[Retracted] Resveratrol reduces acute lung injury in a LPS­induced sepsis mouse model via activation of Sirt1.

Following the publication of this paper, it was drawn to the Editor's attention by a concerned reader that the ß­actin control western blotting data shown in Fig. 3D on p. 1893 were very similar to the contol data shown in Fig. 4A on p. 1894; furthermore, the data shown for the MMP­9 and the INOS protein bands in Fig. 4C were remarkably similar to the data shown for the IL­1ß and IL­6 proteins, respectively, albeit the backgrounds surrounding the bands were different. Moreover, various of the western blotting data shown in these figures were strikingly similar to data that had already been published in different form in other articles written by (largely) different authors at different research institutes. Owing to the fact that the contentious data in the above article had already been published prior to its submission to Molecular Medicine Reports, and due to the number of apparent duplications of strikingly similar data between Figs. 3 and 4, the Editor has decided that this paper should be retracted from the Journal. The authors were asked for an explanation to account for these concerns, but the Editorial Office did not receive a satisfactory reply. The Editor apologizes to the readership for any inconvenience caused. [Molecular Medicine Reports 7: 1889­1895, 2013; DOI: 10.3892/mmr.2013.1444].

Epigenetic Inheritance From Normal Origin Cells Can Determine the Aggressive Biology of Tumor-Initiating Cells and Tumor Heterogeneity.

The acquisition of genetic- and epigenetic-abnormalities during transformation has been recognized as the two fundamental factors that lead to tumorigenesis and determine the aggressive biology of tumor cells. However, there is a regularity that tumors derived from less-differentiated normal origin cells (NOCs) usually have a higher risk of vascular involvement, lymphatic and distant metastasis, which can be observed in both lymphohematopoietic malignancies and somatic cancers. Obviously, the hypothesis of genetic- and epigenetic-abnormalities is not sufficient to explain how the linear relationship between the cellular origin and the biological behavior of tumors is formed, because the cell origin of tumor is an independent factor related to tumor biology. In a given system, tumors can originate from multiple cell types, and tumor-initiating cells (TICs) can be mapped to different differentiation hierarchies of normal stem cells, suggesting that the heterogeneity of the origin of TICs is not completely chaotic. TIC's epigenome includes not only genetic- and epigenetic-abnormalities, but also established epigenetic status of genes inherited from NOCs. In reviewing previous studies, we found much evidence supporting that the status of many tumor-related "epigenetic abnormalities" in TICs is consistent with that of the corresponding NOC of the same differentiation hierarchy, suggesting that they may not be true epigenetic abnormalities. So, we speculate that the established statuses of genes that control NOC's migration, adhesion and colonization capabilities, cell-cycle quiescence, expression of drug transporters, induction of mesenchymal formation, overexpression of telomerase, and preference for glycolysis can be inherited to TICs through epigenetic memory and be manifested as their aggressive biology. TICs of different origins can maintain different degrees of innate stemness from NOC, which may explain why malignancies with stem cell phenotypes are usually more aggressive.

Re-Recognizing the Cellular Origin of the Primary Epithelial Tumors of the Liver.

The primary epithelial tumors of the liver (PETL) are composed of a series of heterogeneous tumors. Although the classification of PETLs has been updated several times by the World Health Organization, the cellular origins of some tumors in this family remain to be precisely depicted. In addition, certain tumors in different categories have similar histology, molecular phenotypes and biological characteristics, suggesting that they may have the same cellular origin. In this work, a narrative review method was adopted to review the relevant papers. By comparing the expression profiles of biomarkers of liver epithelium at different lineages and stages of differentiation, the cells-of-origin of some major members of the PETL family were reassessed. We propose that 1) hepatic adenomas, hepatocellular carcinomas (HCCs) and pure fetal hepatoblastomas (HBs) share the same spectrum in their cellular origin including the hepatocytic-committed progenitors (HCP) and their differentiated descendants. 2) Bile duct adenomas, peribiliary cysts and intrahepatic cholangiocellular carcinomas (ICCs) can share the same spectrum in their cellular origin including the cholangiocytic-committed progenitors (CCP) and their differentiated descendants. 3) The cells-of-origin of embryonal HBs include liver stem cells (LSCs), hepatoblasts, and transitional cells between them. Embryonal HB with small cell element, small cell undifferentiated HB and small cell neuroendocrine carcinoma of the liver can have the same or similar cells-of-origin from LSC. Embryonal HB lacking the small cell component of the LSC phenotype and presenting both hepatocytic and bile duct/ductule components may originate from actual hepatoblasts/hepatic progenitor cells (HPCs) as the combined HCC-ICC does. 4) Teratoid hepatoblastoma and mixed epithelial/mesenchymal HBs can be derived from the LSCs or even less committed extrahepatic pluripotent stem cell. 5) Many members of the PETLs family, including those derived from LSCs, hepatoblasts/HPCs, early HCPs and CCPs, have neuroendocrine potentiality. Except for those primary hepatic neuroendocrine tumor (PHNET) exhibit hepatocytic and/or cholangiocytic phenotypes, other PHNETs subtype may be derived from the descendants of LSC that differentiate towards the upper digestive tract, pancreas or other lineages.

