Immunohistochemical assessment of angiogenesis in hepatocellular carcinoma and surrounding cirrhotic liver tissues

AIM: To investigate whether vascular endothelial growth factor (VEGF) was over-expressed in hepatocellular carcinoma (HCC) or in surrounding cirrhotic liver tissues.METHODS: Immunohistochemistry was performed to investigate the expression of VEGF proteins in HCC tissues from 105 consecutive patients undergoing curative resection for HCC. The immunostaining results and related clinicopathologic materials were analyzed with statistical methods. Kaplan-Meier method was used to calculate survival curves, and Log-rank test was performed to compare differences in survival rates of the patients with positive HCC staining and negative VEGF.RESULTS: VEGF-positive expression was found in 72 of105 HCC patients (68.6%). Capsular infiltration (P= 0.005),vascular invasion (P = 0.035) and intrahepatic metastasis(P=0.008) were observed more frequently in patients with VEGF-positive expression than in those with VEGFnegative expression. Kaplan-Meier curves showed that VEGF-positive expression was associated with a shorter overall survival (P = 0.014). VEGF-positive expression was found in 47 of tissues 68 HCC (69.1%), and VEGF-positive expression was found in 54 of 68 surrounding cirrhotic liver tissues (79.4%). VEGF-positive expression was significantly higher in surrounding cirrhotic liver tissues than in HCC (P= 0.017).CONCLUSION: VEGF may play an important role in the angiogenesis and prognosis of HCC, as well as in the angiogenesis of liver cirrhosis.

VEGF in hepatocellular carcinoma and surrounding cirrhotic liver tissues

We read with a great interest the recent work of Deli and colleagues. in the World Journal of Gastroenterology reporting vascular endothelial growth factor （VEGF） expression in hepatocellular carcinoma （HCC） and cirrhotic liver tissues. This well-documented work shows that VEGF was significantly higher in surrounding cirrhotic liver tissues than in HCC. Authors assessed VEGF expression using immunohistochemistry. The immunohistochemical staining is an efficient tool to assess the percentage of cells stained positively for VEGF but is not really efficient to estimate their true VEGF content. Evaluation of the VEGF protein by an enzyme-linked immunosorbent assay 0ELISA） has been reported, by us and others, to be an efficient tool in order to assess tissue VEGF expression. We have, thus, tested whether the ELISA method might be an efficient tool in order to confirm data reporting higher amounts of VEGF in surrounding cirrhotic liver tissues than in HCC. Deli and colleagues. also correctly pointed out that basic fibroblast growth factor （bFGF） has been reported to act cooperatively on VEGF expression. We have, thus, also assessed bFGF tissue levels in order to search for a putative link between VEGF and bFGF levels in cirrhotic tissues.

mPGES-1 expression in non-cancerous liver tissue impacts on postoperative recurrence of HCC

AIM:To investigate whether microsomal prostaglandin E synthase-1 (mPGES-1) expression in hepatocellular carcinoma (HCC) and in non-cancerous liver affects HCC prognosis after hepatectomy. METHODS: The relationship between patient clinical prof iles, tumor factors, surgical determinants, and mPGES-1 expression and the recurrence-free survival rate were examined in 64 patients who underwent curative hepatectomy between March 2003 and December 2006. RESULTS: The scores for mPGES-1 expression were higher in well differentiated and moderately differentiated HCC tissues than in poorly differentiated HCC tissues (well differentiated, 5.1 ± 2.7; moderately differentiated, 5.1 ± 1.7; poorly differentiated, 3.0 ± 1.8). In noncancerous liver tissues, the mPGES-1 levels were higher in injured liver tissues than in normal tissues. Cirrhotic livers had higher mPGES-1 levels than livers with chronic hepatitis (normal livers, 3.3 ± 0.7; chronic hepatitic livers, 5.4 ± 1.9; cirrhotic livers, 6.4 ± 1.6). A univariate analysis revealed that the recurrence-free survival rate was signif icantly lower in patients with vascular invasion,a higher mPGES-1 level in non-cancerous liver tissue,a larger tumor diameter (≥5 cm), and a lower serum albumin level (≤3.7 g/dL). The mPGES-1 expression in HCC tissues did not correlate well with postoperative recurrence. A multivariate analysis demonstrated that the presence of vascular invasion and higher mPGES-1 levels were statistically significant independent predictors for early postoperative recurrence of HCC.CONCLUSION: Increased mPGES-1 expression in noncancerous liver tissues is closely associated with the early recurrence of HCC after curative resection.

