Objective To observe the pathological changes of tissues of the injured bile duct, and to provide theoretical basis for bile duct repair. Methods Dog models of obstructive biliary injury were established.Sixty dogs were equally divided into five groups according to the duration of biliary obstruction: biliary obstruction for 5 days (BDL5 group), 10 days (BDL10 group), 15 days (BDL15 group), 20 days (BDL20 group) and 30 days (BDL30 group). The morphological and pathological changes of bile duct and local tissues were observed, and biliary-enteric Roux-en-Y anastomosis was applied to repair the injured bile duct and postoperative complications were observed. All data were analyzed by LSD test, independent sample t test, one-way analysis of variance and chi-square test. Results Proximal bile duct rapidly expanded as the pressure increased in the early stage, and the bile duct expanded to ( 15.6 ± 1.8)mm in the BDL10 group. The expansion rate decreased in the later stage,and the bile duct expanded to (18.9 ± 1.9)mm in the B DL15 group. Acute inflammation was observed in injured local tissues. The acute inflammation was severe in the BDL5 group with white blood cell count of 54 ± 6, and the acute inflammation was relatively mild in the BDL15 group with white blood cell count of 42 ± 7. There was a significant difference between the BDL5 group and BDL15 group in the degree of acute inflammation (t =4. 688,P ＜ 0. 05). The content of the collagen was increased in the injured bile duct as time passed by. Bile duct repair was successfully performed on 57 dogs. Ten dogs ( three in the BDL5 group, four in the BDL10 group, one in the BDL15 group, one in the BDL20 group and one in the BDL30 group) died of bile leakage after the operation. The incidences of bile leakage was 30% (7/23) within 10 days and 9% (3/34) beyond 10 days, with a significant difference between the two groups (x2 =4.429, P＜0.05). Conclusion Ten days after obstructive biliary injury,an obvious reduction of bile duct expansion and edema of the bile duct is observed, the difficulty of the operation is reduced and the incidence of bile leakage is low, so 10 days after the incidence of obstructive biliary injury is the proper timing for the surgical repair.

Objectives To evaluate the effect of probiotics enteral nutrition therapy on the inflammatory reaction and immune function in acute necrotizing pancreatits(ANP)in rats.Methods 100 SD rats were divided randomly into control group(C group,n=20),ANP gruop,enteral nutrition group (EN group,n=20),probiotics group(P group,n=20)and enteral nutrition plus probiotics group(PEN group,n=20),ANP were induced by retrograde injection of 5% sodium tautocholate(1.5ml/kg)into the biliary and pancreatic duct.The nasojejunal tube was placed via gastric route.The rats were sacrificed at 12h,24h,48h and 72h after ANP modeling.Serum MIF and amylase levels,as well as CD_3~+,CD_4~+,CD_8~+were measured and MIF was determined by immunohistochemistry and the histopathological changes in pancreatic tissue were observed.Results Serum MIF and amvlase levels were all increase when compared with that in C group.72h after ANP modeling,srum MIF leves in EN,P,PEN groups were(117.59±1.86)μg/L,(108.39±1.99) μg/L and (95.33±1.96) μg/L,respectivdy;serum amylase levels were (2799±161)U/L,(2482±140)U/L and (2146±572 )U/L,respectively;the values in P and PEN groups were significantly lower than that of EN group(P＜0.05),the values in PEN groups were significantly lower than that of P group (P＜0.05 ).After ANP induction,CD_4~+/CD_8~+ values in EN,P and PEN groups were 0.93±0.12,1.31±0.13,1.51±0.10,respectively;the values in P and PEN groups were significantly higher than that of EN group(P＜0.05 ),the values in PEN groups were significantly higher than that of P group (P＜0.05 ).MIF positive rates in C,ANP,EN,P and PEN group were 45%,96%,95%,65% and 60% ,respectively;the values in P and PEN groups were significantly lower than those of ANP and EN group (P＜0.05 ).At 72 h,the damage of pancreatic tissue was more severe in EN and P groups than that in PEN group.Conclusions Probioties enteral nutrition could effectively regulate inflammation mediator and cytokines in rats with ANP,and enhance immune function.

In order to further investigate the mechanisms of action of berberine (Ber), we assessed the effects of Ber on the mRNA expression of nitric oxide synthases (NOS) in rat corpus cavernosum. After incubation with Ber for 1 or 3 h respectively, the levels of NOS mRNA were examined by reverse transcription polymerase chain reaction (RT-PCR). Our results showed that there were iNOS and eNOS mRNA expressions in rat corpus cavernosum. Ber enhanced eNOS mRNA expression in rat penis, but exhibited no effect on the expression of iNOS mRNA (P > 0.05). The present study indicated that the relaxation of Ber involved the NO-cGMP signal transduction pathway. The enhancing effect of Ber on eNOS mRNA expression might associated with its relaxation of corpus cavernosum.
Berberine/*pharmacology ; Connective Tissue/physiopathology ; Nitric Oxide Synthase/*biosynthesis ; Nitric Oxide Synthase/genetics ; Nitric Oxide Synthase Type I/biosynthesis ; Nitric Oxide Synthase Type I/genetics ; Nitric Oxide Synthase Type III/biosynthesis ; Nitric Oxide Synthase Type III/genetics ; Penile Erection/*physiology ; Penis/*metabolism ; Penis/physiology ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics

