Hepatocellular carcinoma (HCC) is a serious threat for human health, the incidence of HCC in China accounts for more than 50% worldwide. There is an urgent need to develop novel anticancer agents for the treatment of HCC patients. Here we characterized the inhibitory effect and the molecular mechanism of protopine on HCC cancer cells. The results of a CCK-8 assay indicated that protopine displays anticancer activities on HCC cells. Flow cytometry and JC-1 staining confirmed that treatment with protopine decreased the mitochondrial membrane potential and induced apoptosis in HCC cells. Western blot analysis showed that protopine was able to increase protein expression in the mitochondrial apoptotic pathway; the level of cytochrome C, apoptotic protease activating factor-1 (Apaf-1), Bax, cleaved-poly ADP-ribose polymerase (cleaved-PARP), cleaved-caspase-3, and cleaved-caspase-9 were increased while the expression of Bcl-2 was suppressed significantly. An in vivo study revealed that protopine significantly suppressed the growth of tumors in nude mice bearing HepG2 cells. Administration of protopine intraperitoneally at a concentration of 50 mg·kg^-1 inhibited tumor growth by 72.46%. Animal experiments were carried out according to the Regulation of the Animal Ethics Committee of Southwest Medical University. This study provides preliminary evidence that there is potential to develop protopine as a lead compound for the treatment of HCC.

Objective To explore the diagnostic sensitivity and specificity of echocardiography parameters ,and the diagnostic value of multiple parameters score . Methods Forty‐nine cases of the fetus checked by fetal echocardiography and were indicated aortic coarctation . According to the results of follow‐up ,the cases were divided into the positive and negative groups . Comprehensive analysis were performed in fetal echocardiography parameters ,such as diameter ratio and velocity ratio of ventricle ,aorta ,pulmonary artery ,ductus artery ,middle cerebral artery and umbilical artery . Results Diameter ratio of left ventricle/right ventricle ( LV/RV ) ,pulmonary artery/aorta ( PA/AO ) ,aortic isthmus/ductus artery ( AI/DA ) ,and velocity ratioof middle cerebral artery/inner umbilical artery ( MCA/inUA ) were calculated ,ROC curve showed their area under ROC were>0 .5 with a significant statistical significance ( P < 0 .05) . The Chi‐square test showed that the consistency of any single ratio was relatively low . When the number of the ratiois increased to 3-4 ,the Kappa value was 0 .687( P = 0 .000) and 0 .649( P = 0 .000) ,respectively , which had relatively high diagnostic value . Conclusions Ultrasonic diagnosis of fetal coarctation of aorta has certain difficulty ,and the value of single hemodynamic parameter of fetal echocardiography in diagnosis of fetal aortic coarctation is lower ,so that the comprehensive evaluation and combined multiple ultrasonic parameters can improve the sensitivity and specificity of ultrasound diagnosis .

BACKGROUND:Stem cel therapy has been used for prevention and treatment of degenerative disc disease. Considering the special microenvironment in the intervertebral disc, the survival rate and differentiation ability of transplanted cels are decreased, which may lead to the poor efficacy of stem cel therapy. How to improve the survival ability and therapeutic effect of the transplanted cels is the focus of stem cel therapy for degenerative disc disease.
OBJECTIVE: To investigate the effects of cobalt chloride combined with hypertonic solution pretreatment on bone marrow mesenchymal stem cels that wil be transplanted for treatment of degenerative disc disease.
METHODS:(1)In vitro cel experiment: bone marrow mesenchymal stem cels were divided into three groups and subjected to normal culture medium (normal control group), 1% hypertonic mother solution (hypertonic group), 100 μmol/L cobalt chloride (hypoxia group), or 1% hypertonic mother solution plus 100 μmol/L cobalt chloride (combined group) for 1 week. Then, 2% hypertonic solution and 200 μmol/L cobalt chloride cobalt chloride were used to simulate the anaerobic and hypertonic environment intervenes in pretreated and untreated bone marrow mesenchymal stem cels for 24 hours. After that, RT-PCR was used to detect the expression of caspase-3 for apoptosis evaluation. (2)In vivo animal experiment: Sprague-Dawley rats were divided into model, cel transplantation and hypertonic plus hypoxic groups. Rat models of intervertebral disc degeneration were made in these three groups. After modeling, rats in these three groups were given no treatment, bone marrow mesenchymal stem cel transplantation or transplantation of bone marrow mesenchymal stem cels which were subjected to hypertonic and hypoxia pretreatments into the intervertebral disc. Two weeks later, immunohistochemistry and RT-PCR methods were used to detect cel distribution and related gene expression, respectively, thereby to evaluate the therapeutic effect of stem cels.
RESULTS AND CONCLUSION: (1)In vitro cel experiment: caspase-3 mRNA expression was significantly reduced in pretreated bone marrow mesenchymal stem cels compared with the untreated cels (P < 0.05). (2)In vivo animal experiment: compared with the control group, the caspase-3 and interleukin-1β in the intervertebral disc and a number of degenerative indexes were decreased in the cel transplantation. Compared with the cel transplantation group, these indicators had better outcomes in the hypertonic plus hypoxic group (P < 0.05). These findings indicate that bone marrow mesenchymal stem cels have therapeutic potential for degenerative disc disease, and have better adaptability and transplantation effects by hypertonic and hypoxia pretreatments.