Construction and characterization of a cDNA library from human liver tissue with chronic hepatitis B

Objective: To construct a cDNA library from human liver tissue with chronic hepatitis B and check its quality for investigating the expression level of liver tissue infected by hepatitis B virus. This will then be used to find the relevant genes and interesting proteins associated with the development of hepatitis B. Methods: The total RNA from liver tissue with chronic hepa- titis B was extracted and the mRNA was purified using TRIZOL method. Switching mechanism at 5′ end of the RNA transcript (SMART) technique and CDS III/3′ primer were used for first-strand cDNA synthesis. Long distance polymerase chain reaction (LD PCR) was then used to synthesize the double-strand cDNA that was then digested by Sfi I and fractionated by CHROMA SPIN-400 column. The longer than 0.4 kb cDNAs were collected and ligated to λTriplEx2 vector. Then λ phage packaging reaction and library amplification were performed. The qualities of both unamplified and amplified cDNA libraries were strictly checked by conventional titer determination. Fourteen plaques were randomly picked and tested using PCR with universal primers derived from the sequence flanking the vector. Results: The titers of unamplifed and amplified libraries were 1.94×106 pfu/ml and 1.49×109 pfu/ml respectively. The percentages of recombinants from both libraries were 98.15% in unamplified library and 98.76% in amplified library. The lengths of the inserts were 1.23 kb in average, 1?2 kb in 64.29%, and 0.5?1.0 kb in 35.71%. Conclusion: A high quality cDNA library from human liver tissue with chronic hepatitis B was successfully constructed.

Hepatitis gene chip in detecting HBV DNA, HCV RNA in serum and liver tissue samples of hepatitis patients

OBJECTIVE: To study the preparation of diagnostic gene chip for detecting hepatitis B virus (HBV) and hepatitis C virus (HCV) and its accuracy in detecting HBV DNA and HCV RNA in serum and liver tissues. METHODS: The probes, which depend on the conservative gene fragment of hepatitis virus, was designed, synthesized and spotted on the modified glass. The probes and some other control probes were assembled on the diagnostic microarray of hepatitis virus. The gene of hepatitis virus, purified from blood or tissue, was labeled with fluorescence and hybridized to the microarray. The hybridized microarry was scanned with microarray scanner and the diagnostic result was analyzed from the scanning data. Fourty patients with hepatitis B virus and 40 healthy people or 40 patients with hepatitis C virus were subjected to detection of HBV DNA and HCV RNA with the hepatitis virus gene chip by the double-blind method. Paraffin liver specimens obtained from 99 cases of posthepatitic cirrhosis were used to detect HBV DNA. The liver tissues and serum from 15 cases of chronic hepatitis B were used to detect HBV DNA. Simultaneously, HBsAg and HBcAg were detected in the serum by fluorescence microparticle quantitation, HBV DNA and HCV RNA in the serum by PCR, and HBcAg in liver tissues by immunocytochemistry or HBV DNA by in situ molecular hybridization. RESULTS: Chip detection of serum specimens showed that 30 patients were HBV DNA positive and 10 HBV DNA negative in the 40 patients with HBV positive, 25 patients were HCV RNA positive and 15 patients were HCV RNA negative in the 40 patients with HCV positive, and all were HBV and HCV negative in the 40 healthy people. In 15 patients with HBV marker positive who were subjected to liver biopsy, 15 patients were detected HBV DNA positive in serum by gene chip, 15 patients HBcAg positive in liver tissues by immunocytochemistry, 14 patients HBV DNA positive in liver tissues by in situ molecular hybridization, and 14 patients HBV DNA positive in liver tissues by gene chip. Paraffin liver tissues specimens from the 99 patients with posthepatitis B cirrhosis showed that 67 patients were detected HBcAg positive by immunocytochemistry, 53 patients HBV DNA positive by in situ molecular hybridization, and 46 patients HBV DNA positive by gene chip. In the 46 patients, 40 patients were detected HBV DNA and HBcAg positive by in situ molecular hybridization and immunocytochemistry, 6 patients only HBcAg positive, and 33 patients HBcAg negative. CONCLUSIONS: The designed diagnostic gene chip can be used to simultaneously detect serum HBV DNA and HCV RNA, but the positive rate of HCV RNA diagnosed by this chip is lower. The gene chip can detect HBV DNA in serum and in liver tissue.