To further investigate the mechanisms of action of icariin (ICA), we assessed the effects of ICA on the in vitro formation of cGMP and cAMP in isolated rabbit corpus cavernosum. Isolated segments of rabbit corpus cavernosum were exposed to increasing concentrations of ICA and the dose-dependent accumulation of cGMP and cAMP was determined in the tissues samples by means of 125I radioimmunoassay. Responses of the isolated tissues preparations to ICA were compared with those obtained with the reference compounds sildenafil (Sild). Furthermore, the effects of ICA on the mRNA expression of specific cGMP-binding phosphodiesterase type V (PDE5) in rat penis were also observed. After incubation with ICA for 6 h or 14 h respectively, the levels of PDE5 mRNA were examined by reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that ICA increased cGMP concentrations directly (P < 0.05), but there was no significant effect on cAMP concentrations (P > 0.05). In the presence of sodium nitroprusside (SNP), a stimulatory agent of cGMP, both ICA and Sild increased cGMP concentrations with increasing dose (P < 0.01). Their EC50 was 4.62 (ICA) and 0.42 (Sild) micromol/L respectively. Under the same condition, ICA and Sild unaltered cAMP level significantly (P > 0.05). There were PDE5A1 and PDE5A2 mRNA expressions in rat corpus cavernosum with PDE5A2 being the dominant isoform. ICA could obviously inhibit these two isoforms mRNA expression in rat penis, and decrease PDE5A1 more pronouncedly (P < 0.01). The present study indicated that the aphrodisiac mechanisms of icariin involved the NO-cGMP signal transduction pathway, with increasing cGMP levels in the corpus cavernosum smooth muscle. The inhibitory effect of icariin on PDE5 mRNA expression, especially on PDE5A1, might account for its molecular mechanisms for its long-term activity.

Animals ; Dendrobium ; chemistry ; Fatigue ; drug therapy ; Mice ; Plant Extracts ; pharmacology

In order to further investigate the mechanisms of action of berberine (Ber), we assessed the effects of Ber on the mRNA expression of nitric oxide synthases (NOS) in rat corpus cavernosum. After incubation with Ber for 1 or 3 h respectively, the levels of NOS mRNA were examined by reverse transcription polymerase chain reaction (RT-PCR). Our results showed that there were iNOS and eNOS mRNA expressions in rat corpus cavernosum. Ber enhanced eNOS mRNA expression in rat penis, but exhibited no effect on the expression of iNOS mRNA (P＞0.05). The present study indicated that the relaxation of Ber involved the NO-cGMP signal thansduction pathway. The enhancing effect of Ber on eNOS mRNA expression might associated with its relaxation of corpus cavernosum.

Obej ctive Knockout mice are widely used in the studies of joint diseases .This article investigated the effects of joint processing methods , collagenase types ,and collagenase digestion time on the number of primary fibroblast -like synoviocytes (FLSs) obtained from mice. Methods The hind legs of 6 of the 12 male mice were cut open from the hip joints , but not those of the other 6.FLSs were isolated using the type-Ⅳcollagenase digestion method and purified by differential digestion .Cell morphology was observed under the inverted microscope .The type, viability, and purity of the cells were determined by flow cytometry . Rse ults Significantly fewer FLSs were obtained from the mice with the hind legs cut open ( 19 133 ±115 ) than from those without (24 933 ± 503) (P<0.05).The numbers of FLSs collected from the cell suspension at 1, 2, 3, 4, 5,6 , and 7 hours after digestion were 700 ±300 , 600 ±100 , 15 200 ±900 , 5100 ±800 ,2700 ±300 , 900 ±200, and 300 ±100, respectively, the highest at 3 hours. There were statistically significant differences in the total number of FLSs obtained by type-Ⅳ and type-Ⅱ collagenase digestions (24900 ±500v s 18 100 ±400, P<0.05). Conclusion For in virt o culture of primary mouse FLSs, it is recommended that the hip joints be not cut open, and type-Ⅳcollagenase be used with cell sus-pension at 2-6hours after digestion .

