重组牛碱性成纤维细胞生长因子滴眼液对白内障术后干眼症的预防效果

目的观察重组牛碱性成纤维细胞生子因子滴眼液(贝复舒滴眼液)对白内障术后干眼症的预防效果。方法选取该院2022年2月至2023年8月接受手术治疗的白内障患者82例,随机分为对照组与试验组各41例。两组均接受超声乳化白内障吸除术治疗,术前以5%聚维酮碘溶液+0.9%氯化钠注射液冲洗结膜囊,试验组在此基础上于术后第1 d开始术眼使用贝复舒滴眼液。比较两组术前与术后第7 d角膜形态、眼表指标变化及术后干眼症发生情况。结果术后第7 d试验组角膜表面规则性指数、角膜表面非对称性指数、泪膜破裂时间、角膜荧光素染色评分及基础泪液分泌试验长度明显优于对照组,术后干眼症发生率(7.3%,3/41)明显低于对照组(24.4%,10/41),差异有统计学意义(P<0.05)。结论贝复舒滴眼液配合5%聚维酮碘溶液,能改善白内障术后患者角膜情况及泪膜功能,预防术后干眼症的发生。

甲状旁腺切除术对维持性血液透析患者成纤维细胞生长因子23的影响

目的探讨甲状旁腺切除加前臂自体移植术（total parathyroidectomv with autotransplantation，tPTX＋AT）对维持性血液透析（maintenance hemodialysis，MHD）合并严重继发性甲状旁腺功能亢进（secondary parathyroidectomy，SPTH）患者的成纤维细胞生长因子23（FGF-23）的影响。方法选取2014．2016年在本院行MHD合并严重SPTH的患者，符合人选条件者按照患者意愿分成手术组与非手术组，手术组行tPTX＋AT，非手术组予药物治疗。对比两组治疗前和治疗后6个月的生化指标、FGF-23水平。结果共纳入48例患者，手术组22例，非手术组26例，两组患者间年龄、透析龄、原发病、术前全段甲状旁腺素（iPTH）、血磷、FGF-23、胆固醇（TCH）、三酰甘油（TG）、白蛋白（ALB）、血红蛋白（HGB）的差异均无统计学意义，手术组术前的血钙与碱性磷酸酶（ALP）水平显著高于非手术组（P〈0．01）。术后6个月手术组血iPTH、血磷、血钙、钙磷乘积水平显著低于非手术组（P〈0．05），而两组间血脂、ALB、HGB、ALP水平差异无统计学意义（均P〉0．05）；手术组血FGF-23术后1周、1个月、3个月、6个月逐渐下降，与术前相比差异均有统计学意义（P〈0．05）；术后6个月手术组血FGF．23水平显著低于非手术组[1462．9（903．7，5826．9）ng／L比12627．9（5488．9，16844．4）ngm，P〈0．01]。结论tPTX＋AT可明显改善MHD合并严重SPTH患者的钙磷代谢紊乱，术后6个月内逐渐降低血FGF-23水平。

