OBJECTIVE

To determine whether pulmonary vascular bed contributes to the development of in situ thrombosis and vascular remodelling in secondary pulmonary hypertension (SPH) via changes in its local secretory activities.

METHODS

Seventy-one patients with the diagnosis of secondary pulmonary hypertension (38 females, mean age 40.36+/-1.05 years) were included in the study. Selective right and left heart catheterization was performed to each patient for diagnostic purposes. Blood samples obtained from left ventricle (LV) and pulmonary artery (PA) of each patient were analyzed for levels of plasminogen activator inhibitor-1 (PAI-1), platelet derived growth factor (PDGF), vascular endothelial growth factor (VEGF), D-dimer, von Willebrand factor (vWF), protein-C, antithrombin-III, fibrinogen, and plasminogen. Results were compared between LV and PA. Correlation analysis between each parameter and mean pulmonary artery pressure (MPAP) was performed.

RESULTS

Although mean level of VEGF in LV and PA were found to be in normal range, it was significantly higher in LV than in PA (p<0.001). Mean PDGF and D-dimer levels, which remained in normal range were also higher in LV (p<0.001 and p<0.001, respectively) than in PA;.vWF showed similar degree of elevation in both LV and PA. Only one parameter, PAI-1, was found to be significantly higher in PA than in LV (p=0.012). Antithrombin-III, protein C, plasminogen, and fibrinogen levels showed no significant differences between two chambers. They also remained in normal range, except for fibrinogen, which was slightly elevated in both LV and PA. Correlation analysis revealed strong positive correlation between D-dimer level in both LV and PA and MPAP (r=0.775, p<0.001 and r=0.649, p<0.001, respectively).

CONCLUSIONS

In SPH, pulmonary vascular bed shows increased thrombotic, hypofibrinolytic, and proliferative activities, which are partially related to the severity of illness.

Platelets are involved in atherogenesis, partly through the release of smooth muscle cell mitogens and chemoattractants such as platelet-derived growth factor (PDGF). The St Thomas' Atherosclerosis Regression Study (STARS) demonstrated that a cholesterol-lowering diet induced angiographic regression of coronary artery disease in 74 men. Serum PDGF concentrations were not different at the end of the study between patients randomised to receive diet, with or without cholestyramine: usual care (UC) median 53.6 pM (interquartile range 15.3), diet (D) 60.0 pM (29.7), diet and cholestyramine (DC) 51.5 pM (13.0), 2p = 0.23. Similarly, mean platelet volume (MPV), a marker of platelet function, did not differ between the three groups: UC 8.0 fl (1.1), D 8.6 fl (0.8), DC 9.0 fl (1.4), 2p = 0.16. PDGF concentrations and MPV did not correlate with lipid concentrations or angiographic indices of regression. These findings suggest that platelet function, as measured by PDGF and MPV, does not change during, or contribute to, dietary-induced regression of coronary artery disease.

