metabolism ; Fungal Proteins ; genetics ; metabolism ; Molecular Sequence Data ; Single-Strand Specific DNA and RNA Endonucleases ; genetics ; metabolism

OBJECTIVETo compare and evaluate the clinical value among transperitoneal, retroperitoneal laparoscopic adrenalectomy and open adrenalectomy for the treatment of adrenal tumours.

METHODSFrom January 1996 to June 2002, transperitoneal laparoscopic adrenalectomy (TLA) was performed for 19 cases, 17 cases were successful (Group A), retroperitoneal laparoscopic adrenalectomy (RLA) for 21 cases, 19 cases were successful (Group B), during the same period 41 patients with adrenal tumours under 6 cm in diameter underwent open adrenalectomy (Group C). The procedure was evaluated and compared in the respects of tumors weight, operation time, blood loss, recovery time of GI function, hospital stay, complications transferable rate and indications among three groups.

RESULTSAverage tumors weight was (13.86) g in group A, (15.66) g in group B and (18.03) g in group C; mean operating time: group A was 17.21 min longer than group B, group A. B was 48.53 min longer and 31.32 min longer han group C respectively, mean blood loss in group A. B was less diminished by 121.55 ml and 137.05 ml than group C, and the blood loss in group B was less diminished by 15.50 ml than group A; mean recovery of Gl function, group A and B was 1.87 day and 2.17 day earlier than group C respectively, group B was 0.3 day earlier than group A, mean hospital stay was 2.60 day and 3.87 day shorter than group C, group B was 1.27 day shorter, than group A; transferable rate of operation was 10.5% (group A) and 9.5% (group B). Postoperative complication rate was 10.5%, 14.3% and 9.7% in group A. B and C respectively.

CONCLUSIONSTLA and RLA were better than open method. On the hand of approach to adrenal gland disturbs to abdominal organ and postoperative recovery on RLA have some slight advantage.

Adrenal Gland Neoplasms ; surgery ; Adrenalectomy ; methods ; Adult ; Aged ; Female ; Humans ; Laparoscopy ; methods ; Laparotomy ; Male ; Middle Aged ; Postoperative Complications ; prevention & control ; Retroperitoneal Space ; Retrospective Studies ; Treatment Outcome

BACKGROUNDAn inflammatory response leading to organ dysfunction and failure continues to be a major problem after injury in many clinical conditions such as sepsis, severe burns, and trauma. It is increasingly recognized that atrial natriuretic peptide (ANP) possesses a broad range of biological activities, including effects on endothelial function and inflammation. A recent study has revealed that ANP exerts anti-inflammatory effects. In this study we tested the effects of human ANP (hANP) on lung injury in a model of oleic acid (OA)-induced acute lung injury (ALI) in rats.

METHODSRats were randomly assigned to three groups (n = 6 in each group). Rats in the control group received a 0.9% solution of NaCl (1 ml × kg(-1) × h(-1)) by continuous intravenous infusion, after 30 minutes a 0.9% solution of NaCl (1 ml/kg) was injected intravenously, and then the 0.9% NaCl infusion was restarted. Rats in the ALI group received a 0.9% NaCl solution (1 ml × kg(-1) × h(-1)) intravenous infusion, after 30 minutes OA was injected intravenously (0.1 ml/kg), and then the 0.9% NaCl infusion was restarted. Rats in the hANP-treated ALI group received a hANP (0.1 µg × kg(-1) × min(-1)) infusion, after 30 minutes OA was injected intravenously (0.1 ml/kg), and then the hANP infusion was restarted. The anti-inflammation effects of hANP were evaluated by histological examination and determination of serum cytokine levels.

RESULTSSerum interleukin (IL)-1β, IL-6, IL-10 and tumor necrosis factor (TNF) α were increased in the ALI group at six hours. The levels of all factors were significantly lower in the hANP treated rats (P < 0.005). Similarly, levels of IL-1β, IL-6, IL-10 and TNF-α were higher in the lung tissue in the ALI group at six hours. hANP treatment significantly reduced the levels of these factors in the lungs (P < 0.005). Histological examination revealed marked reduction in interstitial congestion, edema, and inflammation.

CONCLUSIONhANP can attenuate inflammation in an OA-induced lung injury in rat model.

Acute Lung Injury ; chemically induced ; drug therapy ; Animals ; Atrial Natriuretic Factor ; therapeutic use ; Disease Models, Animal ; Inflammation ; chemically induced ; drug therapy ; Male ; Oleic Acid ; toxicity ; Rats ; Rats, Wistar

BACKGROUNDPediatric patients are susceptible to lung injury that does not respond to traditional therapies. Total liquid ventilation has been developed as an alternative ventilatory strategy for severe lung injury. The aim of this study is to investigate the effect of total liquid ventilation on oleic acid (OA)-induced lung injury in piglets.

METHODSTwelve Chinese immature piglets were induced acute lung injury by OA. Twelve piglets were randomly treated with conventional gas ventilation (control group) or total liquid ventilation (study group) for 240 minutes. Samples for blood gas analysis were collected before, and at 60-minute intervals after OA-induced lung injury. The degree of lung injury was quantified by histologic examination. The inflammatory cells and the levels of IL-1β, IL-6, IL-10 and TNF-α in plasma, tissue and bronchoalveolar lavage were analyzed.