Prediction of Early Recurrence of Solitary Hepatocellular Carcinoma after Orthotopic Liver Transplantation.

Hepatocellular carcinomas(HCC) consisted of heterogeneous subtypes with different recurrence probabilities after liver transplantation(LT). Our study aimed to develop an improved model for predicting the recurrence of solitary HCC after LT. In this retrospective study, 151 solitary HCC patients who received orthotopic LT over a period of 10 consecutive years were included. All recipients received graft from deceased donors. The first eligible 50 patients were used as validation cohort and others were utilized to construct the model. A two-tailed P < 0.05 was considered to indicate statistical significance for all analysis. Based on the maximisation of the Youden's index, the optimal cutoff values for alpha-fetoprotein(AFP) and tumor diameter were 261.6 ng/mL and 3.6 cm, respectively. Vascular involvement includes gross and microscopic vascular invasion. Variables potentially affecting recurrence-free survival(RFS) were examined using univariate and multivariate Cox regression analysis. Univariate and multivariate analysis revealed that AFP, tumor diameter, vascular invasion and cytokeratin-19/glypican-3 sub-typing were independent prognostic factors for RFS, thus comprised the risk scoring model. The AUC values of the model in the cohorts were significantly higher than that of the Milan, UCSF, Fudan and Hangzhou criteria. These findings suggest the model has high performance in predicting early recurrence of solitary HCC patients after LT.

Histopathological characteristics of needle core biopsy and surgical specimens from patients with solitary hepatocellular carcinoma or intrahepatic cholangiocarcinoma.

BACKGROUND: Pathological manifestations of hepatic tumours are often associated with prognosis. Although surgical specimens (SS) can provide more information, currently, pre-treatment needle core biopsy (NCB) is increasingly showing important value in understanding the nature of liver tumors and even in diagnosis and treatment decisions. However, the concordance of the clinicopathological characteristics and immunohistochemical (IHC) staining between NCB and SS from patients with hepatic tumours were less concerned. AIM: To introduce a more accurate method for interpreting the IHC staining results in order to improve the diagnostic value of hepatic malignancy in NCB samples. METHOD: A total of 208 patients who underwent both preoperative NCB and surgical resection for hepatocellular carcinoma (HCC) or intrahepatic cholangiocarcinoma (ICC) between 2008 and 2015 were enrolled in this study. The expression of CK19, GPC3, and HepPar1 were detected by IHC staining. Clinicopathological, NCB, and surgical data were collected and analysed using χ 2 and kappa statistics. RESULTS: Morphologically, the presence of compact tumour nests or a cord-like structure in NCB was considered the primary cause of misdiagnosis of HCC from ICC. The kappa statistic showed a moderate agreement in histomorphology (k = 0.504) and histological grade (k = 0.488) between NCB and SS of the tumours. A 4-tier (+++, ++, +, and -) scoring scheme that emphasized the focal neoplastic cell immunoreactivity of tumour cells revealed perfect concordance of CK19, GPC3 and HepPar1 between NCB and SS (k = 0.717; k = 0.768; k = 0.633). Furthermore, with the aid of a binary classification derived from the 4-tier score, a high concordance was achieved in interpreting the IHC staining of the three markers between NCB and final SS (k = 0.931; k = 0.907; k = 0.803), increasing the accuracy of NCB diagnosis C (k = 0.987; area under the curve = 0.997, 95%CI: 0.990-1.000; P < 0.001). CONCLUSION: These findings imply that reasonable interpretation of IHC results in NCB is vital for improving the accuracy of tumour diagnosis. The simplified binary classification provides an easy and applicable approach.

