Objective To investigate the expressions of platelet derived growth factor (PDGF) and survivin in hepatocellular carcinoma (HCC) with portal vein tumor thrombosis (PVTT) and to evaluate the relationships among the expressions of PDGF, survivin and the proliferation of cancer cells. Methods The expression of PDGF mRNA in 16 cases in HCC with PVTT group was observed by in situ hybridization and the results were compared with that in HCC tissue group. The expressions of PDGF and survivin protein in 36 cases in HCC with PVTT group were detected with immunohistochemistry and it was found that there was a correlation between them. Flow cytometry (FCM) was used to measure the proliferation of cancer cells and it was also used to analyze the relations among PDGF, survivin and the proliferation of cancer cells. Results The expression level of PDGF mRNA in HCC with PVTT group was significantly higher than that in HCC tissue group (P＜0.01). There was a positive correlation between the expressions of PDGF and survivin protein in HCC with PVTT (P＜0.01). The degree of proliferation of cancer cells in PDGF and survivin protein positive group was significantly higher than that in negative group (P＜0.01). Conclusion PDGF and survivin gene over-expressed in HCC with PVTT group and there is a positive correlation between the expressions of PDGF and survivin protein. The proliferation of cancer cells increases as the expressions of PDGF and survivin increase.
Objective To investigate the mechanismof lung injury caused by paraquat poisoning by observing the changes of fibrogenic cytokines in acute paraquat poisoned rats and the effects of pyrrolidine dithiocarbamate ( PDTC) . Methods Sprague-Dawley rats were randomly divided into three groups, ie. acontrol group ( n =6) , a PDTC group ( n =36) , a paraquat group ( n = 36) , and a paraquat + PDTC group( n =36) . The rats in the PDTC group, the paraquat group, and the paraquat + PDTC group were subdivided into 6 subgroups sacrificed respectively on 1st, 3rd,7th,14th, 28th and 56th day after the treatment. The levels of transforming growth factor-β1( TGF-β1 ) , platelet-derived growth factor ( PDGF) , insulin-like growthfactor-1 ( IGF-1) in serum were measured. Meanwhile the expression of connective tissue growth factor ( CTGF) and hydroxyproline in lung tissues were detected. The relationship of above cytokines with hydroxyproline was analyzed. Results The destructive phase in early ( 1 ～7 d) was characterized by hemorrhage, alveolar edema, and inflammatory cell infiltration. The proliferous phase in later stage ( 14 ～56 d) was characterized by diffused alveolar collapse with fibroblast proliferation and patchy distribution of collagen fibers. Compared with the control group, the level of TGF-β1 on all time points, the level of PDGF from7th to 56th day, the level of IGF-1 from3rd to 56th day in the paraquat group all significantly increased ( P lt;0. 01) . Immunohistochemistry results showed CTGF positive cells mainly located in aleolar epithelialcells, endothelial cells,macrophages in early stage, and fibroblasts were main positive cells on the 28th and the 56th day. The expression of CTGF in the paraquat group increased gradually compared with the control group on different time points ( P lt; 0. 05 or P lt; 0. 01) . Meanwhile, the levels of above cytokines were positively correlated with the level of hydroxyproline. Noteworthy, PDTC treatment led to significant decreases of above cytokines compared with the paraquat group in corresponding time points ( P lt;0. 05 or P lt;0. 01) .Conclusions Over expressions of IGF-1, TGF-β1 , PDGF, IGF-1 and CTGF may play important roles in lung fibrosis of paraquat poisoned rats. PDTC, as a b NF-κB inhibitor, may inhibits NF-κB activity and further significantly decreases expressions of cytokines, leading to significantly attenuated pulmonary inflammation and fibrosis. However, the mechanisms of PDTC intervention still remain to be explored.
Objective To study the function and mechanism of G protein coupled receptor kinase interacting protein 1(GIT1) RNA hairpin (GIT1-RNAh) in osteoblast migration. Methods The sixth passage osteoblasts were divided into 2 groups and were infected by GIT1-RNAh (experimental group) and green fluoresence protein RNA hairpin (GFP-RNAh) (control group) adenovirus for 12 hours respectively. Each group was further classfied into two groups according to with or without platelet-drived growth factor (PDGF) stimulation. The GIT1 expression and Paxillindistribution was analyzed by immunofluorescence staining. Paxillin phosphorylation was detected by Western Blot. The localization of Paxillin was determined by co-immunofluorescence staining after transfection with cyanine fluorescence protein tagged GIT1RNAh (CFP-GIT1-RNAh)(experimental group) and GFP-RNAh (CFP-GFP-RNAh)(control group). The role of GIT1-RNAh (experimental group) and GFP-RNAh (control group) adenovirus in osteoblasts migration was determined by wound healing assay. Results Immunofluorescence staining results showed that the GIT1-RNAh significantly inhibited endogenous GIT1 expression, interfered Paxillin distribution.Western Blot results showed that Paxillin phosporylation was obviously inhibited in osteoblasts infected with GIT1-RNAh adenovirus (P＜0.05). The wound healing assay results howed that GIT1-RNAh adenovirus significantly inhibited osteoblast migration induced by PDGF. Conclusion GIT1-RNAh inhibits osteoblasts migration by interfering paxillin distribution and decrease Paxillin phosphorylation.