ObjectiveTo explore the effect of different nutritional support mdoes on humoral immunity and outcomes after esophagectomy in patients with esophageal carcinoma.MethodsForty-six patients with middle or low thoracic esophageal carcinoma underwent Ivor Lewis esophagectomy.The patients were randomized into enteral nutrition group ( EN,n =23 ) and enteral combined parenteral nutrition group ( EN + PN,n =23 ) based on the nutrition support modes.Serum levels of immunoglobulin (IgG,IgA,IgM,IgE,κ/λ light chain) and comphments (C3/C4) were assayed and compared on the 1st pre-operative day and at 18 hours as well as 3rd and 7th day after operation.The clinical outcomes including infection-related complications and hospital stay were compared between two group s.ResultsThere was no significant difference in all humoral immunity indicators between two groups at the eachpost-operative time point.In both two groups,the levels ofIgG [ (8.90 ± 1.75),(7.53 ±1.41) g/Land (8.64±2.44),(7.48±2.16) g/L],κ [ (2.14±0.46),(1.78±0.41) g/L,and (2.15 ±0.63),( 1.86 ± 0.62) g/L] and λ light chain [ ( 1.34 ± 0.45 ),( 1.11 ± 0.31 ) g/L and ( 1.20 ± 0.32),( 1.08 ± 0.35 ) g/L] were significantly lower 18 hours and 3rd day after operation than the pre-operative levels [ (12.15±2.86)and (11.11±2.96) g/L,(2.90±0.77) and (2.77±0.79) g/L,(1.79±0.57) and (1.56±0.41) g/L] (P=0.000,P=0.000,and P=0.004,P=0.000,and P=0.000,P=0.000,and P=0.011,P=0.000,and P=0.004,P=0.000,and P =0.008,P =0.000),and returned to the preoperative levels by the postoperative 7th day (P＞0.05),except for the level of κ light chain 7th day after operation in EN group [ ( 2.42 ± 0.69) g/L] ( P =0.027 ).The levels of IgA,IgE,and C3 were not significantly different during the perioperative period ( P ＞ 0.05 ).The level of IgM was not significantly different during the perioperative period in EN group (P ＞0.05),and was significantly lower on the 3rd post-operative day [ ( 1.00 ±0.53) g/L] than the pre-operative level [ ( 1.47 ±0.76) g/L] in the EN + PN group (P =0.031 ),and were not significantly different on the other time points (P ＞ 0.05 ).In the EN group,the C4 level was significantly lower at the postoperative 18 hours [ (0.24 ±0.08) g/L] than the pre-operative level [ (0.37 ±0.36) g/L] (P =0.030),and were not significantly different at the other time points ( P ＞ 0.05 ).In the EN + PN group,the C4 level was not significantly different during the perioperative period ( P ＞ 0.05 ).There was no significant difference in the infection-related complications and hospital stay between these two groups ( P =0.300,P =0.371 ).ConclusionsThe effects of EN or EN + PN on humoral immunity and outcomes after esophagectomy in patients with esophageal carcinoma are not different.Both these two nutritional support modes can not completely alleviate the harm to the humoral immunity.The EN is more cost-effective.