Obej ctive Abnormal proliferation of T cells plays an important role in the development of autoimmune diseases. The article aimed to study the inhibitory effect of small molecule compound BD691 on T cell proliferation and its mechanism. Methods Human peripheral blood T-lymphocytes were isolated and purified by the immunomagnetic microbeads,then T cells were ac-tivated with anti-CD3/CD28 mAbs or alloantigen.The inhibitory effect of BD691 on activated T cell proliferation, the cytotoxic effect BD891 on resting T cells and the expression of activated T cells marker CD25 were measured by flow cytometry.Furthermore, ELISA was used to detect the secretion of cytokines associated with T cell differentiation. Results BD691 significantly inhibited the prolif-eration of T cells being stimulated by anti-CD3/CD28 mAb or alloantigen in a dose-dependent manner, and IC50 values are (8.5 ± 1.5)μmol/L and (7.2 ±1.3)μmol/L, respectively.However, BD691 had no obvious cytotoxic effects on resting T cells and periph-eral blood mononuclear cells, even at a high concentration ( up to 100μmol/L) .In T cells which were not activated by anti-CD3/CD28 mAb, the percentage of CD25+T cells is only 1.6%of the total cells, while the number increased to 68% after activating treatment.Mean-while, in T cells which were activated by 0, 3.3, 10, 30μmol/L BD691, no obvious change of CD25 expression were observed, while immunosuppressant FK506 (0.1μmol/L) significantly decreased the expression of CD25 +T cells (14.9%).In unactivated T cells, 95.6%cells were at G0/G1 phase, while after activation, the percentage of cells at G0/G1 phase reduced to 57.7%.In addition, BD691 inhibited the secretion of IFN-γ, IL-6 and IL-17 in activated T cells, but had no effects on the secretion of IL-2, IL-4 and IL-10. Co nclusion BD691 exerts no effects on T cell activation, but it inhibits T cell proliferation by inducing T cell cycling arrest at G0/G1 phase.Moreover, BD691 inhibits the secretion of key cytokines (such as IFN-γ, IL-6, IL-17) closely related to the differ-entiation of Th1 and Th17 cells.The results suggest that BD 691 is a potential lead compound to develop a new immunosuppressant for the inhibition of abnormal proliferation and differentiation of T cells.

Objective To explore the optimal timing of operation for experimental obstructive jaundice in a dog model. Method A dog model of bile duct stricture (BDS) was established. Dogs were divided into (n = 12 in each group) 6 groups, ie control, BDS days 5, 10, 15, 20, and 30. In each dog,the morphology and local histopathology of the bile duct, and the liver function in different periods were observed. At the time of surgery biopsy was taken and Roux-en-Y hepaticojejunostomy performed. Surgical complications and survival were evaluated. Result After bile duct obstruction, the proximal bile duct dilated continuously. The diameter of bile duct was 15.6 ± 1.7 mm at the 10th day. The injury bile ductshowed the acute inflammation change. In the early time (in 10 days), inflammatory cells increased in the tissues, mucous edema aggravated, the wall was edematous thickening, it was most severe ( WBC counting 54 ±6) in the 5th day. In the later period (10 -30 days), inflammatory cells reduced, bile duct wall became fibrosis, which was most obvious in the 15th day (42 ± 7 vs 54 ± 6, P ＜ 0.05 ). During the development of jaundice, serum bilirubin reached the highest level in the early period ( BDS days 5 group),then presented a platform time, and then rised extremely at the last stage of the experiment ( BDS day 30 group) . Changes of ALT and AST paralleled that of bilirubin before the 20th day of obstruction and then plummeted. BDS was repaired successfully in 57 dogs. Ten dogs died postoperatively due to bile leakage within 10 days, 3 dogs in BDS days 5 group (3/11), 4 in BDS days 10 group (4/12), one each in other groups. Postoperatively 13 BDS dogs died of malnutrition and organ failure within 3 months, including one each in days 5 and days 10 group, two each in days 15 and days 20 group, and 7 in days 30 group (P＜0. 05). Conclusion Considering the changes of morphology, physical function and result of follow up.The period between 10 and 20 days after acute bile duct injury is optimal for surgical repair.

ObjectiveTo explore the histopathological changes of bile duct,liver and local tissue for injurious biliary stricture(IBS). MethodTo observe the morphological and pathological changes of bile duct, local tissue and liver in different periods with dogs as the established animal model for IBS. ResultBile duct obstruction due to injury can expand the proximal bile duct up to 18.91 ±1.85 mm as the pressure goes up. Damage to local tissue triggers acute inflammation. In early injury phase (within 10 d), inflammatory cell infiltration and proliferation appears on the wall of the duct with increased mucosal edema as well as thickening of the biliary ductile wall. In the late injury phase (15 d), the degree of infiltration of inflammatory cells, edema and mucosal thickness were reduced whereas fibroblast and collagen tissue were proliferated extensively. The wall of biliary duct also becomes fibrotic and thickens. Quantitative analysis of the inflammatory edema shows the most severe outcome on the 5th day (HE staining WBC count of 54.2±5.8 unit) and its severity progressively subsides on the 15th day. (HE staining WBC count of 41.7±7.2 vs 54.2±5.8 a, P＜0.0,5). In the early obstruction (5 d and 10 d), the liver cells showed mild to moderate swelling and its degeneration is often associated with steatosis and sinusoidal expansion and congestion. As the obstruction time increases in the 20 d and 30 d group, liver cells starts to show extensive vacuolation and sinusoidal occlusion. ConclusionsEarly phase (5 days) of acute bile duct obstruction due to injury shows rapid expansion of the bile duct, edema in the bile duct itself as well as its surrounding tissue and liver damage. After 15 days, the local inflammatory edema is greatly reduced and is replaced by hyperplasia of fibers and collagen. Liver damage appears to be irreversible after 20 days. Considering local environmental and systemic conditions, the optimal time frame to repair obstruction of bile duct surgically is between 10-20 days.