RESULTSNeutrophil and macrophage counts in bronchoalveolar lavage were significantly decreased in the study group (P < 0.05). The total lung injury score was also reduced in the study group (P < 0.05). The concentrations of IL-1β, IL-6, IL-10 and TNF-α in plasma, tissue and bronchoalveolar lavage were significantly reduced in the study group (P < 0.05).

CONCLUSIONSTotal liquid ventilation reduces biochemical and histologic OA-induced lung injury in piglets.

Acute Lung Injury ; chemically induced ; metabolism ; therapy ; Animals ; Interleukin-10 ; metabolism ; Interleukin-1beta ; metabolism ; Interleukin-6 ; metabolism ; Liquid Ventilation ; methods ; Oleic Acid ; toxicity ; Swine ; Tumor Necrosis Factor-alpha ; metabolism

BACKGROUNDThe interaction between activated microglia and T lymphocytes can yield abundant pro-inflammatory cytokines. Our previous study proved that thymus immune tolerance could alleviate the inflammatory response. This study aimed to investigate whether intrathymic injection of myelin basic protein (MBP) in mice could suppress the inflammatory response after co-culture of T lymphocytes and BV-2 microglia cells.

METHODSTotally, 72 male C57BL/6 mice were randomly assigned to three groups (n = 24 in each): Group A: intrathymic injection of 100 μl MBP (1 mg/ml); Group B: intrathymic injection of 100 μl phosphate-buffered saline (PBS); and Group C: sham operation group. Every eight mice in each group were sacrificed to obtain the spleen at postoperative days 3, 7, and 14, respectively. T lymphocytes those were extracted and purified from the spleens were then co-cultured with activated BV-2 microglia cells at a proportion of 1:2 in the medium containing MBP for 3 days. After identified the T lymphocytes by CD3, surface antigens of T lymphocytes (CD4, CD8, CD152, and CD154) and BV-2 microglia cells (CD45 and CD54) were detected by flow cytometry. The expressions of pro-inflammatory factors of BV-2 microglia cells (interleukin [IL]-1β, tumor necrosis factor-α [TNF-α], and inducible nitric oxide synthase [iNOS]) were detected by quantitative real-time polymerase chain reaction (PCR). One-way analysis of variance (ANOVA) and the least significant difference test were used for data analysis.

RESULTSThe levels of CD152 in Group A showed an upward trend from the 3rd to 7th day, with a downward trend from the 7th to 14th day (20.12 ± 0.71%, 30.71 ± 1.14%, 13.50 ± 0.71% at postoperative days 3, 7, and 14, respectively, P < 0.05). The levels of CD154 in Group A showed a downward trend from the 3rd to 7th day, with an upward trend from the 7th to 14th day (10.00 ± 0.23%, 5.28 ± 0.69%, 14.67 ± 2.71% at postoperative days 3, 7, and 14, respectively, P < 0.05). The ratio of CD4+/CD8 + T in Group A showed a downward trend from the 3rd to 7th day, with the minimum at postoperative day 7, then an upward trend from the 7th to 14th day (P < 0.05). Meanwhile, the levels of CD45 and CD54 in Group A were found as the same trend as the ratio of CD4+/CD8 + T (CD45: 83.39 ± 2.56%, 82.74 ± 2.09%, 87.56 ± 2.11%; CD54: 3.80 ± 0.24%, 0.94 ± 0.40%, 3.41 ± 0.33% at postoperative days 3, 7, and 14, respectively, P < 0.05). The expressions of TNF-α, IL-1β, and iNOS in Group A were significantly lower than those in Groups B and C, and the values at postoperative day 7 were the lowest compared with those at postoperative days 3 and 14 (P < 0.05). No significant difference was found between Groups B and C.

CONCLUSIONSIntrathymic injection of MBP could suppress the immune reaction that might reduce the secondary immune injury of brain tissue induced by an inflammatory response.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Antigens, Surface ; analysis ; Brain Injuries, Traumatic ; drug therapy ; CD4-CD8 Ratio ; Coculture Techniques ; Male ; Mice ; Mice, Inbred C57BL ; Microglia ; immunology ; Myelin Basic Protein ; administration & dosage ; pharmacology ; T-Lymphocytes ; immunology

OBJECTIVETo investigate the feasibility and therapeutic effect of acellular cadaveric dermis (ACD)-assisted immediate breast reconstruction.

METHODSFrom Sep. 2009 to May 2010, 10 cases received ACD-assisted immediate breast reconstruction. During the operation, the ACD was used to cover inferior and lateral portion of the implants in 2 cases and expanders in 8 cases.

RESULTSThe patients were followed up for an average period of 4 months with satisfactory breast appearance. The complications included infection in 2 cases and dehiscence in 2 cases. But no implant or expander was taken out.

CONCLUSIONSThe ACD-assisted immediate breast reconstruction is a technically simple procedure with minimal morbidity. Satisfactory clinical outcome can be achieved with appropriate candidates.