Ectopic liver tissue (choristoma) on the gallbladder: Acomprehensive literature review

BACKGROUND Liver tissue situated outside the liver with a hepatic connection is usually calledan accessory liver, and that without a connection to the mother liver, is calledectopic liver tissue.AIM To identify studies in the literature on ectopic liver tissue located on thegallbladder surface or mesentery.METHODS We present two patients and review published articles on ectopic liver tissuelocated on the gallbladder surface accessed via PubMed, MEDLINE, GoogleScholar, and Google databases. Keywords used included accessory liver lobe,aberrant liver tissue, ectopic liver tissue, ectopic liver nodule, heterotopic livertissue, hepatic choristoma, heterotopic liver tissue on the gallbladder, and ectopicliver tissue on the gallbladder. The search included articles published before June2020 with no language restriction. Letters to the editor, case reports, reviewarticles, original articles, and meeting presentations were included in the search.Articles or abstracts containing adequate information on age, sex, history of liverdisease, preliminary diagnosis, radiologic tools, lesion size, surgical indication,surgical procedure, and histopathological features of ectopic liver tissue wereincluded in the study.RESULTS A total of 72 articles involving 91 cases of ectopic liver tissue located on the gallbladder surface or mesentery were analyzed. Of these 91 patients, 62 werefemale and 25 were male (no gender available for 4 patients), and the age rangewas 5 d to 91 years. Forty-nine patients underwent surgery for chroniccholecystitis or cholelithiasis, and 14 patients underwent surgery for acutecholecystitis. The remaining 28 patients underwent laparotomy for other reasons.Cholecystectomy was laparoscopic in 69 patients and open in 11 patients. Theremaining 19 patients underwent various other surgical procedures such asautopsy, liver transplantation, living donor hepatectomy, Whipple procedure, andliver segment V resection. Histopathologically, hepatocellular carcinoma wasdetected in the ectopic liver tissue of one patient.CONCLUSION Ectopic liver tissue is a rare developmental anomaly which is usually detectedincidentally. Although most studies suggest that ectopic liver located outside thegallbladder has a high risk of hepatocellular carcinoma, this is not reflected instatistical analysis.

Raman and UV-Vis Spectroscopy Applied to the Analysis of Liver Tissues from Rats with Myocardial Ischemia Induced by Isoproterenol

