Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Carcinoma, Hepatocellular ; pathology ; Cell Division ; drug effects ; Cell Line, Tumor ; Humans ; Liver Neoplasms ; pathology ; Organoplatinum Compounds ; pharmacology

OBJECTIVETo investigate whether there is intercellular transfer of functional P-glycoprotein(P-gp) from P-gp-positive cells to P-gp-negative cells in vitro.

METHODSHepG2/GFP cells, a HepG2 cell line stably expressing GFP, were co-cultured with HepG2/ADM cells, an adriamycin-resistant cell line derived from HepG2 cells. The distribution of P-gp in hepatocellular carcinoma cell was observed under laser scanning confocal microscope (LSCM). Immunomagnetic beads were used to separate HepG2/GFP cells from the mixed culture. The abundance of P-gp was analyzed by western blot, and the expression of mdr1 mRNA was detected by qRT-PCR.

RESULTSYellow fluorescence was detected in HepG2/aqMDR cells, green fluorescence was detected in HepG2/GFP cells, red fluorescence was detected in HepG2/ADM cells by LSCM. The level of P-gp protein in HepG2/aqMDR cells was lower than that in HepG2/ADM cells, but higher than that in HepG2/GFP cells (q = 35.07, P < 0.05) and HepG2 cells (q = 36.87, P < 0.05). The expression of mdr1 mRNA in HepG2/ADM cells was higher than that in HepG2/aqMDR, HepG2 and HepG2/GFP cells, but there was no significant difference in mdr1 mRNA among HepG2/aqMDR, HepG2 and HepG2/GFP cells (F = 2.30, P > 0.05).

CONCLUSIONSP-gp can transfer from drug resistant hepatocellular cells to sensitive hepatocellular carcinoma cells. This study suggests a novel mechanism of multidrug resistance in hepatocellular carcinoma.

ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Carcinoma, Hepatocellular ; genetics ; metabolism ; pathology ; Coculture Techniques ; methods ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; genetics ; Genes, MDR ; Green Fluorescent Proteins ; Hep G2 Cells ; Humans ; Liver Neoplasms ; genetics ; metabolism ; pathology ; Plasmids ; Protein Transport ; RNA, Messenger ; genetics ; metabolism ; Transfection

OBJECTIVETo investigate the relationship between activation of nuclear factor-K-gene binding (NF-κB) and apoptosis induced by matrine(MT) in transplanted tumor of human hepatocellular carcinoma in nude mouse.

METHODSTumors were established by injection of hepatocellular carcinoma cell line HepG2 into the back of nude mice. The mice were divided randomly into four groups: Control group, MT group (35 mg/kg), PDTC group (120 mg/kg) and Combination group: PDTC + MT group (120 mg/kg + 35 mg/kg), the reagents were injected peritoneally. The tumor growth curve of nude mice bearing transplanted tumor were observed and the inhibition ratios were evaluated. Apoptosis of carcinoma cells was analyzed by TUNEL. The DNA-binding activity of NF-κB was determined by electrophoretic mobility shift assay (EMSA). Expression of bcl-2 and bax in carcinoma tissue were detected by immunohistochemical method. NF-κB mRNA, bcl-2 mRNA and bax mRNA in carcinoma tissue were detected by RT-PCR.

RESULTSPyrrolidine dithiocarbamate (PDTC) could enhance the inhibition of matrine on carcinoma proliferation (P < 0.05). The apoptosis and activation of NF-κB in carcinoma cells could be induced by matrine. PDTC significantly suppressed NF-κB activation induced by matrine in carcinoma cells from 93.64 ± 2.95 to 65.78 ± 5.65 (F = 124.754, P < 0.01). Meanwhile, PDTC increased the apoptosis induced by matrine from 55.9% ± 2.8% to 74.3% ± 4.8% (P < 0.05).A positive correlation observed between the expressions of NF-κB and of bcl-2 (Pearson correlation coefficient = 0.983, P < 0.01).