Objective To study the function of platelet-derived growth factor (PDGF) in inducing phosphorylation extracellular signalregulated kinase 1/2 (pERK1/2) localization in osteoblasts. Methods Primary osteoblasts were isolated and cultured from cranial bone of 10 mice atthe age of 3 days, weighting 6-9 g without limitation in male and female.The sixth passage osteoblasts were incubated in 1% serum for 12 hours and divided into 2 groups: treated with DMSO(control group) or with PP2(experimentalgroup) for 30 minutes. Each group was further divided into 2 subgroups according to with or without PDGF (20ng/ml) stimulation for 10 minutes. pERK1/2 localization was analysized by immunofluorescence staining in osteoblasts pretreated with or without Src inhibitor PP2. The sixth passage osteoblasts were divided into 2 groups treated with DMSO (control group) or with PP2 (experimental group) for 30 minutes. Each group was further divided into two subgroupsaccording to with or without PDGF (20 ng/ml) stimulation for 10 mintues. The ability of osteoblast migration was determined by wound healing assay. The sixth passage osteoblasts were divided into 2 groups treated with DMSO (control group) or 10 μmol/L PP2 (experimental group) for 30 mintues. Each group was further divided into 2 subgroups according to with or without PDGF (20 ng/ml) stimulation.The pERK1/2 was determined by Western blot in osteoblastic cytoskeleton inducedby PDGF. Results Immunofluorescence staining showed pERK1/2 localization in osteoblastic nuclears and focal adhesions after PDGF stimulation. PP2 significantly inhibited ERK1/2 localization in focal adhesions, but not in nuclears. The wound healing assay results showed that PP2 significantly inhibited osteoblast migration induced by PDGF. The result of Western blot demonstrated that pERK1/2 in osteoblastic cytoskeleton was significantly inhibitedSrc activation is required for pERK1/2 translocalization to focal adhesions and osteoblasts migration.
Objective To investigate the possible signaling mechanisms by which recombinant human plateletderived growth factor (rhPDGF) accelerated healingof cutaneous wound in diabetic rats. Methods Four full-thickness skin woundswere incised in the back of 26 male Wistar diabetic rats. The wounded rats were divided into 3 groups (7 or 8 rats each group). One group without treatmentwas used as a control, and the other 2 groups were treated with rhPDGF at a dose of 7.0 μg/cm2 wound or vehicle (DMSO/09% NaCl, vol/vol 1∶1) from 1 to14 days. The wound healing was evaluated by the measurements of the wound volume and area. Immunofluorescent and immunohistochemical staining were used to examine the phosphorylation of extracellular signalregulated kinase 1/2 (ERK1/2) andthe expression of proliferative cell nuclear antigen (PCNA), respectively. Results Granulation tissue appeared in the bed of wound after injury. The number of blood capillary buds and fibroblasts was greater in the rhPDGF-treated group than that in the other 2 groups. A lot of inflammatory cells infiltration and collagen deposition were observed in the wound. The wound-volume in the rhPDGF-treated group was smaller than that in control group (Plt;0.05). The reepithelialization rate in rhPDGF-treated group was higher than that inthe other 2 groups at 7 days after injury (Plt;0.05). The expression of PCNA in reparative cells was higher in rhPDGF-treated group than in control group or vehicle-treated group at 3,7 days after injury(Plt;0.05). The phosphorylation of ERK1/2 was ber in rhPDGF-treated group than that in control group or vehicle group at 7 and 14 days after injury(Plt;0.05). Conclusion These results suggest that rhPDGF accelerates wound healing and improves healing quality by increasingthe phosphorylation of ERK1/2.
Objective To examine an effect of the locally-used platelet derived growth factor-BB (PDGF-BB) on the healing of the medial collateral ligament (MCL) in the knee joints of rats. Methods Forty-eight rats were equally randomly divided into 2 groups: the experimental group (group A) and the control group(group B). MCL of all the rats were ruptured to establish the wound models. In group A, 5 μg of PDGF-BB was locally injected in the wound of each rat and then the wound was sutured; but in group B, the wound was only sutured. After 2 weeks, histological evaluations were performed to determine whether PDGF-BB could promote the healing of MCL. Results There were significantly more fibroblasts formed during the ligament healing process in group A than in group B (213.44±15.32 vs. 180.42±12.78, Plt;0.01). The fibroblasts were more mature andmore regularlyarranged in group A than in group B. The type, content, and crosslink of the collagen were improved to a greater extent in group A than in group B (Plt;0.01). Conclusion PDGF can promote the healing of the injured ligament.