Prediction early recurrence of hepatocellular carcinoma eligible for curative ablation using a Radiomics nomogram.

BACKGROUND: Predicting early recurrence (ER) after radical therapy for HCC patients is critical for the decision of subsequent follow-up and treatment. Radiomic features derived from the medical imaging show great potential to predict prognosis. Here we aim to develop and validate a radiomics nomogram that could predict ER after curative ablation. METHODS: Total 184 HCC patients treated from August 2007 to August 2014 were included in the study and were divided into the training (n = 129) and validation(n = 55) cohorts randomly. The endpoint was recurrence free survival (RFS). A set of 647 radiomics features were extracted from the 3 phases contrast enhanced computed tomography (CECT) images. The minimum redundancy maximum relevance algorithm (MRMRA) was used for feature selection. The least absolute shrinkage and selection operator (LASSO) Cox regression model was used to build a radiomics signature. Recurrence prediction models were built using clinicopathological factors and radiomics signature, and a prognostic nomogram was developed and validated by calibration. RESULTS: Among the four radiomics models, the portal venous phase model obtained the best performance in the validation subgroup (C-index = 0.736 (95%CI:0.726-0.856)). When adding the clinicopathological factors to the models, the portal venous phase combined model also yielded the best predictive performance for training (C-index = 0.792(95%CI:0.727-0.857) and validation (C-index = 0.755(95%CI:0.651-0.860) subgroup. The combined model indicated a more distinct improvement of predictive power than the simple clinical model (ANOVA, P < 0.0001). CONCLUSIONS: This study successfully built a radiomics nomogram that integrated clinicopathological and radiomics features, which can be potentially used to predict ER after curative ablation for HCC patients.

Prediction of early recurrence of hepatocellular carcinoma within the Milan criteria after radical resection.

Approximately 50% hepatocellular carcinoma patients meeting the Milan criteria utilized to develop an improved prognostic model for predicting the recurrence in these patients. Using univariate and multivariate analysis, cytokeratin-19 and glypican-3 expression patterns, tumor number and histological grading from eight putative prognostic factors comprised the risk factor scoring model to predict the tumor recurrence. In the training cohort, the area under roc curve (AUC) value of the model was 0.715 [95% confidence interval (CI) = 0.645-0.786, P<0.001], which was the highest among all the parameters. The performance of the model was assessed using an independent validation cohort, wherein the AUC value was 0.760 (95% CI=0.647-0.874, P<0.001), which was higher than the other factors. The results indicated that model had high performance with adequate discrimination ability. Moreover, it significantly improved the predictive capacity for the recurrence in patients with hepatocellular carcinoma within the Milan criteria after radical resection.

Simple Risk Score for Prediction of Early Recurrence of Hepatocellular Carcinoma within the Milan Criteria after Orthotopic Liver Transplantation.

Ten to twenty percent of the hepatocellular carcinoma (HCC) patients fulfilling the Milan criteria (MC) recurred within three years after orthotopic liver transplantation (OLT). We therefore utilize a training cohort to develop an improved prognostic model for predicting the recurrence in these patients. By univariate and multivariate analysis, AFP level [cut-off value: 321 ng/mL, area under the curve (AUC) = 0.724, 95% confidence interval (CI) = 0.604-0.843, P < 0.001] and cytokeratin-19 (CK19) and glypican-3 (GPC3) expression pattern from nine putative prognostic factors were entered in risk factor scoring model to conjecture the tumor recurrence. In the training cohort, the AUC value of the model was 0.767 (95% CI = 0.645-0.890, P < 0.001), which was the highest among all the elements. The model's performance was then assessed using a validation cohort. In the validation cohort, the AUC value of the model was 0.843 (95% CI = 0.720-0.966, P < 0.001) which was higher than any other elements. The results indicated that model had high performance with good discrimination ability and significantly improved the predictive capacity for the recurrence of HCC patients within MC after OLT.

The significance of glypican-3 expression profiling in the tumor cellular origin theoretical system for hepatocellular carcinoma progression.