Objective: To detect the effect of brain cytoplasmic RNA 1 (BCYRN1) on the proliferation and migration of airway smooth muscle cells (ASMCs) in rat model of asthma. Methods: Male SD rats were randomly divided into control group and asthma group (n=10 each). The ovalbumin (OVA) model was constructed in asthma group. Real time-qPCR was performed to detect the level of BCYRN1 in the ASMCs separated from the airway tissue of these rats. Then 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2, 4-disulfophenyl)-2H-tetrazolium (WST-1) assay, roche real-time cell analyzer assay and Transwell cell migration assay were performed to detect the viability/proliferation and migration of ASMCs which were transfected with Ad-BCYRN1.Platelet-derived growth factor (PDGF)-BB was used to treat ASMCs to induce proliferation and migration, and the level of BCYRN1 was examined.The viability/proliferation and migration of ASMCs treated with PDGF-BB and transfected with si-BCYRN1 were detected. Inspiratory resistance and expiratory resistance were measured in rats with BCYRN1 knockdown.Briefly, rats were randomly divided into four groups: control (group A), sensitization + Ad-GFP (group B), sensitization + AdSM22α-siBCYRN1 (group C), control + Ad-SM22α-siBCYRN1 (group D) (n=10 each). The corresponding adenovirus vectors were sent to lung of group B, group C and group D through nasal spray. The OVA model was constructed in group B and group C. The rats in group A and group D were treated with saline.After 24 h of the last treatment with OVA or saline, rats of each group were given tracheal intubation, connected with breathing machine. Rats were injected with methacholine to measure the inspiratory resistance and expiratory resistance. Results: The level of BCYRN1 in ASMCs separated from rats in asthma group and in ASMCs treated with PDGF-BB was 3.60±0.45 and 3.53±0.35, respectively, significantly higher than those of the corresponding control (both P<0.01). Ad-BCYRN1 significantly increased the expression of BCYRN1 in ASMCs. The cell viability and proliferation rates of ASMCs transfected with Ad-BCYRN1 increased 1.75-and 1.47-fold compared to those of the control group, respectively (P<0.01); mobility increased 2.42-fold compared to that of the control group (all P<0.01). BCYRN1 knockdown reversed the increasing proliferation and migration of ASMCs induced by PDGF-BB. The cell proliferation rate and cell migration number in the PDGF-BB treatment group were (4.87±0.21)% and 80.00±5.00, respectively, which were significant higher than those in the si-BCYRN1 transfected group ((3.63±0.21)% and 25.33±2.52, all P<0.01). BCYRN1 knockdown reduced the inspiratory resistance and expiratory resistance in sensitization + Ad-SM22α-siBCYRN1 group. When the concentration of acetylcholine reached 1 mg/kg, the inspiratory resistance in the group A, group B, group C, and group D were 8.27±0.21, 25.40±0.56, 12.07±0.67 and 8.40±0.46 cmH2O·s·ml-1, and expiratory resistance were 13.30±0.56, 38.37±1.33, 16.40±0.56 and 13.40±0.46 cmH2O·s·ml-1, respectively (all P<0.01). Conclusion: Overexpression of BCYRN1 promotes the proliferation and migration of ASMCs in rat model of asthma.

To observe the effect of Fengshi Qutong Capsules(FSQTC) on angiogenesis of rat aortarings and in knee joint synovium of type Ⅱ collagen-induced arthritis(CIA) rats. The blood vessel of aorta rings of normal SD rats were induced by vascular endothelial growth factor(VEGF) 20 μg·L~(-1 )in vitro, and were treated with FSQTC(0.02, 0.1 and 0.5 μg·L~(-1)) continuously for 9 days. The number, length and area of neovascularization of the vascular ring were measured. SD rats were immunized to establish collagen-induced arthritis. CIA rats were treated with FSQTC(0.25, 0.5, 1 g·kg~(-1)·d~(-1)) and methotrexate(0.2 mg·kg~(-1)·d~(-1)) daily for 19 days. Histopathological examination(HE) was performed to observe the vascular morphology and vascular density in the synovial membrane of the inflamed joint. Immunohistochemistry was performed to observe the expression of platelets-endothelial cell adhesion molecule(CD31), VEGF and VEGF receptor 2(VEGFR_2)in the synovium. Immunofluorescence was performed to observe the expression of CD31 and α smooth muscle actin(αSMA) in synovial membrane.TGF-β, PDGF and VEGFR_2 in serum were detected by enzyme-linked immunosorbent assay. The number, branch length and area of blood vessels of aorta rings were significantly increased induced by VEGF, and FSQTC could significantly reduce the number, branch length and area of blood vessels. Compared with the normal group, the vascular density, CD31 positive expression, CD31~+/αSMA~- immature and total vascular positive expression in the synovial membrane of the model group were significantly increased, and so as VEGF and VEGFR_2 in the synovium. The VEGFR_2, TGF-β and PDGF in sera were also significantly increased in model group. FSQTC reduced the synovial vascular density and inhibited the positive expression of CD31, CD31~+/αSMA~- immature blood vessels and total vascular. FSQTC has no significant effect on CD31~+/αSMA~+mature blood vessels. FSQTC also negatively inhibited the expression of VEGF, VEGFR_2, TGF-β and PDGF in synovial membrane and/or sera. The effect of methotrexate is similar with to the high dose group. Our results demonstrated that FSQTC could inhibit the angiogenesis of synovial tissue in CIA rats and of aortaring in rats, which is related to the reduction of angiogenesis regulatory factor.