bFGF对LASIK后角膜上皮、内皮和神经修复作用的临床研究

目的临床观察重组碱性成纤维细胞生长因子(recombinant basic fibroblast growth factor,rbFGF)对准分子激光原位角膜磨镶术(laser assisted in situ keratomileusis,LASIK)后主观症状缓解、角膜上皮、内皮和神经修复的作用。方法一定时间段内抽取LASIK术后患者,随机分为试验组和对照组,各30例(60眼)。术后常规给予1g·L-1氟米龙滴眼液+3g·L-1氧氟沙星滴眼液,每天4次,试验组加rbF-GF(贝复舒)滴眼液,对照组加玻璃酸钠(爱丽)滴眼液每天4次,各治疗3个月。术后第1天、第2周、第1个月、第3个月对患者主观症状、客观体征及顺应性进行平行对照并评价整体疗效。主观指标包括视疲劳、模糊感、异物感、干涩感、刺痛感。客观指标包括视力、泪膜破裂时间、泪液分泌试验(Schirmer’s TestⅠ)、角膜荧光染色、新生血管、基质混浊(haze)、角膜内皮计数、角膜知觉等。所有数据均采用SPSS11.5软件进行处理。结果入选病例中性别、年龄、术前裸眼视力2组间无统计学差异(P>0.05)。各组用药效果:除眼干涩感P>0.05外,其余各主观症状好转,2组统计学均有显著性差异(P<0.01)。术后早期角膜染色2组均增加,用药治疗后减少,统计学有显著性差异(P<0.01);试验组用药后Schirmer’s TestⅠ较用药前增加,差异有统计学意义(P<0.05);对照组Schirmer’s TestⅠ及2组泪膜破裂时间、角膜内皮计数用药前后无统计学差异(P>0.05);术后早期角膜知觉下降,以后逐渐恢复,各组治疗前后比较有显著统计学差异(P<0.01)。各组术后未见新生血管增加或加重病例,均未见haze。2组顺应性好,均未出现副作用。2组间用药效果的比较:术后2周刺痛感对照组较试验组轻,二者比较有统计学差异(P<0.05,差值法),其余各项主观症状2组间无显著差异(P>0.05);而用药2周后对术后5项症状的评分总和缓解程度试验组优于对照组,差异有显著统计学意义。术后第1天、第2周、第1个月和第3个月的裸眼视力试验组分别为0.99±0.28、1.06±0.25、1.11±0.21和1.14±0·21,对照组分别为1.05±0·28、1.16±0.20、1.22±0.19和1·14±0.17,各时间点2组比较无统计学差异(P>0.05)。术后2周起Schirmer’s TestⅠ试验组均优于对照组,比较有统计学差异(P<0.05),其余各时间点比较无差异(P>0·05);其他客观体征2组间差异无统计学意义(P>0.05)。结论bFGF安全、顺应性好,可缓解LASIK术后眼表不适症状,并对LASIK后角膜上皮及神经修复、泪液分泌、角膜内皮保护有一定作用,且rbFGF治疗3个月后未见角膜新生血管生成或加重、haze以及其他毒副作用发生。

Effects of basic fibroblast growth factor on ischemic gut and liver injuries

AIMS To explore the possible effects of basic fibrob- last growth factor (bFGF) on ischemic gut and liver in- juries after trauma. METHODS Animal model of super mesenteric artery (SMA) occlusion was used in this study. Seventy-two Wistar rats were divided into three groups of 24 rats each. Each animal in group 1 (bFGF treated) was in- jected with 4 μg of bFGF in 0.15 ml of normal saline solution containing 0.1%(w/v) heparin through the jugular vein at the onset of reperfusion. Animals in group 2 (saline treated) received the same vehicle, but without bFGF. Group 3 (sham-operated) ani- mals were treated with the same operations,but without SMA occlusion. Liver function parameters, serum TNFα,bacterial examination and pathological study were used to evaluate the results. RESULTS In group 1,the amounts of ALT and AST and serum TNFα were reduced significantly at 6,24 and 48 hours as compared with group 2. Bacterial ex- amination showed that the bacterial translocation from gut to liver,spleen and MLN in group 1 was much lower than that in group 2. The pathological results support the concept of significant protecting effects of bFGF. CONCLUSIONS Venous administration of bFGF may help reduce gut and liver injuries after ischemia and reperfusion. Its mechanism of action may involve the mitogenic and non-mitogenic effects of bFGF.

Involvement of MAPK/ERK kinase-ERK pathway in exogenous bFGF-induced Egr-1 binding activity enhancement in anoxia-reoxygenation injured astrocytes

Objective Intravenous administration of basic fibroblast growth factor （bFGF） is effective to reduce the volume of cerebral infract due to ischemia. This study was designed to investigate the molecular mechanism, especially the signal transduction pathways, involved in this protective role of bFGF. Methods Anoxia-reoxygenation treated atrocytes were used to study the role of mitogen-activated protein kinase/extracellular signal-regulated kinase kinase （MAPK/ERK kinase, MEK）-ERK signaling pathway after exogenous bFGF administration by Western blot. Electrophoretic mobile shift assay was used to detect the binding activity of early growth response factor-1 （Egr-1）, an important transcription factor for endogenous bFGF. Results bFGF could protect some signal transduction proteins from the oxygen-derived free radicals induced degradation. ERK1/2 was activated and involved in Egr-1 binding activity enhancement induced by exogenous bFGF. Conclusion MEK-ERK MAPK cascade may be an important signal transduction pathway contributed to bFGF induced enhancement of Egr-1 binding activity in anoxia-reoxygenation injured astrocytes.