BACKGROUND AND AIM: Glypican-3 (GPC3) expression is correlated with poor prognosis and progression in hepatocellular carcinoma (HCC). HCC progression can be associated with the differentiation status of tumor cell before malignant transformation. Our aim was to investigate the dynamic expression of GPC3 during tumor cells differentiation and to explore the role and theoretical significance of GPC3 in malignant essence of HCC. METHODS: The expressions of tissue GPC3 and alpha fetoprotein (AFP) were detected by immunohistochemical staining. The tumor size, lymph node involvement, and metastasis were determined by pathological and imaging studies. HepG2 cells were induced to differentiate by all-trans retinoic acid (ATRA). Differentiation was evaluated by cytokeratin 19, gamma glutamyl transferase, and AFP through reverse transcription-polymerase chain reaction and real-time polymerase chain reaction. GPC3 staining was analyzed through flow cytometry. RESULTS: Based on the immunohistochemical staining, the enrolled 316 cases were divided into two subtypes, namely, GPC3+ HCC and GPC3- HCC. Significant differences in morphology, histology variations, AFP expression, TNM staging, and overall survival curves were observed between two subtypes. During HCC differentiation induced by ATRA, the mean value of GPC3 expression treated with ATRA was much lower than the ones in placebo. There were significant differences between GPC3+ HCC and GPC3- HCC for cumulative intrahepatic and extrahepatic recurrence in early stage HCC (P = 0.009, P = 0.010). CONCLUSIONS: Glypican-3 is correlated with the clinical malignant behavior of HCC. Moreover, GPC3 phenotype changes from positive to negative during tumor cells differentiation. Meanwhile, GPC3 plays a significant role in tumor cellular origin theoretical system, which can better reflect the malignant essence of tumors.

Worse or better?-Cirrhosis with hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) accounts for about 90% of all malignant tumors of liver, ranking fifth in the worldwide incidence of malignant tumors and the third in fatality. More and more evidences suggest that cancer is a metabolic-related disease. From the analysis of recent clinical research data, we found that as the severity of the cirrhosis aggravated, patients with HCC and end-stage liver cirrhosis had a flat energy metabolism which was better than it in patients with simple end-stage liver cirrhosis. Based on these clinical phenomenon, the major aim of this study is to present a new hypothesis: "compensated liver function mechanism" for patients with HCC and liver cirrhosis, cancer cells may play a role to compensate liver function. In this study, we elaborated relevant content about this novel standpoint combined with tumor energy metabolism reprogramming mechanism and tumor cell origin as well as cell exchange mechanism.

CK19 and Glypican 3 Expression Profiling in the Prognostic Indication for Patients with HCC after Surgical Resection.

This retrospective study was designed to investigate the correlation between a novel immunosubtyping method for hepatocellular carcinoma (HCC) and biological behavior of tumor cells. A series of 346 patients, who received hepatectomy at two surgical centers from January 2007 to October 2010, were enrolled in this study. The expressions of cytokeratin 19 (CK19), glypican 3 (GPC3), and CD34 were detected by immunohistochemical staining. The clinical stage was assessed using the sixth edition tumor-node-metastasis (TNM) system (UICC/AJCC, 2010).Vascular invasion comprised both microscopic and macroscopic invasion. The tumor size, lymph node involvement, and metastasis were determined by pathological as well as imaging studies. Recurrence was defined as the appearance of new lesions with radiological features typical of HCC, seen by at least two imaging methods. Survival curves for the patients were plotted using the Kaplan-Meier method, and differences between the curves were assessed using the log-rank test. Significant differences in morphology, histological grading, and TNM staging were observed between groups. Based on the immunohistochemical staining, the enrolled cases were divided into CK19+/GPC3+, CK19-/GPC3+ and CK19-/GPC3- three subtypes. CK19+/GPC3+ HCC has the highest risk of multifocality, microvascular invasion, regional lymph node involvement, and distant metastasis, followed by CK19-/GPC3+ HCC, then CK19-/GPC3-HCC. CK19+/GPC3+ HCC has the shortest recurrence time compared to other immunophenotype HCCs. CK19 and GPC3 expression profiling is an independent prognostic indicator in patients with HCC, and a larger sample size is needed to further investigate the effect of this immunosubtyping model in stratifying the outcome of HCC patients.

Pathological characteristics of liver biopsies in eight patients with hepatic epithelioid hemangioendothelioma.