Increased airway smooth muscle (ASM) cell mass and secretory functions are characteristics of airway inflammatory diseases, such as asthma. To date, there are no effective therapies to combat ASM cell proliferation, which contributes to bronchoconstriction and airway obstruction. Growth factors such as platelet-derived growth factor (PDGF) and the activation of the ERK1/2 are major regulators of ASM cell proliferation and airway remodeling in asthma. However, given the ubiquitous expression and multiple functions of ERK1/2, complete inhibition of ERK1/2 using ATP-competitive inhibitors may lead to unwanted off-target effects. Alternatively, we have identified compounds that are designed to target substrate docking sites and act as function-selective inhibitors of ERK1/2 signaling. Here, we show that both function-selective and ATP-competitive ERK1/2 inhibitors are effective at inhibiting PDGF-mediated proliferation, collagen production, and IL-6 secretion in ASM cells. Proteomic analysis revealed that both types of inhibitors had similar effects on reducing proteins related to TGF-β and IL-6 signaling that are relevant to airway remodeling. However, function-selective ERK1/2 inhibitors caused fewer changes in protein expression compared with ATP-competitive inhibitors. These studies provide a molecular basis for the development of function-selective ERK1/2 inhibitors to mitigate airway remodeling in asthma with defined regulation of ERK1/2 signaling.-Defnet, A. E., Huang, W., Polischak, S., Yadav, S. K., Kane, M. A., Shapiro, P., Deshpande, D. A. Effects of ATP-competitive and function-selective ERK inhibitors on airway smooth muscle cell proliferation.

<AbstractText>To observe the change of neovascular morphology an<em>d</em> angiogenesis relate<em>d</em> factors in the ischemic cerebral area after cerebral infarction an<em>d</em> the intervention effect of electroacupuncture (EA).</AbstractText><p>(<em>d</em>iv><b>METHODS</b></<em>d</em>iv>Wistar rats were ran<em>d</em>omly <em>d</em>ivi<em>d</em>e<em>d</em> into mo<em>d</em>el group(<i>n</i>=90), EA group(<i>n</i>=90), sham operation group(<i>n</i>=90) an<em>d</em> control group(<i>n</i>=10). The first three groups were further <em>d</em>ivi<em>d</em>e<em>d</em> into 1 h, 3 h, 6 h, 9 h, 12 h, 24 h, 3 <em>d</em>, 7 <em>d</em> an<em>d</em> 12 <em>d</em> subgroups(<i>n</i>=10 in each subgroup). The cerebral infarction mo<em>d</em>el was establishe<em>d</em> by mi<em>d</em><em>d</em>le cerebral artery occlusion(MCAO). EA(15 Hz, 2 mA) was applie<em>d</em> to "Shuigou"(GV26) for 20 min in the EA group. The 1, 3, 6, 9, 12, 24 h subgroups were treate<em>d</em> imme<em>d</em>iately after mo<em>d</em>eling, the 3, 7, 12 <em>d</em> subgroups were treate<em>d</em> once <em>d</em>aily for 3, 7 or 12 <em>d</em>ays. The neovascular en<em>d</em>othelial cells were <em>d</em>isplaye<em>d</em> by immunofluorescence <em>d</em>ouble labeling staining. Quantitive real-time PCR an<em>d</em> Western blot were applie<em>d</em> to <em>d</em>etect the changes of basic fibroblast growth factor (bFGF), angiogenin (Ang) -1, 2, platelet-<em>d</em>erive<em>d</em> growth factor b (<em>PDGF</em>-b) in ischemic brain tissue, respectively.</p><p>(<em>d</em>iv><b>RESULTS</b></<em>d</em>iv>After mo<em>d</em>eling, CD31 an<em>d</em> Ki67 positive cells were first observe<em>d</em> at 24 h in the mo<em>d</em>el group, an<em>d</em> reache<em>d</em> the peak at 3 <em>d</em>, <em>d</em>ecrease<em>d</em> at 7 <em>d</em>. While in the EA group, the CD31 an<em>d</em> Ki67 positive cells were first observe<em>d</em> at 12 h, an<em>d</em> reache<em>d</em> the peak at 3 <em>d</em>, an<em>d</em> gra<em>d</em>ually <em>d</em>ecrease<em>d</em> until 12 <em>d</em>. Compare<em>d</em> with the control group, the mRNA expressions of bFGF at 9 h-12 h, Ang-1 at 12 h-12 <em>d</em>, Ang-2 at 1 h-12 <em>d</em> an<em>d</em> <em>PDGF</em>-b at 1 h, 6 h, 9 h, 24 h-12 <em>d</em> were increase<em>d</em> in the mo<em>d</em>el group(<i>P</i><0.01, <i>P</i><0.05). After EA, the mRNA expressions of bFGF at 24 h-12 <em>d</em>, Ang-1 at 3 <em>d</em>-12 <em>d</em>, Ang-2 at 3 h-24 h an<em>d</em> <em>PDGF</em>-b at 3 h, 6 h, 3 <em>d</em>-12 <em>d</em> were increase<em>d</em>(<i>P</i><0.05, <i>P</i><0.01). In comparison with the control group, the proteins of bFGF at 24 h, Ang-1 at 6 h-12 <em>d</em>, Ang-2 at 1 h-12 <em>d</em> an<em>d</em> <em>PDGF</em>-b at 1 h-7 <em>d</em> were increase<em>d</em> in the mo<em>d</em>el group(<i>P</i><0.05, <i>P</i><0.01). After EA, the proteins of bFGF at 3 <em>d</em>-12 <em>d</em>, Ang-1 at 3 <em>d</em>-12 <em>d</em>, Ang-2 at 3 h-12 h an<em>d</em> <em>PDGF</em>-b at 6 h, 3 <em>d</em>-12 <em>d</em> were increase<em>d</em> compare<em>d</em> with the mo<em>d</em>el group(<i>P</i><0.05, <i>P</i><0.01).</p><AbstractText>EA can up-regulate the expression of angiogenesis-relate<em>d</em> factors in MCAO rats, which has an important role in the establishment of bloo<em>d</em> vessel regeneration an<em>d</em> collateral circulation, an<em>d</em> thus promote the recovery of neurological function.</AbstractText>