The application of the laser Raman spectroscopic（LRS） technique for the analysis of liver tissues from rats with myocardial ischemia induced by isoproterenol（ISO） was described.Animal model of myocardial ischemia was established for rats induced by ISO.Rats were randomly divided into four groups as normal group and myocardial ischemia groups.We observed the successful myocardial ischemia model via serum enzymes levels and hematoxylin-eosin（HE） staining,and detected the liver tissue of the rats from normal group and liver tissue of the rats from myocardial ischemia groups via UV-Vis spectroscopy（UV-Vis） and LRS,and the changes of the absorbance spectra were compared in the above four different groups.The results show that ISO can induce rat myocardial ischemia successfully.The spectrum of normal liver tissue supernatant exhibits a strong absorption band at 968 nm,but no absorption band appears in the spectra of liver tissue supernatant solutions from the rats with myocardial ischemia induction after 2,12 and 72 h presented at 968 nm.LRS results show that Raman intensities of the precipitates suffered from ISO-treatment after 2,12 and 72 h were obviously increased compared with that of the precipitate of the liver tissue of the normal rats suffered from 0.9 g/L normal saline（NS） treatment.These results indicate that LRS and UV-Vis can be harmless,nondestructive,rapid and effective methods for analyzing different pathological specimens of liver tissue from myocardial ischemia rats.

Ultrasonic characterization of porcine liver tissue at frequency between 25 to 55 MHz

AIM： To study the relation between acoustic parameters and histological structure of biological tissue and to provide the basis for high-resolution image of biological tissues and quantitative ultrasonic diagnosis of liver disease. METHODS： Ultrasonic imaging and tissue characterization of four normal porcine liver and five cirrhotic liver tissue samples were performed using a high frequency imaging system. RESULTS： The acoustic parameters of cirrhotic liver tissue were larger than those of normal liver tissue. The sound velocity was 1577 m/s in normal liver tissue and 1631 m/s in cirrhotic liver tissue. At 35 MHz, the attenuation coefficient was 3.0 dB/mm in normal liver tissue and 4.1 dB/mm in cirrhotic liver tissue. The backscatter coefficient was 0.00431 dB/Srmm in cirrhotic liver tissue and 0.00303 dB/Srmm in normal liver tissue. The backscatter coefficient increased with the frequency. The high frequency images coincided with their histological features. CONCLUSION： The acoustic parameters, especially the sound backscatter coefficient, are sensitive to the changes of liver tissues and can be used to differentiate between the normal and pathological liver tissues. High frequency image system is a useful device for highresolution image and tissue characterization.

Joint Effects of Mexidol and Nitroglycerine on Nitric Oxide Formation in Animal Liver Tissues

This work is investigating Mexidol (2-ethyl-6-methyl-3-hydroxy pyridine succinate) effect on the formation of nitric oxide (NO) in animal liver tissues, which is a regulator of many physiological processes and plays an important role in the vascular relaxation, neurotransmission and immune system functioning. Analyses performed by EPR spectroscopy revealed Hem-NO complex signals from paramagnetic centers in arbitrary units;produced nitrogen oxide amount in liver tissues was determined by method of double integration signals from nitrosyl complexes.

Endoscopic ultrasound-guided tissue acquisition for the diagnosis of focal liver lesion

In patients with liver tumors,the histopathology examination can assist in diagnosis,staging,prognosis,and therapeutic management strategy.Endoscopic ultrasound(EUS)-guided tissue acquisition using fine needle aspiration(FNA)or more newly fine needle biopsy(FNB)is a well-developed technique in order to evaluate and differentiate the liver masses.The goal of the EUS-FNA or EUS-FNB is to provide an accurate sample for a histopathology examination.Therefore,malignant tumors such as hepatocarcinoma,cholangiocarcinoma and liver metastasis or benign tumors such as liver adenoma,focal hyperplastic nodular tumors and cystic lesions can be accurately diagnosed using EUS-guided tissue acquisition.EUS-FNB using 19 or 22 Ga needle provide longer samples and a higher diagnostic accuracy in patients with liver masses when compared with EUS-FNA.Few data are available on the diagnostic accuracy of EUS-FNB when compared with percutaneously,ultrasound,computer tomography or transjugulary-guided liver biopsies.This review will discuss the EUS-guided tissue acquisition options in patients with liver tumors and its efficacy and safety in providing accurate samples.The results of the last studies comparing EUS-guided liver biopsy with other conventional techniques are presented.The EUS-guided tissue acquisition using FNB can be a suitable technique in suspected liver lesions in order to provide an accurate histopathology diagnosis,especially for those who require endoscopy.