We aim to investigate the pathological characteristics of liver biopsies and their implications for the prognosis of hepatic epithelioid hemangioendothelioma (HEHE). Clinical data of eight patients (5 male, 3 female) with HEHE were analyzed retrospectively. Expression of CD34, FVIII, AE1/AE3, Hepa-par1, GPC3, CK19 and the proliferation index marker Ki-67 were determined by immunohistochemical staining. The clinical pathological features and effects of treatment on prognosis were investigated. Among the eight patients, four did not exhibit significant symptoms, while four showed symptoms such as abdominal distension, aversion to greasy food and mild fever. Two patients had single liver lesions, while multiple lesions were observed in six cases, in which the tumor cells exhibited spindle, irregular or epithelioid morphology, with scattered, streaked and nested distribution. Individual luminal cells were also visible, containing red cells and accompanied by mucoid or fibrous stroma. All cases were CD34 positive, one case was FVIII factor negative, two cases were AE1/AE3 positive, Ki-67 staining exceeded 15% in two cases, and nuclear fission was visible in two cases. Patients with nuclear fission and Ki-67 > 15% died within 2 years after artery embolization, liver transplantation without relapse was observed in two cases and one case survived with the tumor. The other patients without cellular atypia, without nuclear fission and with Ki-67 < 10% did not relapse during the 2-5 years of follow-up. HEHE can be diagnosed according to hematoxylin and eosin morphology and immunohistochemical characteristics in biopsies before treatment allowing the selection of different treatment protocols based on pathological characteristics.

Resveratrol reduces acute lung injury in a LPS­induced sepsis mouse model via activation of Sirt1.

The development of acute lung injury (ALI) during sepsis almost doubles the mortality rate of patients. The efficacy of current treatment strategies is low as treatment is usually initiated following the onset of symptoms. Inflammation is one of the main mechanisms of autoimmune disorders and is a common feature of sepsis. The suppression of inflammation is therefore an important mechanism for the treatment of sepsis. Sirtuin 1 (Sirt1) has been demonstrated to play a role in the regulation of inflammation. Resveratrol, a potent Sirt1 activator, exhibits anti­inflammatory properties. However, the role of resveratrol for the treatment of ALI during sepsis is not fully understood. In the present study, the anti­inflammatory role of Sirt1 in the lipopolysaccharide (LPS)­induced TC­1 cell line and its therapeutic role in ALI was investigated in a mouse model of sepsis. The upregulation of matrix metalloproteinase-9, interleukin (IL)­1ß, IL­6 and inducible nitric oxide synthase was induced by LPS in the mouse model of sepsis and the TC­1 cell line, and resveratrol suppressed the overexpression of these proinflammatory molecules in a dose­dependent manner. Resveratrol decreased pulmonary edema in the mouse model of sepsis induced by LPS. In addition, resveratrol improved lung function and reduced pathological alterations in the mouse model of sepsis. Knockdown of Sirt1 by RNA interference resulted in an increased susceptibility of TC­1 cells to LPS stimulation and diminished the anti­inflammatory effect of resveratrol. These results demonstrated that resveratrol inhibits LPS­induced ALI and inflammation via Sirt1, and indicated that Sirt1 is an efficient target for the regulation of LPS­induced ALI and inflammation. The present study provides insights into the treatment of ALI during sepsis.

[The effect of 5-fluorouracil on enriching cancer stem cells of hepatoma cell line BEL-7402].

OBJECTIVE: To investigate the effect of 5-FU (5-fluorouracil) on enriching cancer stem cells of HCC cell line BEL-7402 and the biological characteristics of enriched cells. METHODS: The enriching concentration of 5-FU was determined by CCK-8 (cell counting kit-8). Flow Cytometry was used to determine the changes in cell cycle and positive expression ratio of surface marker CD56, CD54, EpCAM and CD133. The self-renewal and differentiation of positive cells were tested by colony formation assay, and were compared with the control group. RESULTS: Enriching concentration of 5-FU was determined as 10 µg/ml with 48 h incubation. After enrichment, G0/G1 phase cells increased from 57.50 %+/-0.98% to 68.70%+/-3.41% (P<0.05). Whereas S phase cells decreased from 40.26%+/-4.12% to 31.80%+/-4.15% (P<0.01); G2/M phase cells disappeared in experimental group, and was 5.80%+/-1.87% in control group (P<0.01). The proportion of the cell cycle changed with significant statistical differences. Meanwhile, positive rate of cell surface makers CD56, CD54, EpCAM and CD133 increased from 0.57%+/-0.12%, 8.10%+/-6.79%, 0.3%+/-0.01% and 3.20%+/-0.99% to 4.13%+/-0.06%, 50.08%+/-1.69%, 0.55%+/-0.07% and 10.51%+/-1.13%, respectively. The difference was significant (P<0.05). The colony forming ratio of CD56, CD54, EpCAM and CD133 negative cells and positive cells were 2.11%+/-0.21%, 3.32%+/-0.31%; 0.86%+/-0.101%, 2.40%+/-0.52 %; 7.19%+/-0.56%, 7.73%+/-0.71%; 2.70%+/-0.26%, 5.75%+/-0.81%, respectively, and significant differences were found between (P<0.05). CONCLUSION: 5-fluorouracil enriched the cancer stem cell population in HCC cell line BEL-7402. CD56 and CD54 can be used as important surface markers in research of liver cancer stem cells.