Effects of Proliferation of CEF with Ginsenoside and Its Derivatives

Objective The research aimed to provide theoretical basis for studying anti-MDV mechanism of ginsenoside and its derivatives in vitro. Method Effects of ginsenoside and its derivatives on proliferation activity of chick embryo fibroblast （CEF） in vitro were determined by using neutral red dye absorption method. Result The results showed the proliferation effects of different drugs are not completely same, and are more obvious with low toxic drugs. Detected at different action times, the differences of OD values was biggest at 72 h when compared with normal control group, while there was no significant difference at 24 h. Conclusion Ginsenoside and its derivatives could promote the proliferation of CEF cells in medium as low concentrations, which have time-dependent characteristic.

Study on Agrobacterium tumefaciens-mediated Transformation of Brassica campestris L. with Fusion Gene Ycoil-bFGF

[ Objective] The study is to generate pharmaceutical protein via plant transgenic technique. [Methed] Using the cotyledons with petiole as transformation receptor, the fusion gene of rapeseed oil-body gene and bFGF was introduced into the rapeseed （ Brassica campestris L. ） by Agrobacterium tumefaciens-mediated transformation; meanwhile regeneration conditions of rapeseed were also optimized, and the regenerated resistant plantlets were detected by PCR and Southern blot. [ Result] This fusion gene had been integrated into rapeseed genome successfully, and the optimized conditions of transformation and regeneration were as follows： explants pre-culture for 2 d, co-culture for 3 d, bacteria solution OD600 for 0.3 and infection time for 5 min. [ Conclusion] The results laid a solid foundation for extraction, isolation and purification of protein in transgenic plant seeds.

Cellular and molecular mechanisms of intestinal fibrosis

Fibrosis is a chronic and progressive process characterized by an excessive accumulation of extracellular matrix (ECM) leading to stiffening and/or scarring of the involved tissue. Intestinal fibrosis may develop in several different enteropathies, including inflammatory bowel disease. It develops through complex cell, extracellular matrix, cytokine and growth factor interactions. Distinct cell types are involved in intestinal fibrosis, such as resident mesenchymal cells (fibroblasts, myofibroblasts and smooth muscle cells) but also ECM-producing cells derived from epithelial and endothelial cells (through a process termed epithelialand endothelial-mesenchymal transition), stellate cells, pericytes, local or bone marrow-derived stem cells. The most important soluble factors that regulate the activation of these cells include cytokines, chemokines, growth factors, components of the renin-angiotensin system, angiogenic factors, peroxisome proliferator-activated receptors, mammalian target of rapamycin, and products of oxidative stress. It soon becomes clear that although inflammation is responsible for triggering the onset of the fibrotic proc-ess, it only plays a minor role in the progression of this condition, as fibrosis may advance in a self-perpetuating fashion. Definition of the cellular and molecular mechanisms involved in intestinal fibrosis may provide the key to developing new therapeutic approaches.