CONCLUSIONSMatrine could induce apoptosis and activation of NF-κB in transplanted tumor. PDTC could increase apoptosis in hepatocellular carcinoma cells might be due to the suppression of NF-κB activation and the enhancement of bcl-2 expression.

Alkaloids ; pharmacology ; Animals ; Apoptosis ; drug effects ; Carcinoma, Hepatocellular ; metabolism ; pathology ; Hep G2 Cells ; Humans ; Liver Neoplasms ; metabolism ; pathology ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; NF-kappa B ; metabolism ; Neoplasm Transplantation ; Pyrrolidines ; pharmacology ; Quinolizines ; pharmacology ; Thiocarbamates ; pharmacology

Objective Mutated inbred animal model is introduced to the practical course of genetic diagnosis in the hope that medical students are able to apply what they have learned to clinical cases, based on a deep understanding of principle and technology on gene mutation detection. Methods We integrated DNA extraction, polymerase chain reaction, agarose gel electrophoresis, and gel imaging analysis into a comprehensive experiment and arranged 4-year-programme undergraduates majoring in preclinical medical sciences to conduct it with the purpose of investigating the internal relations between phenotype and genotype in a hairless Uncv mouse model. Subsequently, the questionnaire aimed at evaluating learning effect on the part of students was handed out and their feedbacks were analyzed. Results More than 90% of respondents are satisfied with the general learning effect. Especially, 98. 7% of students support the enhancing effect of the new teaching mode on their research skills and 96% consider the practical course helpful to their problem-solving ability. Conclusions The introduction of mutated inbred animal model to the practical system of molecular diagnostics proves beneficial to boost students' learning effect and scientific research quality. Our practice also provokes thoughts on the further utilization of animal models in teaching system of medical sciences.

Objective To explore the safety of collection of peripheral blood stem cells(PBSCs) from low-weight infants with osteopetrosis(OP) and its clinical significance. Methods One case of low-weight infants with OP received PBSCs collection using a continuous-flow blood cell separator,and the safety of collection process was observed.The amount of monocyte cell(MNC) and CD34+ cell were noted and its clinical significance was analyzed.Results Low-weight infants with OP could tolerate collection process,the number of collection MNC and CD34+ cells were 10.06?108/kg,2.74?106/kg.Conclusion Adequate PBSCs can be collected from OP who need not be mobilized,thus can offer backup for graft failure.PBSCs collection from low-weight infants is safe.

Abdomen ; Hernia ; Humans ; Prostheses and Implants ; Prosthesis Implantation ; Treatment Outcome

OBJECTIVETo investigate clinicopathologic features of solitary plasmacytoma of bone (SPB) and the role of immuno-phenotype and immunoglobulin gene rearrangement detection in the diagnosis and differential diagnosis of SPB.

METHODSA total of 21 cases of SPB were selected during a period from 1990 to 2008. A retrospective clinicopathologic study and immunohistochemistry (EnVision or EliVision methods) of 17 antigens were performed. In addition, universal IgH (FR3A/LJH/VLJH) primers and BIOMED-2 PCR multiplex tubes were used for IgK and IgL rearrangement analysis.

RESULTSThe age of patients ranged from 36 to 72 years with a media of 50 years. Axial skeleton was the most common site of involvement, accounting for 66.7% of the cases (14 of 21), followed by the extremities of 33.3% (7 cases). Low serum level of M-components was found in 5 cases, including two of IgG type (21.4 g/L) and three of IgA type. Clinical manifestations were closely related to the anatomic sites involved, such as pain due to bone destruction, symptoms and signs caused by compression of spinal cord or nerve root, and pathological fracture. All cases presented as a solitary osteolytic lesion. According to the histological grading criteria, grade I tumor was seen in 12 of 21 cases (57.1%). The remaining were grade II (5 cases, 23.8%) and grade III (4 cases, 19.0%). Immunohistochemically, the neoplastic cells expressed two or more plasma cell antigens, including CD138, CD38 and PC, but no CD19 and CD20. CD79a expression detected in 23.8%(5/21) of the cases. Expression of CD56, CD27 and CD44v6 were 57.1% (12/21), 15.0% (3/20) and 23.8% (5/21), respectively. Follow-up data were available in 12 of the 21 patients (57.1%). Five patients were alive and 7 died. Three patients developed multiple myeloma (MM) and died of the tumor.