[Changes and mechanism of apoptosis-related gene expression in T lymphocytic leukemia JM cells induced with matrine].

This study was purposed to investigate the changes of apoptosis-related gene expression in T lymphocytic leukemia JM cells induced with matrine, and its possible mechanism. JM cells was induced with 0.4 mg/ml matrine for 4 days, the total RNA was extracted from JM cells before and after matrine induction, the differential expression of apoptosis-related genes were screened with cDNA Expression Array Kit, the expression change of a part of gene was checked by Western blot. The results indicated that after induction of JM cells with matrine, differential expression of 31 genes were found by gene chip hybridization, the expression of caspase 8 was up-regulated more than 5 times. Western blot analysis showed that the up-regulation of caspase 8 gene expression positively correlated with induction time. It is concluded that differential expressions of many apoptosis-related genes in JM cells can be induced by matrine, in which gene expression of caspase 8 is up-regulated notably.

Glucosamine sulfate-induced apoptosis in chronic myelogenous leukemia K562 cells is associated with translocation of cathepsin D and downregulation of Bcl-xL.

Cathepsin D (cat D) reportedly plays an important role in certain apoptotic processes, the downstream pathways of which involve release of cytochrome c (cyt c) from mitochondria and activation of the caspase cascade. Previous studies revealed that the B-cell lymphoma 2 (Bcl-2) family members Bax or Bid play important roles in apoptotic signal transduction between cat D and mitochondria. Here, we show that glucosamine sulfate (GS) inhibits the proliferation and induces apoptosis of human chronic myelogenous leukemia K562 cells in vitro. GS interfered with the maturation of cat D. Activation of caspase-3, cleavage of poly-(ADP-ribose)-polymerase, release of cyt c, and downregulation of Bcl-xL accompanied GS-induced apoptosis, and these processes were inhibited by the cat D inhibitor pepstatin A. However, we did not detect any altered gene expression of Bcl-2, Bax, or Bid during apoptosis. Translocation of cat D from the lysosome to the cytosol was observed in GS-treated K562 cells. These findings suggest that GS-induced K562 cell apoptosis involves the translocation of cat D from the lysosome to the cytosol. Furthermore, our findings suggest that downregulation of Bcl-xL (but not Bcl-2, Bax, or Bid) connects cat D and the mitochondrial pathway, which causes the release of cyt c and activation of the caspase cascade during GS-induced apoptosis of K562 cells.

[Study on the preparation of resveratrol chitosan nanoparticles with free amino groups on the surface].

OBJECTIVE: To prepare resveratrol chitosan nanoparticles with free amine groups on the surface so as to conjugate ligands, which will actively target to special tissues or organs. METHOD: The chitosan nanoparticles with free amine on the surface was prepared by sodium chloride precipitation. Nanoparticles with different solidification degrees were studied on turbidity, in vitro release, encapsulation efficiency, drug loading and diameter. RESULT: The turbidity of nanoparticles with various solidification degrees decreased at different rates after ultrasonic or water bath heating treatment. All nanoparticles mentioned above obviously shew sustained release. The rate of release was slowed down with the increase of solidification agents. Solidification had no obvious effects on the encapsulation efficiency and drug loading. The diameter of chitosan nanoparticles with 200 microL solidification agents was 487 nm. The polydispersion was 0.144. CONCLUSION: The diameter of the prepared nanoparticles was relatively small. The amine on the surface was free, which offered the possibility of designing the acive target drug delivery system.

[Effects of human fibroblastoid stromal cell line on proliferation of HL-60 cells and expression of VEGF].