Platelet-derived growth factor (PDGF) D has been reported to be active in fibroblasts, and in areas of myocardial infarction. In this longitudinal study we evaluated the association between PDGF-D polymorphism and cardiovascular mortality, and attempted to discover whether specific genotype differences regarding risk could be observed, and if gender differences could be seen.

Four hundred seventy-six elderly community participants were included in this study. All participants underwent a clinical examination, echocardiography, and blood sampling including PDGF-D single nucleotide polymorphism (SNP) analyses of the rs974819 A/A, G/A and G/G SNP. The follow-up time was 6.7 years.

No specific genotype of rs974819 demonstrated increased cardiovascular mortality in the total population, however, the male group with genotypes A/A and G/A demonstrated an increased risk that persisted in a multivariate evaluation where adjustments were made for well-known cardiovascular risk factors (2.7 fold compared with the G/G genotype). No corresponding finding was observed in the female group.

We report here for the first time that the genotypes G/A or A/A of the SNP rs974819 near PDGF-D exhibited a 2.7 fold increased cardiovascular mortality risk in males. Corresponding increased risk could not be observed in either the total population and thus not in the female group. However, the sample size is was small and the results should be regarded as hypothesis-generating, and thus more research in the field is recommended.

OBJECTIVE

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.

METHODS

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.

RESULTS

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).

CONCLUSIONS

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.

OBJECTIVE

To investigate the clinical implication of platelet-derived growth factor (PDGF)-D and PDGF-beta in IgA nephropathy in childhood.

METHODS

Forty-seven children with IgA nephropathy and 26 controls were enrolled for study, and their serum, urine and renal biopsy specimens were examined. The patients were divided into control group [including serum, urine specimens of 13 healthy children and 13 renal biopsy samples of non-IgA nephropathy in children], mild proliferation (MP) group (13 patients), focal proliferation (FP) group (19 patients), and proliferation sclerosis (PS) group (15 patients). Enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry were used to determine contents of PDGF-D, PDGF-beta and PDGF-B in blood, urine and renal tissues. The levels of 24-hour urinary protein excretion, serum albumin (Alb), serum blood urea nitrogen (BUN) and creatinine (Cr) were also determined.

RESULTS

Compared with control group, levels of PDGF-D and PDGF-B were progressively elevated in blood and urine of IgA nephropathy children with increase in severity of glomerular damage (all P<0.01). Serum as well as urinary PDGF-D and PDGF-B levels were positively correlated with 24-hour urinary protein excretion (PDGF-D blood: r=0.546, urine: r=0.760; PDGF-B blood: r=0.634, urine: r=0.577, respectively, P<0.01), while negatively correlated with serum Alb levels in IgA nephropathy patients (PDGF-D blood: r=-0.649, urine: r=-0.528; PDGF-B blood: r=-0.613, urine: r=-0.531, respectively, P<0.01). Contents of PDGF-D and PDGF-beta in renal tissue were much higher than those of control group (P<0.01). Along with the increase in severity of glomerular pathology, their contents increased gradually. PDGF-B was only significantly expressed in renal tissue in FP group and PS group.