Innovations in 3D bioprinting and biomaterials for liver tissue engineering:Paving the way for tissue-engineered liver

The liver is a pivotal organ that maintains internal homeostasis and actively participates in multiple physiological processes.Liver tissue engineering(LTE),by which in vitro biomimetic liver models are constructed,serves as a platform for disease research,drug screening,and cell replacement therapies.3D bioprinting is used in tissue engineering to create microenvironments that closely mimic authentic tissues with carefully selected functional biomaterials.Ideal functional biomaterials exhibit characteristics such as high biocompatibility,mechanical strength,flexibility,processability,and tunable degradability.Biomaterials can be categorized into natural and synthetic biomaterials,each with its own advantages and limitations,and their combinations serve as a primary source of 3D bioprinting materials.It is noteworthy that the liver decellularized extracellular matrix(dECM),obtained by removing cellular components from tissues,possesses traits such as bioactivity,biocompatibility,and non-immunogenicity,making it a common choice among functional biomaterials.Furthermore,crosslinking of biomaterials significantly impacts the mechanical strength,physicochemical properties,and cellular behavior of the printed structures.This review covers the current utilization of biomaterials in LTE,focusing on natural and synthetic biomaterials as well as the selection and application of crosslinking methods.The aim is to enhance the fidelity of in vitro liver tissue models by providing a comprehensive coverage of functional biomaterials,thereby establishing a versatile platform for tissue-engineered livers.

Con A affinity glycoproteomics of normal human liver tissue

In order to establish the novel high throughput, high efficiency and low cost technological platform for the research of N-glycoproteomics, to resolve the significance of characteristic expression profile of glycoprotein and to find the proteins with biological functional importance, the glycoproteins with high-mannose core and the two antennary types were purified and enriched by the Con A affinity chromatography. Con A affinity protein expression profiles of normal human liver tissue were gener- ated by using SDS-PAGE, two-dimensional electrophoresis (2-DE) followed by fast fluorescence stain- ing based on multiplexed proteomics (MP) technology. 301 visible protein spots on the gel were de- tected and 85 of glycoproteins were further successfully identified via peptide mass fingerprinting (PMF) by a matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF- MS/MS) and annotated to IPI databases. Identified glycoproteins definitely take part in the regulation of cell cycle and metabolic processes. The glycosylation sites were predicted with NetNGlyc 1.0 and NetOGlyc 3.1 software, meanwhile they were classified according to the geneontology methods. The construction of Con A affinity glycoprotein database of normal human liver tissue would contribute to the subsequent research.

Polyethylene glycol crosslinked decellularized single liver lobe scaffolds with vascular endothelial growth factor promotes angiogenesis in vivo

Background: Improving the mechanical properties and angiogenesis of acellular scaffolds before transplantation is an important challenge facing the development of acellular liver grafts. The present study aimed to evaluate the cytotoxicity and angiogenesis of polyethylene glycol(PEG) crosslinked decellularized single liver lobe scaffolds(DLSs), and establish its suitability as a graft for long-term liver tissue engineering. Methods: Using mercaptoacrylate produced by the Michael addition reaction, DLSs were first modified using N-succinimidyl S-acetylthioacetate(SATA), followed by cross-linking with PEG as well as vascular endothelial growth factor(VEGF). The optimal concentration of agents and time of the individual steps were identified in this procedure through biomechanical testing and morphological analysis. Subsequently, human umbilical vein endothelial cells(HUVECs) were seeded on the PEG crosslinked scaffolds to detect the proliferation and viability of cells. The scaffolds were then transplanted into the subcutaneous tissue of Sprague-Dawley rats to evaluate angiogenesis. In addition, the average number of blood vessels was evaluated in the grafts with or without PEG at days 7, 14, and 21 after implantation. Results: The PEG crosslinked DLS maintained their three-dimensional structure and were more translucent after decellularization than native DLS, which presented a denser and more porous network structure. The results for Young’s modulus proved that the mechanical properties of 0.5 PEG crosslinked DLS were the best and close to that of native livers. The PEG-VEGF-DLS could better promote cell proliferation and differentiation of HUVECs compared with the groups without PEG cross-linking. Importantly, the average density of blood vessels was higher in the PEG-VEGF-DLS than that in other groups at days 7, 14, and 21 after implantation in vivo. Conclusions: The PEG crosslinked DLS with VEGF could improve the biomechanical properties of native DLS, and most importantly, their lack of cytotoxicity provides a new route to promote the proliferation of cells in vitro and angiogenesis in vivo in liver tissue engineering.

