BACKGROUND: The activation of hypothalamus-pituitary-adrenal cortex axis may play key role in the increasing expression of hypothalamic corticotropin-re-leasing hormone (CRH) during stress reaction. However by what way to induce the CRH expression in hypothalamic neuron, and whether CRH can activate hypothalamic neurons are still not very clear.OBJECTIVE: To observe the changes of intracellular cyclic adenosine monophosphate (cAMP) and cytoplasmic Ca2+ concentration in the hypothalamic neurons cultured in vitro due to exogenous CRH stimulation.DESIGN: Comparative observation experiment.SETTING: Research Institute of Surgery, Daping Hospital, Third Military Medical University of Chinese PLA; Department of Neurosurgery , Tiantan Hospital, Capital University of Medical Sciences MATERIALS: This experiment was carried out in the Research Institute of Surgery, Daping Hospital, Third Military Medical University of Chinese PLA between December 1999 and March 2002. Hypothalamus was obtained from fetus rat at pregnancy of 17 days for the in vitro culture of hypothalamic neurons.METHODS: Hypothalamic neurons were co-cultured with exogenous CRH,with or without pretreatment with specific CRH 1 receptor antagonist -CP-154526. hypothalamic neurons were randomized into: ① CRH (10-12,10-10, 10-8, 10-6 mol/L) stimulation group. ② CP-154526(500 μmol/L)pretreatment aud CRH ( 10-12, 10-10, 10-8,10-6 mol/L) stimulation group. ③Hypothalamic neurons in corresponding normal control group were exposed to the isotonic saline stimulation. PTI fluorescence image system was used to determine and analyze the change of cytoplasmic free Ca2+ concentration in hypothalamic neurons due to exogenous CRH stimulation and RIA was used to detect the neuronal cAMP content.MAIN OUTCOME MEASURES: ①Cytoplasmic free Ca2+ concentration in hypothalamic neurons. ②cAMP content in hypothalamic neurons.RESULTS: The cytoplasmic free Ca2+ concentration and cAMP content were relatively lower in the hypothalamic neurons in normal control group,which obviously increased due to CRH stimulation [(240±22),(153±11)nmol/L; (3.26±0.19),(0.44±0.02) pmol/dish,P ＜ 0.01];CP-154526 could remarkably suppress the CRH (10-6 mol/L)induced increase in cytoplasmic free Ca2+ concentration and cAMP content in hypothalamic neurons [Ca2+ concentration: (240±22),(171±16)nmol/L; cAMP content:(3.26±0.19), (2.33±0.21) pmol/dish, P ＜ 0.01].CONCLUSION: CRH can directly act on hypothalamic neurons via type 1-receptor,thereby increase the cytoplasmic free Ca2+ concentration and cAMP content in hypothalamic neurons,playing the key role in the modulation of the synthesis and secretion of CRH during the activation of hypothalamic neurons.

BACKGROUND: Through what signal pathway does corticotropin release hormone (CRH) regulate hypothalamic neuronal neuroendocrine activity during acute stress?OBJECTIVE: To probe into the regulatory effects of CRH on CREB secretion in hypothalamic neurons.DESIGN: Repetitive measurement design.SETTING: At Field Surgery Research Institute of Daping Hospital, Third Military Medical University of Chinese PLA; Neurosurgery Department,Tiantan Hospital Affiliated to the Capital University of Medical Sciences.MATERIALS: This experiment was carried out in Daping Hospital of Third Military Medical University of Chinese PLA between December 1999and March 2002. Rat fetuses were selected from Wister rats of 17-day gestation.METHODS: In vitro cultured cells were divided into the following groups:① CRH (10-12, 10-10, 10-8 and 10-6 mol/L) stimulation groups. ② Pretreated with nimodipine (5 μmol/L) or CP-154526 (500 μmol/L) followed by CRH (10-12, 10-10, 10-8 and 10-6 mol/L ) stimulation groups. ③ Corresponding control groups stimulated with isotonic physiological saline. PTI fluorescence imaging system was used to detect the changes of neuronal cytoplasmic free calcium concentration; meanwhile, Western blot technique was used to determine the changes of neuronal P-CREB content.MAIN OUTCOME MEASURES: ① Changes of neuronal cytoplasmic free calcium concentration. ② Changes of neuronal P-CREB content.RESULTS: The content of cytoplasmic free calcium in hypothalamic neurons was lower in normal control group, and it increased immediately after exogenous CRH stimulation. However, such increase could be suppressed by pretreatment with nimodipine or CP-154526 before CRH stimulation,and the increase of neuronal P-CREB content was also obviously suppressed.CONCLUSION: During acute stress, the combination of CRH with hypothalamic neuronal CRH 1 receptor leads to the opening of membrane Ltype calcium ions channels, thus enhancing the influx of calcium ions and increasing cytoplasmic free calcium ions content, which would further activate P-CREB signal transduction pathway in neurons. It suggests that CRH may play a vital role in hypothalamic neuronal activation.