CONCLUSIONS

PDGF-D might significantly enhance the development of mesangial proliferation and tubulointerstitial fibrosis. In comparison with PDGF-B, PDGF-D appears to reflect more sensitive to the severity and prognosis of IgA nephropathy.

This study aimed to develop a functionally graded membrane (FGM) to prevent infection and promote tissue regeneration. Poly(l-lactide-co-d,l-lactide) encapsulating platelet-derived growth factor (PDLLA-PDGF) or metronidazole (PDLLA-MTZ) was electrospun to form a nanofibrous layer on the inner or outer surface of a clinically available collagen membrane, respectively. The membrane was characterized for the morphology, molecule release profile, in vitro and in vivo biocompatibility, and preclinical efficiency for alveolar ridge regeneration. The PDLLA-MTZ and PDLLA-PDGF nanofibers were 800-900 nm in diameter, and the thicknesses of the functional layers were 20-30 μm, with sustained molecule release over 28 days. All of the membranes tested were compatible with cell survival in vitro and showed good tissue integration with minimal fibrous capsule formation or inflammation. Cell proliferation was especially prominent on the PDLLA-PDGF layer in vivo. On the alveolar ridge, all FGMs reduced wound dehiscence compared with the control collagen membrane, and the FGM with PDLLA-PDGF promoted osteogenesis significantly. In conclusion, the FGMs with PDLLA-PDGF and PDLLA-MTZ showed high biocompatibility and facilitated wound healing compared with conventional membrane, and the FGM with PDLLA-PDGF enhanced alveolar ridge regeneration in vivo. The design represents a beneficial modification, which may be easily adapted for future clinical use.

Objective: To observe the effect of electroacupuncture (EA) of "Weizhong" (BL40) on the expression of platelet-derived growth factor (PDGF)-CC, PDGF receptor (PDGFR)α and matrix metalloproteinase-1 (MMP-1) in rats with lumbar multifidus muscle injury (LMMI) so as to study its mechanisms underlying improvement of skeletal muscle injury.

Methods: Fifty-four male SD rats were randomly divided into normal group (n=6), model group (n=24) and EA group (n=24), and the latter two groups were further divided into four subgroups (1, 3, 5 and 7 days), with 6 rats in each group. The LMMI model was established by injection of 0.5% bupivacaine (BPVC, 100 μL×4) into the multifidus along the L4 and L5 spinous process. EA (2 Hz/50 Hz, 1 mA) was applied to bilateral "Weizhong"(BL40) for 20 min, once daily for 1, 3, 5 and 7 days respectively, from the first day on after modeling. Histopathological changes of the left multifidus muscle were observed after H.E. staining, and the expression of PDGF-CC, PDGFR-α and MMP-1 proteins in the right multifidus was observed by Western blot.

Results: Compared with the normal group, the expression levels of PDGF-CC protein in the model subgroup 1 d, 3 d and 7 d were significantly decreased (P<0.05), and those of PDGFR-α and MMP-1 proteins in the model subgroup 5 d and 7 d, and PDGF-CC protein in the model subgroup 5 d significantly increased (P<0.05). In comparison with the model subgroups, the expression levels of PDGF-CC in the EA subgroup 3 d, 5 d and 7 d, PDGFR-α in the EA subgroup 5 d, and MMP-1 in the EA group 3 d and 5 d were significantly increased or significantly further increased (P<0.05). H.E. staining showed different shapes and uneven sizes, with large area of damage, enlarged muscle space and inflammatory cell infiltration in the model group, which was relatively milder in the EA subgroups particularly in subgroup 5 d and 7 d.

Conclusion: EA stimulation of BL40 for about 5 days has a positive effect in promoting the repair of the injured multifidus muscle in LMMI rats, which may be related to its function in up-regulating the expression of muscular PDGF-CC, PDGFR-α and MMP-1 proteins.