Acellular Dermis ; Adult ; Breast Implants ; Dermis ; transplantation ; Female ; Humans ; Mammaplasty ; methods ; Middle Aged ; Skin Transplantation ; Surgical Flaps ; Tissue Expansion Devices ; Treatment Outcome

OBJECTIVEAIM To establish a drug screening model based on transcriptional regulation of estrogen responsive element (ERE) and use it to screen compounds for discovering new ligands of estrogen receptor (ER) subtypes.

METHODA recombinant reporter vector pERE-TAL-SEAP was constructed by inserting a synthetic sequence composed of five tandem copies of EREs upstream of promoter of the reporter vector pTAL-SEAP. The pERE-TAL-SEAP and the internal control plasmid pCMV were transiently co-transfected into Hela cells expressing ER subtype or ER subtype, and the effects of pure ER agonists 17estradiol, phytoestrogen genistein and pure ER antagonist ICI182, 780 on reporter gene SEAP expression were observed.

RESULTIn the Hela cells expressing ER alpha or ER beta subtype, the expression of SEAP gene were induced in a dose dependent manner by 17-estrodiol with a maximal effect at approximately 10 nmol.L-1 and with EC50 of (80.58 +/- 8.51) pmol.L-1 and (103.90 +/- 5.29) pmol.L-1, respectively, so done by phytoestrogen genistein with a maximal effect at 1 mumol.L-1 and with EC50 of (10.86 +/- 0.75) nmol.L-1 and (39.38 +/- 2.26) nmol.L-1, respectively. The maximal level induced by estrodiol and genistein were about 7-14 fold higher than that of vehicle. The pure antiestrogen ICI182, 780 at concentration of 1 mumol.L-1 completely blocked the inductions of 17-estrodiol and genistein.

CONCLUSIONThe cellular drug screening model can be established by transfecting reporter vector pERE-TAL-SEAP in Hela cell lines expressing ER alpha or ER beta. The cell lines can be used to screen compounds with estrogenicity by testing SEAP activity in the culture media of cells growing in microtitier wells. The system should provide an efficient model for screening and analyzing the activity of large numbers of ligands of ER.

Drug Evaluation, Preclinical ; methods ; Estradiol ; pharmacology ; Estrogen Receptor alpha ; Estrogen Receptor beta ; Gene Expression Regulation ; drug effects ; Genes, Reporter ; Genistein ; pharmacology ; HeLa Cells ; Humans ; Ligands ; Promoter Regions, Genetic ; Receptors, Estrogen ; genetics ; Transfection

Objective To investigate the value of magnetic resonance imaging(MRI)T2-mapping in evaluating the activity of Graves ophthalmopathy(GO).Methods A total of 64 patients with GO in the Department of Endocrinology,the First Affiliated Hospital of Chongqing Medical University from July 2019 to January 2021 were collected.Simple random grouping was performed by computer,with 49 cases as observation subjects,and 15 patients for diagnostic test.According to clinical activity score(CAS),49 GO patients were divided into active group(CAS≥3 points,48 eyes)and inactive group(CAS<3 points,50 eyes).Normal control group(NC group)included 31 patients(62 eyes).All subjects underwent 3.0T orbital MRI T2-mapping.Measuring the T2 relaxation time(T2RT)of superior rectus,inferior rectus,medial rectus,and lateral rectus on five layers behind the eyeball on T2-mapping coronal images,and select the maximum value of T2RT in the five layers for each extraocular muscle to represent the T2RT of this extraocular muscle.Finally,select the maximum T2RT values of the four extraocular muscles,expressed as extraocular muscle maximum T2RT.Compare the differences of the above 5 indicators(superior rectus T2RT,inferior rectus T2RT,medial rectus T2RT,lateral rectus T2RT,extraocular muscle maximum T2RT)between active group,inactive group and NC group.ROC curve was used to analyze the diagnostic value of the above 5 indicators for GO activity assessment,and the diagnostic threshold was obtained.Then,another 15 GO patients were performed for diagnostic tests evaluation to determine the indicators of high diagnostic efficacy and the threshold of diagnostic activity.Results The T2RT of all extraocular muscles in active group were significantly higher than those in inactive group and NC group,the difference was statistically significant(P<0.001).The threshold value of the five indicators were obtained by ROC curve analysis.The maximum T2RT cut-off values of superior rectus muscle,inferior rectus muscle,medial rectus muscle,lateral rectus muscle and extraocular muscles for judging activity were 80.200 ms,97.045 ms,94.355 ms,85.750 ms and 101.385 ms respectively.Another 15 GO patients were performed for diagnostic tests,the indexes with relatively high sensitivity,specificity,positive predictive value and negative predictive value were inferior rectus T2RT and extraocular muscle maximum T2RT,the cut-off values of GO activity were 97.045 ms and 101.385 ms,respectively;the sensitivity were 91.7％and 93.8％,respectively;the specificity all were 80.0％.Conclusions MRI T2-mapping sequence has a good value in assessment of GO activity.The inferior rectus T2RT and extraocular muscle maximum T2RT can be choosed to evaluate the activity of GO.