Influence of ginsenoside Rg1, a panaxatriol saponin from Panax notoginseng, on renal fibrosis in rats with unilateral ureteral obstruction

Total saponins of Panax notoginseng （PNS） have been shown to ameliorate renal interstitial fibrosis. Ginsenoside Rg 1, a panaxatriol saponin, is one of the major active molecules from PNS. The present study was undertaken to investigate the effect of ginsenoside Rgl on renal fibrosis in rats with unilateral ureteral obstruction （UUO）. The rats were randomly divided into 3 groups： sham-operation （n=15）, UUO （n=15） and UUO with ginsenoside Rgl treatment （n=15, 50 mg per kg body weight, intraperitoneally （i.p.） injected）. The rats were sacrificed on Days 7 and 14 after the surgery. Histological examination demonstrated that ginsenoside Rgl significantly inhibited interstitial fibrosis including tubular injury as well as collagen deposition, u-smooth muscle actin （α-SMA） and E-cadherin are two markers of tubular epithelial-myofibroblast transition （TEMT）. Interestingly, ginsenoside Rgl notably decreased α-SMA expression and simultaneously enhanced E-cadherin expression. The messenger RNA （mRNA） of transforming growth factor-β1 （TGF-β1）, a key mediator to regulate TEMT, in the obstructed kidney increased dramatically, but was found to decrease significantly after administration of ginsenoside Rg 1. Further study showed that ginsenoside Rgl considerably decreased the levels of both active TGF-β1 and phosphorylated Smad2 （pSmad2）. Moreover, ginsenoside Rgl substantially suppressed the expression of thrombospondin-1 （TSP-1）, a cytokine which can promote the transcription of TGF-β1 mRNA and the activation of latent TGF-β1. These results suggest that ginsenoside Rgl inhibits renal interstitial fibrosis in rats with UUO. The mechanism might be partly related to the blocking of TEMT via suppressing the expression of TSP-1.

Effect of basic fibroblast growth factor(bFGF) on the treatment of exposure of the orbital implants

Objective: To evaluate the efficacy and the indication of basic fibroblast growth factor (bFGF) in the treatment of exposure of orbital implants. Design: Retrospective and observational case series. Methods: We reviewed 41 patients (41 eyes) suffering exposure of orbital implants from Jan. 2000 to June 2006. The study group patients with mild exposure received com-bined treatment with bFGF and antibiotic drops, and while the control group patients with mild exposure were treated with anti-biotic drops only. The study group patients with moderate and severe exposure received combined treatment with bFGF and antibiotic drops, and after 2 months they were subjected to amniotic membrane transplantation, while the control group patients with moderate and severe exposure underwent amniotic membrane transplantation after using antibiotic drops. Observation of the growth of conjunctival epithelium and comparison of the healing rate of the two groups. Results: The healing rates of the mild, moderate and severe exposure study group were 100% and 92.3%. The healing rates of the mild, moderate and severe exposure control group were 55.6% and 66.7% respectively. The difference of the healing rates of the mild exposure study group and the control group was significant (P=0.033). And the difference of the healing rates of the moderate and severe exposure study group and the control group was not significant (P=0.167). Conclusion: bFGF may promote obviously the healing of orbital implant exposure, particularly it can be the first choice for the treatment of mild degree exposure. For the moderate and severe cases, it can be administered before surgical repair to enhance neovascularization and will tend to increase the success rate of surgical repair.

Toxicarioside A inhibits SGC-7901 proliferation,migration and invasion via NF-κB/bFGF signaling

AIM:To investigate the inhibitory role of toxicarioside A on the gastric cancer cell line human gastric cancer cell line(SGC-7901) and determine the underlying molecular mechanism.METHODS:After SGC-7901 cells were treated with toxicarioside A at various concentrations(0.5,1.5,4.5,9.0 μg/mL) for 24 h or 48 h,cell viability was determined by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl2H-tetrazolium bromide assay,and the motility and invasion of tumor cells were assessed by the Transwell chamber assay.Immunofluorescence staining,reverse transcription polymerase chain reaction and Western blotting were performed to detect the expression of basic fibroblast growth factor(bFGF) and fibroblast growth factor receptor-1(FGFR1),and nuclear factorkappa B(NF-κB) activation was examined by electrophoretic mobility shift assay.RESULTS:The results showed that toxicarioside A was capable of reducing cell viability,inhibiting cell growth,and suppressing cell migration and invasion activities in a time-and dose-dependent manner in SGC-7901 cells.Further analysis revealed that not only the expression of bFGF and its high-affinity receptor FGFR1 but also the NF-κB-DNA binding activity were effectively blocked by toxicarioside A in a dose-dependent manner compared with the control group(P < 0.05 or P < 0.01).Interestingly,application of the NF-κB specific inhibitor,pyrrolidinedithiocarbamate(PDTC),to SGC-7901 cells significantly potentized the toxicarioside A-induced down-regulation of bFGF compared with the control group(P < 0.05).CONCLUSION:These findings suggest that toxicarioside A has an anti-gastric cancer activity and this effect may be achieved partly through down-regulation of NF-κB and bFGF/FGFR1 signaling.