To investigate the effects of normal human bone m arrow fibroblastoid stromal cell line (HFCL) on the proliferation of acute myeloid leukemia cell line HL-60 and expression of vascular endothelial growth factor (VEGF), establishing coculture system of leukemia cell line HL-60 and HFCL, growth data was obtained by cell counting. Mitotic index (MI) was observed under Wright-Giemsa staining. Flow cytometry and Western blot were used as assays for cell cycle and expression of proliferating cell nuclear antigen (PCNA) separately. VE GF levels were evaluated by using commercial ELISA kits. The results showed that compared with HL-60 cells without HFCL cells, the proliferation of HL-60 cells in direct contact with HFCL cells and with HFCL cells separated by transwell was inhibited. The MI of HL-60 cells without HFCL cells was highest followed by HL-60 cells separated by transwell and HL-60 cells in direct contact with HFCL cells. The expression of PCNA in HL-60 cells with HFCL cells were lower than HL-60 cells without HFCL cells. Meanwhile, the percentage of HL-60 cells in G1 phase cocultured with HFCL cells was higher than that without HFCL cells while the percentage of Sphase cells was lower. The levels of VEGF in HL-60 cells with HFCL cells were lower than that in HL-60 cells alone. In conclusion, the normal bone marrow fibroblastoid stromal cells inhibited the proliferation of HL-60 cells as well as the expression of VEGF.

[Inclusion bodies of human cytomegalovirus are composed of the DNA and immediately early and early antigens of the virus].

BACKGROUND: To study the composition and significance of the inclusion bodies of human cytomegalovirus (HCMV). METHODS: Microdissection of inclusion bodies, PCR and Southern blot were adopted to detect DNA, and immunohistochemistry method and catalyzed signal amplification (CSA) were used to detect the different antigens of HCMV. RESULTS: The inclusion bodies of HCMV were separated from the tissue section of human salivary gland. The fragments amplified by PCR from these dissected inclusion bodies were confirmed to be the DNA of HCMV. With the immunohistochemical method CSA, the immediately early and early antigens of HCMV were detected with monoclonal antibodies DDG9/CCH2, while matrix protein AAC10 was negative in the inclusion bodies. CONCLUSION: The ingredient of inclusion bodies of HCMV included the DNA and the antigens expressed in specific stage of the virus.

[Matrine effects on JM cells by inhibiting proliferation and inducing apoptosis].

OBJECTIVE: To study effects of matrine on JM cell strain. METHOD: Morphologic changes were observed under light microscope with Wright-Giemsa staining, fluorescence microscope with Hoechst 33,258 staining and electron microscope. Alteration of cell cycle of different dose treating groups at the fourth day and 0.8 mg.mL-1 treatment group at the first, second, third, fourth day was analyzed by Flow cytometry. DNA ladder was detected with gel electrophoresis. RESULT: From the third day after treatment of matrine, typical apoptosis features of cells were observed under light microscope and electron microscope in all test groups, and the features were more prominent with the time prolonging. At fourth day, flow cytometry analysis showed that there were sub-G1 peaks in all groups. From 0.1, 0.2, 0.4, 0.6 to 0.8 g.L-1 treatment groups, the rate of apoptotic cells to total cells were 3.1%, 2. 5%, 13.3%, 40.4%, 48.6%, respectively, and what in the control group was 1.4%; the rate of S phase cells to total cells was 28.9%, 26.1%, 27.7%, 0.9%, 14.2%, what in the control group was 30.4%; the rate of G1 phase cells to total cells was 63. 2%, 67.5%, 68.1%, 75.2%, 83.6%, what in the control group was 41.8%; From the first, second, third to fourth day, the rate of apoptotic cells to total cells of 0.8 mg.mL-1 treatment group were 3.0%, 3.7%, 9.1%, 48.6%, respectively; the rate of S phase cells to total cells was 28.6%, 17.5%, 19.1%, 14.2%; the rate of G1 phase cells to total cells were 45.5%, 77.3%, 77.2%, 83.6%. Gel electrophoresis displayed "DNA ladder" in 0.4, 0.6, 0.8 g.L-1 groups, while 0.1 and 0.2 g.L-1 groups didn't show such result. CONCLUSION: Matrine can repress DNA synthesis and arrest JM cell strain at G1 phase, sequentially inhibiting the proliferation of the cell. Besides, this alkaloid can induce the apoptosis of JM cells.