Objective To analyze the short-term and long-term effectiveness of radiofrequency ablation combined with transcatheter arterial chemoembolization (TACE) for the treatment of hepatocellular carcinoma (HCC).Methods The clinical data of 70 HCC patients who had received thermal ablation (group A) done or in combination with TACE (group B) were retrospectively analyzed.Results The rate of intrahepatic distant recurrence in group B (25 cases) was lower than that in group A (45 cases) (X2 =3.845,P =0.046) and the tumor-free survival rate was higher than group A (X2 =5.020,P =O.030).There were no differences in the local tumor progression rate (X2 =0.853,P =0.374) and overall survival (x2 =2.316,P =0.154) between two groups.Incidence of bone marrow suppression in group B was higher than that of group A (X2 =5.642,P =0.042).Major complications didn't occur in any group(X2 =2.016,P =0.183).The costs was higher(t =7.738,P ＜0.001) and the hospital stay was longer (t =5.921,P =0.003) in group B than group A.Conclusions Compared with ablation alone,combined therapy is able to reduce short-term recurrence,and improve tumor-free survival.Combine therapy is safe and effective method for small liver carcinoma.

Objective To construct the bioartificial liver by bone mesenchymal stem cell (BMSC) and alginate scaffold. Method Alginate scaffold was used as the cell carrier for the cultivation of BMSC and the differentiation from BMSC into hepatic like cells was induced by the cell factors of HGF, EGF and FGF-4 in the scaffold in vitro. The compatibility of the cells and the scaffold was observed by microscopy and the function of the differentiatd cells was tested. The gene of AFP and ALB was detected by RT-PCR. The secretion of ALB and the urea synthesis of the cells were tested by ALB kit and urea kit respectively. The glycogen synthesis and the CK-18 was tested by the glycogen stanning method and the immunofluorescence test. Results BMSC was able to attach, grow and proliferate well in the alginate scaffold, the well compatibility was observed by microscopy. ALB and urea were detected in the cultivating medium, the gene of ALB and AFP was identified by RT-PCR. The glycogen synthesis ability and the expression of CK-18 were induced during the differentiation. Conclusion The three dimensional atginate scaffold exhibited well compatibility with BMSC, BMSC could be differentiated into the hepatic like cell in the scaffold. BMSC and the alginate scaffold could be used to construct the bioartificial liver for the hepatic tissue engineering.

Objective To evaluate the effect of preoperative transarterial chemoembolization (TACE) for resectable hepatocellular carcinoma (HCC).Methods HCC patients undergoing up-front hepatectomy (group A) were compared with those receiving TACE before hepatectomy (group B).Results Tumor size decreased significantly after TACE (t =3.3 1,P =0.021).The rates of tumor encapsulation and liver adhesions were significantly more often seen in group B.There were fewer tumor-residual and more frequent necrosis in group B.Operative time in group B was longer (t =2.71 ,P =0.046).The average blood loss and complication rate were of no difference between the two groups.The occurrence of pleural effusion and intrahepatic recurrence rate in group A was higher than group B (x2 =3.85 ,P =0.031) (x2 =2.76,P =0.046).The overall survival rate from the second year postoperative in group B was higher than group A (x2 =3.37, P =0.043).Conclusions TACE could diminish tumor, advance encapsulation and reduce tumor-residual.Preoperative TACE does not improve 1-, 2-, and 3-year tumor-free survival rates but improve 1-, 2-, and 3-year overall survival rates.