Significant roles of anti-aging protein klotho and fibroblast growth factor23 in cardiovascular disease

The klotho gene has been identified as an aging suppressor that encodes a protein involved in cardiovascular disease （CVD）. The inac- tivation of the klotho gene causes serious systemic disorders resembling human aging, such as atherosderosis, diffuse vascular calcification and shortened life span. Klotho has been demonstrated to ameliorate vascular endothelial dysfunction and delay vascular calcification. Fur- thermore, klotho gene polymorphisms in the human are associated with various cardiovascular events. Recent experiments show that klotho may reduce transient receptor potential canonical6 （TRPC6） channels, resulting in protecting the heart from hypertrophy and systolic dys- function. Fibroblast growth factor23 （FGF23） is a bone-derived hormone that plays an important role in the regulation of phosphate and vi- tamin D metabolism. FGF23 accelerates urinary phosphate excretion and suppresses 1,25-dihydroxy vitaminD3 （1,25（OH）2D3）synthesis in the presence ofFGF receptorl （FGFR1） and its co-receptor ldotho, principally in the kidney. The hormonal affects of circulating klotho pro- tein and FGF23 on vascular and heart have contributed to an understanding of their roles in the pathophysiology of arterial stiffness and left ventricular hypertrophy. Klotho and FGF23 appear to play a critical role in the pathogenesis of vascular disease, and may represent a novel potential therapeutic strategy for clinical intervention.

Transforming growth factor-β1 induces intestinal myofibroblast differentiation and modulates their migration

AIM： To investigate the effects of transforming growth factor β1 （TGF-β1） on the differentiation of colonic lamina propria fibroblasts （CLPF） into myofibroblasts in vitro.METHODS： Primary CLPF cultures were incubated with TGF-β1 and analyzed for production of m-smooth muscle actin （α-SMA）, fibronectin （FN） and FN isoforms. Migration assays were performed in a modified 48-well Boyden chamber. Levels of total and phosphorylated focal adhesion kinase （FAK） in CLPF were analyzed after induction of migration.did not change α-SMA levels, while TGF-β1 treatment for 6 d significantly increased α-SIVlA production. Short term incubation （6 h） with TGF-β1 enhanced CLPF migration, while long term treatment （6 d） of CLPF with TGF-β1 reduced migration to 15%-37% compared to untreated cells. FN and FN isoform mRNA expression were increased after short term incubation with TGF-β1 （2 d） in contrast to long term incubation with TGF-β1 for 6 d. After induction of migration, TGF-β1-preincubated CLPF showed higher amounts of FN and its isoforms and lower levels of total and phosphorylated FAK than untreated cells.CONCLUSION： Long term incubation of CLPF with TGF-β1 induced differentiation into myofibroblasts with enhanced α-SMA, reduced migratory potential and FAK phosphorylation, and increased FN production. In contrast, short term contact （6 h） of fibroblasts with TGF-β1 induced a dose-dependent increase of cell migration and FAK phosphorylation without induction of α-SMA production.