Objective To observe the effects of transplantation of autologous adipose-derived stem cells (ASCs) on osteoporosis (OP) in a rabbit ovariectomy (OVX) model.Methods A total of fifteen 6-month-old female New Zealand white rabbits were randomly divided into two groups:ovariectomy group (group A,n =12) and sham operation group (group B,n =3).All rabbits were subjected to bilateral ovariectomy in the group A.Six months later,bone mineral density (BMD) of group A and group B were measured by dual energy X-ray absorptiometry (DXA) to check the result of OVX-OP.ASCs harvested from adipose of OP rabbits were cultured to be expanded and differentiated in osteogenic medium in vitro.Osteogenesis was evaluated by alizarin red staining,alkaline phosphatase (ALP) staining and quantitative assays of osteocalcin (OCN).Autologous osteo-induced ASCs were mixed in calcium alginate hydrogel (CAH) and then transplanted in the left distal femurs,while CAH was transplanted in the right distal femurs of OP rabbits.At 12 weeks after implantation,BMD,micro-CT and histomorphological analyses were performed on these rabbits.Results The BMD of femurs in group A rabbits were obviously lower than that of group B rabbits (P ＜ 0.05) at 6 months after OVX.Compared with control group,ASCs cultured in osteo-induction medium had similar proliferation rate as the non-induced cells,but displayed positive ALP and alizarin red staining and OCN contents.At 12 weeks after implantation,the cell-treated femurs displayed higher BMD,bone trabecula number,trabecula thickness and separation than those of control group,while the structure model index and porosity were lower (P ＜ 0.05).Histological examination indicated that the trabecular thickness increased with complete CAH resorption in cell-treated group,while CAH remained in control group.Conclusions Transplantation of autologous ASCs can help strengthen osteoporotic bone in OVX-OP rabbits,providing a novel approach to OP treatment.

ObjectiveTo explore the relationship of trefoil factor family 3 (TFF3),vascular endothelial growth factor (VEGF) and hypoxia-inducible factor (HIF)-1α in gastric cancer SGC-7901 cells under hypoxic condition and try to investigate the mechanism of TFF3 in the genesis and development of gastric cancer. Methods The hypoxic model of gastric cancer SGC-7901 cell was induced by CoCl2 Gastric cancer cell line SGC-7901 cells were transfccted with pU6-siTFF3 plasmid which carrying RNAi targeted to human TFF3 and pU6-mock.Puromycin was selected as screening medicine.The stable and specific TFF3 inhibited gastric cancer cell line was established. Gastric cancer cell line SGC-7901 and TFF3 RNAi targeted gastric cancer cell line SGC 7901 were cultured under hypoxic condition and normoxic condition. The expression of TFF3,VEGF and HIF-1a at protein and mRNA level were detected by RT-PCR,Western blot and ELISA assay.The distribution and expression of TFF3 and HIF-1α in gastric cancer cell line SGC-7901 cells uuder normoxia and hypoxic condition were determined with immunofluorescence staining.Results The expressions of HIF-1a,TFF3 and VEGF in gastric cancer SGC-7901 cell increased under CoCl2 induced hypoxic condition (33.4 =1.8,14.8 ± 1.1 and 15.1 ± 1.2,respectively). Under hypoxie condition,the expression of VEGF and HIF-1α protein reduced in stable TFF3 RNAi SGC-7901cells.Conclusion TFF3 mediated the regulation of VEGF and HIF-1α expression under hypoxic condition.TFF3might be a potential anti-angiogenic target in gastric cancer treatment.

Objective To investigate the effects of anaesthetic concentration of sevoflurane on TM3 mouse leydig cell viability.Methods TM3 mouse leydig cells were randomly divided into 3 groups ( n =24 dishes each):control group (group C),2％ and 5％ sevoflurane groups (groups SEV1 and SEV2 ).The cells were collected after being exposed to sevoflurane or 95 ％ room air + 5 ％ CO2 for 2,4 or 6 h (T1-3) for microscopic examination with optical binocular inverted microscope.The number of live cells was counted by using cell counting kit8.Gene chips were used to indentify differentially expressed genes between group C and group SEV2 after being exposed to air and 5 ％ sevoflurane for 6 h respectively.Results The leydig cell viability was significantly decreased at T3 in group SEV2 as compared with groups C and SEV1.Morphological changes were found only in group SEV2.A total of 45 genes were identified to be differentially expressed in group SEV2 as compared with group C.The level of expression of prostaglandin-endoperoxidase synthase 2 gene (Ptgs2),chemokine (C-C motif) ligand 2(CCL2) gene and dual specificity phosphatase1 (Dusp1) gene increased by at least 4 times in group SEV2.Conclusion Sevoflurane can inhibit the cell viability of TM3 mouse leydig cell in concentration dependent manner through abnormal expression of Ptgs2,CCL2 and Dusp1 genes.