CONCLUSIONSSPB is a rare tumor with bone pain as the most common presenting symptom due to bone destruction. The diagnosis of EMP can only be established after exclusion of an extramedullay invasion by MM. Immunophenotype and IgH gene rearrangement analysis play important roles in the diagnosis of SPB.

ADP-ribosyl Cyclase 1 ; metabolism ; Adult ; Aged ; Bone Neoplasms ; genetics ; metabolism ; pathology ; surgery ; Diagnosis, Differential ; Female ; Follow-Up Studies ; Gene Rearrangement, B-Lymphocyte, Heavy Chain ; Humans ; Immunophenotyping ; Lymphoma, Large B-Cell, Diffuse ; metabolism ; pathology ; Lymphoma, Large-Cell, Anaplastic ; metabolism ; pathology ; Male ; Melanoma ; metabolism ; pathology ; Middle Aged ; Multiple Myeloma ; pathology ; Plasmacytoma ; genetics ; metabolism ; pathology ; surgery ; Retrospective Studies ; Survival Rate ; Syndecan-1 ; metabolism

OBJECTIVESTo study the clinicopathologic features of Rosai-Dorfman disease (RDD), expression of various antigens, human herpes virus type 8 (HHV8), human papillomavirus (HPV)-DNA and Epstein-Barr virus (EBV)-mRNA, and compare the findings with those in the literature.

METHODSThe clinicopathologic findings of 16 Rosai-Dorfman disease cases were retrospectively reviewed. Immunohistochemical study for S-100 protein, CD68 (PG-M1), CD163, CD21, CD1a, CD20, CD45RO, CD4, CD8, M-CSF and HHV8 was carried out in 9 of the 16 cases. In-situ hybridization for EBV-mRNA and HPV-DNA was also performed.

RESULTSThe male-to-female ratio of the patients was 4.33:1. Amongst the 16 cases studied, 62.5% (10/16) presented nodal RDD, with cervical lymph node predominantly involved. Half of these cases had affected lymph nodes in more than one anatomic site. Extranodal RDD represented 37.5% (6/16) of the cases. The relapse rate of extranodal RDD was higher than that of nodal RDD. Histologically, nodal RDD was characterized by dilated sinuses filled with large polygonal histiocytes which contained lymphocytes and plasma cells. For extranodal lesions, various degrees of stromal fibrosis were seen in association with mixed inflammatory cells (especially plasma cells). The large polygonal histiocytes varied in number and were distributed in clusters or patches. Immunohistochemical study showed that the abnormal histiocytes were strongly positive for S-100 protein. They also expressed CD68, CD163 and M-CSF, but were negative for CD1a, CD21 and HHV8. The lymphocytes in cytoplasm of these histiocytes were positive for both T and B cell markers (with T cell predominance, including a mixture of CD4- and CD8-positive cells). HPV-DNA and EBV-mRNA were not detected by in-situ hybridization. To date, 62 cases of RDD have been reported in mainland China, including 34 cases of nodal RDD and 18 cases of extranodal RDD. The remaining 10 cases involved both lymph nodes and extranodal sites. Compared with overseas reports, RDD occurring in China tended to affect older patients and with slight male predilection.

CONCLUSIONSRosai-Dorfman disease is relatively rare in China. Pathologic diagnosis of extranodal RDD may be difficult. The demographic data of RDD in China, including age and sex of patients, are different from those in the literature.