Targeted systemic therapies for hepatocellular carcinoma:Clinical perspectives,challenges and implications

Hepatocellular carcinoma （HCC） is a lethal disease in most patients, due to its aggressive course and a lack of effective systemic therapies for advanced disease. Surgical resection and liver transplantation remain the only curative options for a small subset of patients. Few patients with HCC are diagnosed early enough to be eli- gible for curative treatment. Angiogenesis inhibition is a natural therapeutic target for all solid tumors, but par- ticularly for the highly vascularized HCC tumors. With the approval of the targeted agent sorafenib, there are now additional options for patients with HCC. Although sorafenib does produce some improvement in survival in HCC patients, the responses are not durable. In addi- tion, there are significant dermatologic, gastrointestinal, and metabolic toxicities, and, as importantly, there is still limited knowledge of its usefulness in special sub- populations with HCC. Other angiogenesis inhibitors are in development to treat HCC both in the first-line set- ting and for use following sorafenib failure; the furthest in development is brivanib, a dual fibroblast growth factor pathway and vascular endothelial growth factor receptor inhibitor. Additional agents with antiangiogenic properties also in phase IT and Ⅲ development for the treatment of patients with HCC include bevacizumab, ramucirumab, ABT-869, everolimus and ARQ 197.

Transplantation of microencapsulated umbilical-cord-blood-derived hepatic-like cells for treatment of hepatic failure

AIM: To investigate intraperitoneal transplantation of microencapsulated hepatic-like cells from human umbilical cord blood for treatment of hepatic failure in rats. METHODS: CD34+ cells in umbilical cord blood cells were isolated by magnetic cell sorting. In the in vitro experiment, sorted CD34+ cells were amplified and induced into hepatic-like cells by culturing with a combination of fibroblast growth factor 4 and hepatocyte growth factor. Cultures without growth factor addition served as controls. mRNA and protein levels for hepatic-like cells were analyzed by reverse transcription-polymerase chain reaction, immunohistochemistry and immunofluorescence. In the in vivo experiment, the hepatic-like cells were encapsulated and transplanted into the abdominal cavity of acute hepatic failure (AHF) rats at 48 h after D-galactosamine induction of acute hepatic failure. Transplantation with PBS and unencapsulated hepatic-like cells served as controls. The mortality rate, hepatic pathological changes and serum biochemical indexes were determined. The morphology and structure of microcapsules in the greater omentum were observed. RESULTS: Human albumin, alpha-fetoprotein and GATA-4 mRNA and albumin protein positive cells were found among cultured cells after 16 d. Albumin level in culture medium was significantly increased after culturing with growth factors in comparison with culturing without growth factor addition (P < 0.01). Compared with the unencapsulated group, the mortality rate of the encapsulated hepatic-like cell-transplanted group was significantly lower (P < 0.05). Serum biochemical parameters, alanine aminotransferase, aspartate aminotransferase and total bilirubin in the encapsulated group were significantly improvement compared with the PBS control group (P < 0.01). Pathological staining further supported these findings. At 1-2 wk post-transplantation, free microcapsules with a round clear structure and a smooth surface were observed in peritoneal lavage fluid, surviving cells inside microcapsules were found by trypan blue staining, but some fibrous tissue around microcapsules was also detected in the greater omentum of encapsulated group by hematoxylin and eosin staining. CONCLUSION: Transplantation of microencapsulated hepatic-like cells derived from umbilical cord blood cells could preliminarily alleviate the symptoms of AHF rats.

Feeder-free maintenance of hESCs in mesenchymal stem cell-conditioned media： distinct requirements for TGF-β and IGF-Ⅱ