Objective To evaluate the short-term and long-term elinical effect of hepatectomy using Tissue-Link & Cusa,compared to the Pringle maneuver.Methods Clinical data of 87 HCC patients who had received hepatectomy by the Pringle's Maneuver (group A) or Tissue-Link & Cusa (group B) were retrospectively analyzed.Results The average amount of bleeding in Group A was more than group B (t =2.030,P =0.023).The time of operation in group A was shorter than group B (t =-2.896,P =0.006).The postoperative supplement of albumin in group A was more than group B,the level of serum total bilirubin on 7th day after operation was higher than group B (P ＜ 0.05).There was no significant difference in postoperation complications and the time in hospital (P ＞0.05).The rate of incisional recurrence and the rate of metastasis in or out of the liver in group A were higher than group B (P =0.029,0.021,0.016).The 2-and 3-year tumor-free survival rates and the 3-year overall survival rates in group A were lower than that in group B (P =0.047,0.036,0.042).Conclusions Hepatectomy using Tissue-Link & Cusa is superior to the Pringle's maneuver for the treatment of primary hepatocellular carcinoma.It has a clear operative field,less operative bleeding,less damage to liver function and a lower relapse rate of incisional margin.

Objective To evaluate the impact of the re-modified Sugiura procedure on portal hemodynamics and liver function in cirrhotic patients with portal hypertension.Methods Forty patients with cirrhosis and portal hypertension who underwent the re-modified Sugiura procedure in the Yichang Second People's Hospital from June 2006 to October 2014 were studied.Changes in the free portal pressure (FPP),portal venous flow (PVF) and liver functions before and after operation were analyzed.Results (1) The FPP at different phases of the operation (after opening the abdomen,after splenectomy,and after devascularization) were (43.2 ± 1.8) cmH2O,(34.8 ± 1.6) cmH2O and (35.2 ± 1.7) cmH2O,respectively.There were significant differences in FPP between the phases of after splenectomy and after opening the abdomen,as well as after devascularization and after opening the abdomen (P ＜ 0.05).There was no significant difference in FPP between after devascularization and after splenectomy (P ＞ O.05).(2) The PVF,which were measured with Doppler sonography at 4 time points (preoperative 1 day,postoperative 10 days,postoperative 6 months,postoperative 18 months),were (1 420.4 ± 137.7) ml/min,(1 205.2 ± 126.7) ml/min,(875.8 ± 118.0) ml/min and (893.8 ± 114.7) n1/min,respectively.There were significant differences in PVF between postoperative 10 days and preoperative 1 day,between postoperative 6 months and postoperative 10 days,as well as between postoperative 18 months and preoperative 1 day (P ＜0.05).There was no significant difference in PVF between postoperative 18 months and postoperative 6 months (P ＞0.05).(3)The liver functions were evaluated using the Child-Pugh score at 4 time points (preoperative 1 day,postoperative 10 days,postoperative 6 months,postoperative 18 months).There were no significant differences among the time points,(P ＞ 0.05).Conclusion The re-modified Sugiura's procedure durably,appropriately and effectively reduced the PVF and FPP,but it did not have any negative effects on the liver functions of patients with cirrhosis.