IMMUNOHISTOCHEMICAL DEMONSTRATION OF CCAAT/ENHANCER BINDING PROTEIN (C/EBP) IN HUMAN LIVER TISSUES OF VARIOUS ORIGIN

C / EBP is a sequence-specific DNA-binding protein. In order to indentify its distribution and localization, immunohistochemical technique (ABC method) was done using anti-C / EBP polypeptide antibodies 1103#, 425# in liver specimens from 20 normal adults, 5 neonates, 6 patients with hepatitis, 25 with liver cirrhosis, 80 with hepatocellular carcinoma (40 cases were associated with surrounding nontumorous tissues) and 26 patients with cholangiocarcinoma (15 cases were associated with surrounding nontumorous tissues). The results showed that C / EBP was diffusely distributed in nuclei and cytoplasm of differentiated liver cells and very low or undetectable in liver cancer cells. The manifestation of C / EBP correlated with degree of differentiation of tumour cells, and was obviously weaker than that in surrounding nontumorous tussues. C / EBP positive staining has also been found in regenerating epithelial cells of bile ductules. The results suggested that C / EBP should play an important role in establishing and maintaining the differentiation of liver cells.

Direct Measurement of Oxygen Free Radicals of the Liver Tissue Following Shock/Reperfusion and Its Significance

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Ultrasonic spectrum characterization of pig's pathological liver tissues

A high-speed digital acquisition and wide-band measurement system was used to acquiring and recording the ultrasonic pulse waveforms backscattered from normal and several pathological pig's fresh liver tissues (including ballooning degeneration, fatty degeneration, cirrhosis) in vitro. Some frequency-domain analyses were performed in which integrated backscattering coefficient (IBSC), frequency dependence of backscattering coefficient (FDBSC),and mean spacing (MS) among adjacent scatterers of the tissues were determined.In the experiments the frequency domain was from 3MHz to 7 MHz. The obtained results show that the differences among normal and various pathological tissues cause distinct changes of the measured parameters. The results show that the ultrasonic spectum technique, which can be used in tissue characterization for pathological liver in clinical, has an exciting prospect.

Liver regeneration using decellularized splenic scaffold: a novel approach in tissue engineering

BACKGROUND： The potential application of decellularized liver scaffold for liver regeneration is limited by severe shortage of donor organs. Attempt of using heterograft scaffold is accompanied with high risks of zoonosis and immunological rejection. We proposed that the spleen, which procured more extensively than the liver, could be an ideal source of decellularized scaffold for liver regeneration. METHODS： After harvested from donor rat, the spleen was processed by 12-hour freezing/thawing ×2 cycles, then circulation perfusion of 0.02% trypsin and 3% Triton X-100 sequentially through the splenic artery for 32 hours in total to prepare decellularized scaffold. The structure and component characteristics of the scaffold were determined by hematoxylin and eosin and immumohistochemical staining, scanning electron microscope, DNA detection, porosity measurement, biocompatibility and cytocompatibility test. Recellularization of scaffold by 5×106 bone marrow mesenchymal stem cells（BMSCs） was carried out to preliminarily evaluate the feasibility of liver regeneration by BMSCs reseeding and differentiation in decellularized splenic scaffold.RESULTS： After decellularization, a translucent scaffold, which retained the gross shape of the spleen, was generated. Histological evaluation and residual DNA quantitation revealed the remaining of extracellular matrix without nucleus and cytoplasm residue. Immunohistochemical study proved the existence of collagens I, IV, fibronectin, laminin and elastin in decellularized splenic scaffold, which showed a similarity with decellularized liver. A scanning electron microscope presented the remaining three-dimensional porous structure of extracellular matrix and small blood vessels. The poros-ity of scaffold, aperture of 45.36±4.87 μm and pore rate of 80.14%±2.99% was suitable for cell engraftment. Subcutaneous implantation of decellularized scaffold presented good histocompatibility, and recellularization of the splenic scaffold demonstrated that BMSCs could locate and survive in the decellularized matrix. CONCLUSION： Considering the more extensive organ source and satisfying biocompatibility, the present study indicated that the three-dimensional decellularized splenic scaffold might have considerable potential for liver regeneration when combined with BMSCs reseeding and differentiation.