A paracrine regulation was recently proposed in human embryonic stem cells （hESCs） grown in mouse embryonic fibroblast （MEF）-conditioned media （MEF-CM）, where hESCs spontaneously differentiate into autologous fibroblastlike cells to maintain culture homeostasis by producing TGF-β and insulin-like growth factor-lI （IGF-Ⅱ） in response to basic fibroblast growth factor （bFGF）. Although the importance of TGF-β family members in the maintenance of pluripotency of hESCs is widely established, very little is known about the role of IGF-Ⅱ. In order to ease hESC cul- ture conditions and to reduce xenogenic components, we sought （i） to determine whether hESCs can be maintained stable and pluripotent using CM from human foreskin fibroblasts （HFFs） and human mesenchymal stem cells （hM- SCs） rather than MEF-CM, and （ii） to analyze whether the cooperation of bFGF with TGF-β and IGF-Ⅱ to maintain hESCs in MEF-CM may be extrapolated to hESCs maintained in allogeneic mesenchymal stem cell （MSC）-CM and HFF-CM. We found that MSCs and HFFs express all FGF receptors （FGFR1-4） and specifically produce TGF-β in response to bFGF. However, HFFs but not MSCs secrete IGF-Ⅱ. Despite the absence of IGF-Ⅱ in MSC-CM, hESC pluripotency and culture homeostasis were successfully maintained in MSC-CM for over 37 passages. Human ESCs derived on MSCs and hESCs maintained in MSC-CM retained hESC morphology, euploidy, expression of surface markers and transcription factors linked to pluripotency and displayed in vitro and in vivo multilineage developmental potential, suggesting that IGF-Ⅱ may be dispensable for hESC pluripotency. In fact, IGF-Ⅱ blocking had no effect on the homeostasis of hESC cultures maintained either on HFF-CM or on MSC-CM. These data indicate that hESCs are successfully maintained feeder-free with IGF-Ⅱ-lacking MSC-CM, and that the previously proposed paracrine mechanism by which bFGF cooperates with TGF-β and IGF-Ⅱ in the maintenance of hESCs in MEF-CM may not be fully extrapolated to hESCs maintained in CM from human MSCs.

Genomic imbalances in esophageal carcinoma cell lines involve Wnt pathway genes

AIM： To identify molecular markers shared across South African esophageal squamous cell carcinoma （ESCC） cell lines using o/togenetics, fluorescence in situ hybridization （FISH） and single nucleotide polymorphism （SNP） array copy number analysis. METHODS： We used conventional cytogenetics, FISH, and multicolor FISH to characterize the chromosomal rearrangements of five ESCC cell lines established in South Africa. The whole genome copy number profile was established from 250K SNP arrays, and data was analyzed with the CNAT 4.0 and GISTIC software. tions involved the following chromosomal regions and genes： 11q13.3 （CCND1, FGF3, FGF4, FGF19, MYEOV）, 8q24.21（C-MYC, FAM84B）, 11q22.1-q22.3 （B[RC2, BIRC3）, 5p15.2 （CTNND2）, 3qll.2-q12.2 （MINA） and 18p11.32 （TYMS, YES1）. The significant deletions included 1p31.2-p31.1 （CTH, GADD45a, DIRAS3）, 2q22.1 （LRPIB）, 3p12.1-p14.2 （FHIT）, 4q22.1-q32.1 （CASP6, SMAD1）, 8p23.2-q11.1 （BNIP3L） and 18q21.1-q21.2 （SMAD4, DCC）. The 3p11.2 translocation breakpoint was shared across four cell lines, supporting a role for genes involved at this site, in particular, the EPHA3 gene which has previously been reported to be deleted in ESCC.CONCLUSION： The finding that a significant number of genes that were amplified （FGF3, FGF4, FGF19, CCND1 and C-MYC） or deleted （SFRP2 gene） are involved in the Wnt and fibroblast growth factor signaling pathways, suggests that these pathways may be activated in these cell lines.

Lithocholic acid induction of the FGF19 promoter in intestinal cells is mediated by PXR

To study the effect of the toxic secondary bile acid lithocholic acid （LCA） on the expression of fibroblast growth factor 19 （FGF19） in intestinal cells and to characterize the pregnane-X-receptor （PXR） response of the FGF19 promoter region. METHODS： The intestinal cell line LS174T was stimulated with various concentrations of chenodeoxy- cholic acid and lithocholic acid for several time points. FGF19 mRNA levels were determined with quantitative realtime RT-PCR. FGF19 deletion promoter constructs were generated and the LCA response was analzyed in reporter assays. Co-transfections with PXR and RXR were carried out to study FGF19 regulation by these factors, RESULTS： LCA and CDCA strongly up-regulate FGF19 mRNA expression in LS174T cells in a time and dose dependent manner. Using reporter gene assays with several deletion constructs we found that the LCA responsive element in the human FGF19 promoter maps to the proximal regulatory region containing two poten- tial binding sites for PXR. Overexpression of PXR and its dimerization partner retinoid X receptor （RXR） and stimulation with LCA or the potent PXR ligand rifampicin leads to a significant induction of FGF19 promoter activ- ity in intestinal cells. CONCLUSION： LCA induced feedback inhibition of bile acid synthesis in the liver is likely to be regulated by PXR inducing intestinal FGF19 expression.