Adolescent ; Adult ; Aged ; Antigens, CD ; metabolism ; Antigens, Differentiation, Myelomonocytic ; metabolism ; Bone Diseases ; metabolism ; pathology ; virology ; Child ; DNA, Viral ; analysis ; Female ; Follow-Up Studies ; Herpesvirus 8, Human ; genetics ; isolation & purification ; Histiocytosis, Sinus ; metabolism ; pathology ; virology ; Humans ; Immunohistochemistry ; Lymph Nodes ; pathology ; Macrophage Colony-Stimulating Factor ; metabolism ; Male ; Middle Aged ; Nose Diseases ; metabolism ; pathology ; virology ; RNA, Viral ; analysis ; Receptors, Cell Surface ; metabolism ; Retrospective Studies ; S100 Proteins ; metabolism ; Skin Diseases ; metabolism ; pathology ; virology ; Young Adult

CD151 is a member of the tetraspanin family that is implicated as a promoter of pathological or physiological angiogenesis. C-Met is expressed on a variety of cells including vascular endothelial cells (VECs) and up-regulated during angiogenesis. In this study, we investigated whether CD151 regulated migration, proliferation, tube formation and angiogenesis of human umbilical VECs (HUVECs) with activation of C-Met. Moreover, we studied whether CD151 could affect the angiogenic molecules such as nitric oxide (NO), vascular cell adhesion molecule-1 (VCAM-1) and vascular endothelial growth factor (VEGF). The expression of CD151 was determined by Western blotting. The cell proliferation assay was performed using the cell counting kit-8 (CCK-8) method and cell migration was assessed in microchemotaxis chambers by using fetal bovine serum (FBS) as the chemotactic stimulus. The angiogenic molecules were evaluated using ELISA. The NO level was detected using NO detection kit. The potential involvement of various signaling pathways was explored using relevant antibodies. We found that proliferation, migration and tube formation of HUVECs were promoted by CD151 with activation of C-Met, FAK and CDC42, while they were suppressed with CD151 knockdown by RNAi. Similarly, the levels of NO, VCAM-1 and VEGF in HUVECs were increased by CD151, but they were inhibited with CD151 knockdown by RNAi. These data suggested that CD151 could promote migration, proliferation, tube formation and angiogenesis of HUVECs, which was possibly related to the C-Met signaling pathways.
Base Sequence ; Human Umbilical Vein Endothelial Cells ; Humans ; Neovascularization, Physiologic ; RNA, Small Interfering ; genetics ; Receptor Protein-Tyrosine Kinases ; metabolism ; Signal Transduction ; Tetraspanin 24 ; genetics ; metabolism

OBJECTIVESTo study the correlation between positive rates of RNA in clinical confirmed severe acute respiratory syndrome (SARS) patients and its appearance in relation to the development of the disease in order to provide scientific basis for early diagnosis, effective prevention and treatment of the disease.

METHODSOne-step reverse transcription-polymerase chain reaction (RT-PCR) was used to amplify the SARS RNA in the clinical specimens from different courses of the disease. The representative amplicons were then sequenced. Chi-square for trend test was performed to study the correlation between positive rates of RT-PCR and at different periods after the onset of the disease.

RESULTSThe fragments amplified from the sputum specimens of SARS patients were shown to share 100% homology with the published SARS-associated coronavirus. Of the different clinical specimens, positive rate in the stools appeared to be the highest (21.55%). Chi-square for trend test revealed that the positive rates of stools and sputa of SARS patients decreased with the development of the disease (chi(2) for trend = 12.55 and 16.408, P = 0.0004 and P = 0.000 05 respectively).

CONCLUSIONOne-step RT-PCR proved to be an effective method for the detection of SARS-associated coronavirus from clinical specimens. Data as indicated that the positive rates of SARS coronavirus were decreasing in SARS patients along with the disease progression.

Adolescent ; Adult ; Aged ; Chi-Square Distribution ; China ; Disease Progression ; Feces ; virology ; Female ; Humans ; Male ; Middle Aged ; Mucus ; virology ; RNA, Viral ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; SARS Virus ; genetics ; isolation & purification ; Severe Acute Respiratory Syndrome ; pathology ; virology