Objective To study the effect of serum rich in growth factors (SRGF) derived from plateletrich plasma (PRP) on the biological function of human and rat osteoblast.Methods PRP and platelet-poor plasma (PPP) obtained from healthy human and SD rat were activated by thrombin toget SRGF and serum poor in growth factors (SPGF). The level of TGFβ1 and PDGF-AB in human-SRGF and SPGF were assayed by enzyme-linked immunoassay(ELISA). Rat and human osteoblast were cultured and identified. Rat osteoblasts were treated with 5% rat-SRGF, 5% rat-SPGF and serumfree F12 medium, respectively. And human osteoblast were treated with 5% human-SRGF, 5% human-SPGF and serumfree DMEM. Cellular mitogenic activity was evaluated by thiazoly blue (MTT) colorimetric assay at 24, 48, 72 and 96hours.Results The level of TGF-β1 in human-SRGF was 307.67±35.57 ng/ml, and that of PDGF-AB was 52.76±7.89 ng/ml. The proliferation of rat and human osteoblast were promoted after treated with rat-SRGFand human-SRGF, respectively. In rat osteoblast groups, there were significant differences in absorbency between ratSPGF group and rat-SRGF group at 48 and 96 hours(Plt;0.05). In human osteoblast groups, the differences between human-SPGF group and human-SRGF group were significant at 48, 72 and 96 hours(Plt;0.05). The proliferation of these two kinds of osteoblasts almost stopped in serum-free medium, and the differences in absorbency , compared with othergroups，were significant (Plt;0.05). Conclusion High quality of PRP can be achieved by the improved method and SRGF is capable of up-regulating the proliferation of rat osteoblast and human osteoblast.
OBJECTIVE: To investigate the expression and distribution of platelet derived growth factor receptor-beta(PDGFR-beta) in normal skin and keloid and to discuss its biological function in keloid formation. METHODS: 1. To detect the expression and distribution of PDGFR-beta in normal skin and keloid tissue by immunohistochemistry; 2. To detect the receptor expression in vitro by Flow cytometry (FCM); 3. To detect the subcellular distribution of receptor by Laser confocal microscope. RESULTS: 1. Immunohistochemistry showed that normal skin and keloid tissue were almost the same in expression but different in distribution of PDGFR-beta; 2. There was more expression of PDGFR-beta in normal fibroblasts than that in keloid fibroblasts in vitro by FCM; 3. Laser confocal microscope revealed that the PDGFR-beta concentrated on the surface of cell membrane in keloid fibroblasts, but in normal skin fibroblasts, the receptors were coagulated on the nuclear membrane and intranucleus. CONCLUSION: Compared with the fibroblasts in vivo, there was a difference of the PDGFR-beta expression in fibroblasts in vitro, more expression of PDGFR-beta in normal fibroblast than that in keloid fibroblast in vitro; and the subcellular distribution of PDGFR-beta was different in normal skin and keloid fibroblasts. The characteristics of the expression and distribution of PDGFR-beta in keloid may contribute to the formation of keloid.
【摘要】 目的 观察t(4; 22)致血小板源性生长因子受体α（the platelet-derived growth factor receptor alpha， PDGFRA）异常的髓系/淋巴系肿瘤的临床特点。 方法 对2010年6月收治的1例t(4; 22)致PDGFRA异常的髓系/淋巴系肿瘤患者的临床资料进行回顾性分析，并对其临床特点、实验室检查、诊断、治疗进行总结。 结果 该疾病临床表现及骨髓涂片检查类似慢性粒细胞白血病（chronic myelogenous leukemia，CML），但无CML特征性Ph染色体和（或）BCR/ABL融合基因，而细胞遗传学检测显示4号与22号染色体易位，诊断为t(4; 22)致PDGFRA异常的髓系/淋巴系肿瘤。采用羟基脲及干扰素治疗后可获得完全血液学缓解。 结论 t(4; 22)致PDGFRA异常的髓系/淋巴系肿瘤是一类罕见疾病，临床表现与CML相似，t(4; 22)及BCR/PDGFRA融合基因阳性是诊断该类疾病的关键。【Abstract】 Objective To observe the clinical features of myeloid and lymphoid neoplasms with t (4; 22) induced abnormalities of the platelet-derived growth factor receptor alpha (PDGFRA) to increase the identification and reduce the misdiagnosis. Methods The clinical data of one patient with myeloid and lymphoid neoplasm with t (4; 22) induced abnormalities of PDGFRA diagnosed in June 2010 was retrospectively analyzed. We summarized the clinical features, morphology, genetics, diagnostic criteria and therapy about this kind of disease. Results The patient had a clinical manifestation and bone marrow smear result of chronic myelogenous leukemia (CML). But the result of genetic analysis found no translocation of chromosomes 9 and 22 juxtaposing BCR and ABL gens. Cytogenetic analysis showed an abnormal karyotype with rearrangement of chromosomes 4 and 22. So the patient was diagnosed myeloid and lymphoid neoplasms with t (4; 22) induced abnormalities of PDGFRA. After receiving interferon and hydroxyurea, the patient achieved complete hematologic remission. Conclusion Myeloid and lymphoid neoplasms with t (4; 22) induced abnormalities of PDGFRA is a rare kind of disease. Its clinical feature is similar to that of CML. The key of diagnosis is genetics.