Fibroblast growth factor-4 and hepatocyte growth factor induce differentiation of human umbilical cord blood-derived mesenchymal stem cells into hepatocytes

AIM： To investigate the differentiation of human umbilical cord blood （HUCB）-derived mesenchymal stem cells （MSCs） into hepatocytes by induction of fibroblast growth factor-4 （FGF-4） and hepatocyte growth factor （HGF）, and to find a new source of cell types for therapies of hepatic diseases. METHODS： MSCs were isolated by combining gradient density centrifugation with plastic adherence. When HUCB-derived MSCs reached 70% confluence, they were cultured in Iscove modified Dulbecco medium （IMDM） supplemented with 10 mL/L FBS, 20 ng/mL HGF and 10 ng/mL FGF-4. The medium was changed every 4 d and stored for albumin, alpha-fetoprotein （AFP） and urea assay. Expression of CK-18 was detected by immunocytochemistry. Glycogen storage in hepatocytes was determined by PAS staining. RESULTS： By combining gradient density centrifugation with plastic adherence, we could isolate MSCs from 25.6% of human umbilical cord blood. When MSCs were cultured with FGF-4 and HGF, approximately 63.6% of cells became small, round and epithelioid on d 28 by morphology. Compared with the control, the level of AFP increased significantly from d 12 to 18.20：1=1.16 μg/L （t = 2.884, P〈0.05） in MSCs cultured with FGF-4 and HGF, and was higher （54.28±3.11 μg/L） on d 28 （t = 13.493, P〈0.01）. Albumin increased significantly on d 16 （t = 6.68, P〈0.01） to 1.02±0.15 μg/mL, and to 3.63±0.30 μg/mL on d 28 （t = 11.748, P〈0.01）. Urea （4.72±1.03 μmol/L） was detected on d 20 （t = 4.272, P〈0.01）, and continued to increase to 10.28±1.06 μmol/L on d 28 （t = 9.276, P〈0.01）. Cells expressed CK-18 on d 16. Glycogen storage was observed on d 24. CONCLUSION： HUCB-derived MSCs can differentiate into hepatocytes by induction of F-GF-4 and HGF. HUCB derived MSCs are a new source of cell types for cell transplantation therapy of hepatic diseases.

Establishment and Characterization of a New Cell Line from the Kidney of Spotted Halibut Verasper variegates

A cell line, SHK, was derived from the kidney of spotted halibut Verasper variegates. The cell line was subcultured more than 40 passages in minimum essential medium （MEM） supplemented with fetal bovine serum （FBS） and 10 ng ml4 basic fibroblast growth factor （bFGF）. Cell morphology from primary culture and subculture was observed continuously by microscopy. The SHK cell line consisted predominantly of fibroblast-like cells. The cell line was able to grow between 20℃ and 30℃ with the optimum growth at 24℃ and with a reduced growth between 12℃ and 20℃. The growth rate of the cells increased as the proportion of FBS increased from 10% to 20% at 28℃ with optimum growth at the concentration of 20%. The doubling time of the cells was determined to be 44.8 h. Chromosome analysis revealed that 52% of the SHK cells maintained a normal diploid chromosome number （2n=46）. The cells were successfully transfected with green fluorescent protein （GFP） reporter plasmids and the expression of GFP gene in the cells indicated the possible utility of the cells in gene expression studies. The cells were infected by lymphosystis disease virus （LCDV） and found to be susceptible to the virus in cytopathic effect （CPE） observation. The infection was confirmed by PCR and electron microscopy experiments, which proved the existence of the viral particles in the cytoplasm of the virus-infected cells.