Objective To analyze gene mutations of a Niemann-Pick disease type C (NPC) proband,and carry out prenatal diagnosis for the family.Methods The coding regions of NPC1 gene in the proband (late-infantile form) and white blood cell (WBC) in peripheral blood of its parents were amplified by polymerase chain reaction and direct DNA sequencing in both directions was performed.The sequencing results were compared with human NPC1 gene sequence (NM_000271) in GenBank,and sequences of mutated exons were determined.Direct sequencing was used on 50 normal Chinese individuals' DNA samples (control) to exclude mutation's single nucleotide polymorphism (SNP).An inter-species alignment of homologous NPC1 proteins was performed using ClustalX 1.81 software.During the second pregnancy of the proband's mother,the amniotic fluid was obtained at 18 weeks of gestation and the amniocytes were cultured for gene mutation analysis.Neonate's DNA of WBC in peripheral blood was also extracted for NPC1 gene analysis.Results Mutation analysis of NPC1 gene revealed two novel heterozygous mutations (c.2284-2287 delCTCT and p.V959G) in the proband,which originated from her father and mother,respectively.These two mutations were absent in the control,suggesting that these mutations were not SNP.While comparing with the amino acid in NPC1 protein of human,mouse,rat,rabbit,cat and pig,it revealed that p.V959 belonged to a conservative amino acid region and the missense mutation of p.V959G may perturb the function of NPC protein.Neither mutation was found in DNA from amniotic fluid or from the cultivated amniocytes in the second pregnancy,suggesting a normal fetus.c.2284-2287 delCTCT and p.V959G mutation were not found in NPC1 gene analysis of WBC in peripheral blood of the neonate,which was consistent with the prenatal diagnosis.Conclusions PCR-direct sequencing could be used as genetic diagnosis for NPC proband and prenatal diagnosis for its family.The mutation p.V959G may be correlated to late infantile form of NPC.

Objective To retrospectively analyzed the echocardiographic characteristics of ventricular diverticula in fetuses ,children and adults ,and discuss the main points of diagnoses and prognoses of ventricular diverticula . Methods Echocardiographic characteristics ,clinical data and intraoperative findings of ventricular diverticula of 8 fetuses ,3 children and 14 adults were summarized . Results The ventricular diverticula in 8 fetuses and 3 children were all single . Twenty five diverticula were found in the 14 adult patients ;in other words ,there were 3 multiple diverticula found in adult group . A statistical difference was found in ratio of diverticula area and corresponding ventricular area ( P = 0 .021 ) ,and corresponding ventricular fractional shortening ( P =0 .003) between fetal and single adult ventricular diverticula . There were 2 cases with ventricular septal defects ,1 with pericardial effusion ,1 with single umbilical artery ,and 2 with atrioventricular valvular regurgitation in fetal cases . Congenital complex cardiac disease and dextrocardia were found in 1 child . There were 1 case with supravalvular aortic stenosis , 1 with nonobstructive hypertrophic cardiomyopathy ,1 with left ventricular noncompaction ,and 1 with mitral reguigitation in adult group . All the 7 diverticula with ventricular arrhythmias ,ST‐T changes and abnormal Q wave were fibrous type . And 2 multiple diverticula were accompanied with ventricular dysfunction . Among the fetal cases ,2 cases were born ,3 cases were terminated pregnancy ,and 3 cases were lost to follow up . Three children and 2 adults were underwent cardiac operations for diverticula with good outcomes . No such complications as cardiac rupture ,thromboembolism ,and sudden death were found . Conclusions Echocardiography is a useful method to diagnose ventricular diverticulum ,especially for fetuses ,and provide the basis for prognosis consultation .

We reported a case of cryptogenic organizing pneumonia ( COP) who was admitted to the hospital in July 2008 and reviewed the Chinese literature of COP from 2003 to 2008. The most common symptoms of COP are fever, cough and exertional dyspnea. The imaging characteristics of COP are similar to those of pneumonia, therefore is often misdiagnosed as pneumonia with a high misdiagnosis rate. Lung biopsy is the main method for pathological diagnosis; polypoid growth of granulation tissue was noted within respiratory bronchioles, small airways and alveolar spaces.