目的：观察电针“委中”穴对腰多裂肌损伤大鼠血小板衍生生长因子CC(PDGF-CC)、血小板衍生生长因子受体α(PDGFR-α)、基质金属蛋白酶-1(MMP-1)表达的影响, 从骨骼肌损伤后修复的角度研究电针及不同治疗周期对其的影响。方法： SD大鼠按随机数字表法分为正常组6只、模型组24只和电针组24只。其中, 模型组和电针组再分1、3、5、7 d 4个亚组, 每组6只, 采用布比卡因致腰多裂肌损伤。电针组电针“委中”穴, 每次治疗20 min, 每日治疗1次, 分别干预1、3、5、7 d。取大鼠左侧多裂肌用于HE染色观察多裂肌损伤变化, 右侧多裂肌用于Western blot检测PDGF-CC、PDGFR-α、MMP-1的含量。结果： 造模后, 与正常组相比, 模型组肌纤维出现大面积变形, 肌间隙增大；同一时点, 电针组肌纤维恢复程度优于模型组；电针5、7 d组肌纤维恢复程度优于电针1、3 d组。与同时点正常组相比, 模型1、3、7 d组PDGF-CC表达降低(P<0.05), 模型5 d组升高(P<0.05)；模型5、7 d组PDGFR-α表达升高(P<0.05)；模型3 d组MMP-1表达降低(P<0.05), 模型5、7 d组表达升高(P<0.05)。与各时点模型组相比, 电针3、5、7 d组PDGF-CC表达升高(P<0.05)；电针5 d组PDGFR-α表达升高(P<0.05)；电针3、5 d组MMP-1表达升高(P<0.05)。电针组各亚组比较, 电针5 d组PDGF-CC、PDGFR-α表达高于电针1、3、7 d组(P<0.05)；电针5、7 d组MMP-1表达高于电针1、3 d组(P<0.05)。结论： 电针“委中”穴能促进损伤多裂肌的修复, 并且在治疗5 d时有较好疗效, 其机制可能与提高多裂肌中PDGF-CC、PDGFR-α和MMP-1的表达相关。.

Keywords: Electroacupuncture; Lumbar multifidus muscle injury; MMP-1; PDGFR-α; Platelet-derived growth factor (PDGF)-CC; stroke.

OBJECTIVE

The aim of this study is to investigate the role of platelet-derived growth factor (PDGF) in the pathogenesis of sinonasal polyps.

METHODS

Study group (groups 1-3) consisted of nasal polyp samples of patients with sinonasal polyps and the control group consisted of inferior turbinate samples of patients without nasal polyp. In group 1, 14 specimens from ethmoid sinus; in group 2, 10 specimens from nasal cavity; in group 3, 10 specimens from maxillary sinus; and in group 4 (control), 9 specimens from inferior turbinate were included. By immunohistochemical staining technique, the PDGF positivity index (PI) in mucosal layers and in the inflammatory cells were assessed at the epithelium (EP), subepithelial layer of lamina propria (SE), and deep paraglandular layer of the mucosa (D).

RESULTS

Polymorphonuclear cell (PMNC)-percentage (%) values of ethmoid and maxillary sinus, and the PDGF PI from all cells of ethmoid sinus and nasal cavity were significantly higher than those of the control group. As mononuclear cell-% (MNC-%) increased, the PDGF_EP_basal PI, PDGF_SE_endothelial PI, and PDGF_D_endothelial PI decreased. As PMNC-PDGF PI increased, the PDGF_D_perivascular PI decreased and PDGF_D_endothelial PI increased. As PDGF-MNC PI increased, the PDGF_EP_apical PI, PDGF_SE_endothelial PI, and PDGF_D_endothelial PI decreased. As PDGF-all cells (PMNCs, MNCs, and fibroblasts) PI increased, the PDGF_EP_basal PI and PDGF_D_endothelial PI decreased, and the PDGF_D_perivascular PI increased.

CONCLUSIONS

We concluded that the PDGF systems play important roles in polyp pathogenesis. Fibroblast-derived PDGF may be more important than MNC-derived PDGF in polyp developing process. Increased perivascular-PDGF-PI in deep layers of the mucosa may result in sinonasal polyp formation by causing an increase in vascular permeability and extracellular edema, and thus promoting migration of inflammatory cells to extracellular area. Tissue oxygenization may be important for the initiation of PDGF release system. Because of this reason, nasal obstruction should be medically treated earlier, and, if necessary, by surgical approaches.

Objective: To observe and compare the cytological and biological differences between human normal and degenerated nucleus pulposus (NP), and to investigate the repair effect of insulin-like growth factor 1 (IFG-1) and platelet derived growth factor (PDGF) on human degenerated NP.

Methods: Human degenerative and normal NP tissues were obtained from operative patients, a portion of which were processed into tissue sections and HE staining was performed to observe the morphological changes of nucleus pulposus cells (NPCs) before and after degeneration of NP. Immunohistochemistry staining was used to determine the expression levels of collagen type Ⅰ, collagen type Ⅱ, B-cell lymphoma 2 (Bcl-2), