Intrasplenic co-transplantation of fetal hepatic progenitor cells and transforming growth factor beta 1 induced hepatic stellate cells ameliorates acute liver injury / 中国组织工程研究

BACKGROUND: Difficulty in long-time survival and continuous proliferation is the main problem for transplanted fetal hepatic progenitor cells and co-transplantation with transforming growth factor beta 1 (TGF-β1)-induced hepatic stellate cells may be a promising way to solve this scientific obstacle. OBJECTIVE: To explore the therapeutic effects of co-transplantation of fetal hepatic progenitor cells and TGF-β1-induced hepatic stellate cells on acute liver injury in mice. METHODS: Over-expression vector pHBLV-CMVIE-TGF-β1 was infected to mouse hepatic stellate cells and transfection efficiency was detected by immunocytochemistory, western blot and qRT-PCR. Hepatic progenitor cells, mHPCs-E14.5, were cultured and identified by immunofluorescence in vitro.The mouse model of acute liver injury was established by intraperitoneal injection of CCl4in combination with 2/3 partial hepatectomy, followed by mHPCs-E14.5transplantation, co-transplantation of mHPCs-E14.5and mHSCs-pHBLV-CMVIE-TGF-β1 (experimental co-transplantation group) or co-transplantation of mHPCs-E14.5and mHSCs-pHBLV-CMVIE-GFP (control co-transplantation group) for cell transplantation assay. Confocal immunofluorescence staining against CK19, ALB, a-SMA was performed to analyze the engraftment and differentiation of transplanted cells in the splenic parenchyma 14 days post-transplantation; serum alanine transferase and aspartate transferase levels were monitored using an automatic biochemistry analyzer. RESULTS AND CONCLUSION: (1) A hepatic stellate cell line that over-expressing TGF-β1 was successfully established and expression levels of TGF-β1 and α-smooth muscle actin were efficiently up-regulated in the over-expression group (P < 0.01). (2) mHPCs-E14.5expressed massive AFP and low levels of ALB and CK19,confirming that this cell line was in complete conformity with fetal hepatic progenitor cells in vitro.(3)CK19 and ALB positive cells existed in the splenic parenchyma in mHPCs-E14.5transplantation group.Highly expressed ALB but less expressed α-SMA and CK19 were observed in the control co-transplantation group, while massive CK19 and a-SMA positive cells as well as less level of ALB positive cells existed in the experimental co-transplantation group. Serum alanine transferase and aspartate transferase levels were decreased remarkably after cell transplantation, and moreover, the decrease was more obvious in the experimental co-transplantation group (P < 0.05). Overall, transplanted fetal hepatic progenitor cells engraft and differentiate into hepatocytes and cholangiocytes in the splenic parechyma successfully in vivo.Importantly,hepatic stellate cells induced by TGF-β1 promote the differentiation of fetal hepatic progenitor cells into cholangiocytes and accelerate recovery from CCl4/partial hepatectomy induced acute liver injury.

Changes of Acin1 expression in congenital cataract mouse during retinal development / 国际眼科杂志(Guoji Yanke Zazhi)

?AlM: To observe the expression of Acin1 ( apoptotic chromatin condensation inducer 1 ) in congenital cataract mouse retina during development and investigate the differences of retinal apoptosis and the connection of lens and retina development between congenital cataract mouse and normal mouse. ?METHODS: There were congenital cataract mice ( 10 female and 5 male) and normal C57BL/6 mice (10 female and 5 male) . One male and two female mice were fed in the same cage randomly. The young mice were divided into two groups: congenital cataract group and normal control group. Five young mice were treated each group on 1, 5, 9, 14, 17, 21, 26, 60d. The left eyes were fixed with 4% neutral formalin to detect AClN1 protein by immunohistochemistry and retinas from right eyes were used to detect the mRNA expression of Acin1. ?RESULTS: Acin1 had sustained expression in each group. AClN1 protein gradually expressed from the ganglion cell layer, inner nuclear layer to the outer nuclear layer following retinal development. lt mainly expressed on ganglion cell layer and inner nuclear layer, but not neuroblastoma layer. AClN1 protein positive cells on P1 ~ P14d increased in normal control group, P17d reduced, after P21d positive cells of each layers decreased. The overall trend was similar in congenital cataract group with normal control group, P1 ~ P14d positive cells count was lower than normal control group, P17-P21d positive cells were flat and higher than the normal control group. Compared with the same day of the two groups, the differences except for P17, P26, P60d were significant (P ?CONCLUSlON: Acin1 exist differential expression of time and space in mouse retina during development, congenital cataract crystal developmental disorder may affect the expression of Acin1 and retinal cell apoptosis and development.

The study on platelet-derived growth factor and proliferating cell nuclear antigen antisense oligodeoxynucleotides together inhibiting the stenosis of transplanted vascular / 中华外科杂志

<p><b>OBJECTIVE</b>To study the effect of platelet-derived growth factor (PDGF) and proliferating cell nuclear antigen (PCNA) antisense oligodeoxynucleotides (AODN) together on inhibiting the proliferation of the stenosis of transplanted vascular.</p><p><b>METHODS</b>The left and right external iliac arteries (length 1.0 cm) of rabbits were transplanted reciprocally. The transplanted vascular were respectively soaked in liposomes, PDGF-AODN, PCNA-AODN and PDGF-AODN adding PCNA-AODN solution about 20 minute, the vascular anastomotic were sutured by 8/0 suture of soaked in AODN solution. Four weeks later, the specimens were harvested for microscopy. The pathological morphology of transplanted vascular were observed under microscope (HE). The intimal thickness and area, stenosis ratio(%) of transplanted vascular were calculate and analysed statistically among group by computer system. The number of positive cells of PDGF's mRNA in transplanted vascular wall were counted with in situ hybridization histo-cytochemistry and the number of positive cells of PCNA's protein in transplanted vascular wall were counted by S-P immunochemistry.</p><p><b>RESULTS</b>The intimal thickness and area, stenosis ratio of transplanted vascular, the number of PDGF and PCNA positive cell in PDGF-AODN adding PCNA-AODN group were significantly lower than those in other group (P < 0.01), and that were lower evidently than PDGF-AODN group and PCNA-AODN group.</p><p><b>CONCLUSION</b>PDGF and PCNA antisense oligodeoxynucleotides together could significantly inhibit the proliferation of vascular smooth muscle cell and stenosis of transplanted vascular.</p>

To study the effects of local co-transfection vascular endothelial growth factor165 and tissue-type plasminogen activator genes on inhibiting intimal hyperplasia after operation injury artery in rabbits / 中华外科杂志

<p><b>OBJECTIVE</b>To observe the effects of local co-transfection vascular endothelial growth factor165 (VEGF165) and tissue-type plasminogen activator genes on inhibiting intimal hyperplasia and restenosis in rabbits artery after operation injury and possible mechanisms.</p><p><b>METHODS</b>Micrology operation injury was used to establish the model of intimal injury of right external iliac artery in rabbits. To select 120 male New Zealand rabbits and were randomly divided into 3 groups (n = 40, in each group): Group A (physiological brine control group), Group B (pBudCE4.1 group), Group C (pBudCE4.1/VEGF165-tPA group). The vas-wall of micrology operation injury were infused respectively physiological brine, pBudCE4.1 and pBudCE4.1/VEGF165-tPA transfection solution by micro-injector. Each group were divided into 5 subgroups (n = 8, in each subgroup) randomly according to the sacrifice times (2 d, 1 week, 2 week, 4 week and 8 week after operation). The injured vascular specimen were harvested for pathology test, electric microscopy study, reverse transcription-PCR examining and immunochemistry detecting.</p><p><b>RESULTS</b>The intimal area and narrow ratio of vases in Group C at every time point after operation were significantly lessened than that in Group A and Group B (P < 0.01). The narrow ratio of vases in Group C at 8 week after operation were decreased respectively by 57.9% and 59.0% than that in Group A and B. The expression of VEGF165 mRNA in Group C were increased significantly than that in Group A and B at every time point after operation (P < 0.01), the expression reached the peak at 1 week and continued to 4 week after operation. Immunohistochemical identified that tPA positive cell in Group C were significantly increased than that in Group A and B (P < 0.01) at every time point after operation.</p><p><b>CONCLUSION</b>Local co-transfection VEGF165 and tPA genes could restrain intimal hyperplasia and restenosis of vas, which lay a foundation for future multi-gene therapy of vascular intimal hyperplasia.</p>

Effects of platelet derived growth factor antisense oligodeoxynucleotides and tissue-type plasminogen activator gene transfection on inhibition of intimal proliferation / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To observe the effects of co-transfection of platelet derived growth factor antisense oligodeoxynucleotides (PDGF-AODN) and tissue-type plasminogen activator (tPA) gene on inhibition of intimal proliferation of auto-transplantion artery.</p><p><b>METHODS</b>One hundred male New Zealand rabbits were randomly divided into four groups (25 in each): Group A (control group), Group B (PDGF-AODN transfection group), Group C (tPA gene transfection group) and Group D (PDGF-AODN and tPA co-transfection group). The left and right external iliac arteries were transplanted reciprocally. The transplanted arteries were respectively soaked in PDGF-AODN, pBudCE4.1/tPA and PDGF-AODN plus pBudCE4.1/tPA solution about 15 minute before transplantation. The rabbits were sacrificed at 3d, 1w, 2w, 4w and 8w after operation. The specimens were harvested for pathologic examination, electron microscopy, chromogenic substrate test, 3H-TdR incorporation test and immunohistochemical staining.</p><p><b>RESULT</b>The scan electron microscopy showed that there were a few thrombocytes on vas-wall of Group C and D without thrombus, whereas there were abundant thrombocytes and thrombus forming on vas-wall of Group A and B. The intimal area, stenosis ratio of transplanted artery, 3H-TdR incorporation,the number of PDGF positive cell in Group D were significantly less than those in Group A (P<0.01),Group B and Group C (both P<0.05). The activity of tPA gene products in transplanted vas-wall of Group D was significantly higher than that of Group A (P<0.01).</p><p><b>CONCLUSION</b>Local co-transfection of PDGF-AODN and tPA gene can effectively inhibit the proliferation of vascular smooth muscle cells, hyperplasia of intima and restenosis of transplanted artery.</p>