Effects of regenerated tissue extracts after liver injury on the proliferation,differentiation,migration and invasion of SK-HEP1 cells

Objective: To study the effects of regenerated tissue extracts after liver injury on the proliferation, differentiation, migration and invasion of SK-HEP1 cells. Methods: Regenerated tissue extracts after liver injury were used to induce SK-HEP1 cells after enrichment, their effects on the proliferation, differentiation, migration and invasion of SK-HEPI cells were observed through in vitro cell culture, MTT, flow cytometry and transwell assays. Results:In response to the action of regenerated tissue extracts after liver injury, SK-HEP1 cells were blocked in G_0/G_1 phase, their growth rate was distinctly reduced. The number of SK-HEP1^(-fj)colonies decreased. The migration ability of SK-HEPI cells showed a decreased trend on day7 and day 11 after induction. SK-HEPl's invasion ability clearly decreased on days 7 and11 after induction, especially on day 7. Conclusions: To a certain extent, regenerated tissue extracts after liver injury can inhibit the proliferation, differentiation, migration and invasion of hepatoma cells, showing an important potential of being a differentiating agent for the treatment of liver cancer.

A NEW METHOD OF ISOLATING KUPFFER CELLS FROM BIOPSY TISSUE OF RAT LIVER

The isolation of a high yield and purity of Kupffer cells has been reported in detail.1 This paper reports into the research about isolation Kupffer cells from biopsy tissue of liver. This method includes 5 important steps: (1) take fresh liver tissue, and mince with scissors. (2) spin at low speed to wash off red blood cells. (3) digest in collagenase for suitable time. (4) isolate Kupffer cells on a percoll density gradient. (5) cell charaterization was observed by N.S.E stain and peroxidatic activity with lumino-meter measurement and phagocytosis with latex beads.2.3

Green Electrospun Silk Fibroin/Galactose Chitosan Composite Nanofibrous Scaffolds for Hepatic Tissue Engineering

The electrospun nanofibrous scaffolds made of proteins and polysaccharides were thought to be able to simulate the structure of natural extracellular matrix well.Silk fibroin(SF)and chitosan(CS)are probably the most widely used natural materials in biomedical fields including liver tissue engineering for their good properties and wide variety of sources.The asialoglycoprotein receptors of hepatocyte were reported to specifically recognize and interact with galactose.In this work,a green electrospun SF/galactosylated chitosan(GC)composite nanofibrous scaffold was fabricated and characterized.The data indicated that the addition of GC greatly influenced the spinning effect of SF aqueous solution,and the average diameter of the composite nanofibers was about 520nm.Moreover,the green electrospun SF/GC nanofibrous scaffolds were demonstrated significantly enhancing the adhesion and proliferation of hepatocyte(RH35)according to our data.The present study did a useful exploration on constructing scaffolds for liver regeneration by green electrospinning,and also laid a good foundation for the further applicative research of this green electrospun scaffolds in liver tissue engineering.