BACKGROUND: There is no accepted treatment for liver fibrosis recently. Bone marrow meaenchymal stern cells (BMSCs) used in the treatment of liver fibrosis has been reported as an effectively treatment, but the mechanism is unclear.OBJECTIVE: To study the regulation of hepatic stellate cells mediated by human BMSCs in vitro.DESIGN, TIME AND SETTING: The cytological in vitro study was performed at the Center for Stem Cells and Tissue Engineering of Sun Yat-sen University and the Central Laboratory of Third Affiliated Hospital of Sun Yat-sen University from June to December 2008.MATERIALS: Human bone marrow masenchymal stem cells were collected from normal youth volunteers; Human hepatic stellate cells and normal liver call line L-O2 were supplied by the Animal Experimental Center of Sun Yat-sen University.METHODS: The purified human BMSCs and hepatic stellate calls were set up in Transwell co-culture system. The incubation density was 2×104cells/well. L-O2 was set up instead of human BMSCs as negative control. Hepatic stellate cells cultured alone served as blank control group. The culture was performed for 72 hours.MAIN OUTCOME MEASURES: Morphology of hepatic stellate cells and results of immunocytochemical staining. Apoptosis of hepatic stellte calls was determined by flow cytometry. Western blot were used to assay the expression of α-actin.RESULTS: Activated hepatic stellate cells presented fiat and thin shape under an inverted microscope. Fat drop was lack in cytoplasm, a -actin located in hepatic stellate calls, with the presence of high tension fibers. Compared with the L-O2 + hepatic stellate cell and hepatic stellate call groups, the apoptotic rate of hepatic stellate cells was significantly increased in the BMSC + hepatic stellate cell group (P < 0.05). α -actin expression was significantly down-regulated.CONCLUSION: Human BMSCs can inhibit activation of hepatic stellate ceils and promote them apoptosis, which may be the anti-hepatic fibrosis mechanism of BMSCs.

ObjectiveTo explore the role of vascular endothelial growth factor(VEGF) in the growth and development of hypertrophic scar.MethodsThe burn wound samples of various stages were selected from transition of wound granulation tissue to scar and in long-persisting post-burn hypertrophic scar, and the concentrations of VEGF protein were detected using enzyme-linked immunosorbent essay (ELISA) method. ResultsThe tissue homogenate concentration of VEGF protein increases gradually from the wound granulation tissue to hypertrophic scar before it achieves summit concentration during 4 to 6 month. The concentration of VEGF degreases gradually after the maturation of hypertrophic scar. The high concentration of VEGF is synonymous with the large amount of capillary of the immature scar.ConclusionsThe abnormal expression of VEGF is related to the growth and development of hypertrophic scar and induces excessive and uncontrollable angiogenesis.

Objective To detect and analyze the resistance at the stomach meridian acupoints in chronic atrophic gastritis (CAG) patients with spleen-stomach deficiency-cold syndromes, summarize the status of asthenia-sthenia at stomach meridian under this condition, in order to guide the clinical therapy with acupuncture and moxibustion.Methods From 2010 January to 2013 December, 83 CAG patients with spleen-stomach deficiency-cold syndromes were collected from No.306 Hospital of the PLA, the surface resistances on the bilateral stomach meridian stomach acupoints were detected,including Lidui, Neiting, Xiangu,Chongyang, Jiexi, Fenglong, Zusanli, Liangqiu, and the non-acupoints 1cm lateral to the point were set as controls. Zhu auricle holographic detector was adopted to detect the resistance at 20 points (about 0.5 cm2around the acupoints) in each acupoint, and the average value was calculated as the resistance value.Results Theresistances on the leftChongyang andright Chongyang were (9.64 ± 2.03) k? and (9.68 ± 2.02) k?, respectively, both were significantly lower than those on other acupoints, with statistically significant difference (allP<0.01); theresistances on the leftLiangqiu andright Liangqiu were (13.44 ± 2.11) k? and (13.68 ± 2.12) k?, respectively, both were significantly higher than those on other acupoints (all P<0.01).Conclusions The resistance on the stomach meridian stomach acupoints in CAG patients with spleen-stomach deficiency-cold syndromes is different, which may reflect the basic pathogenesis of CAG being the weakness of the spleen and stomach, qi stagnation and blood stasis.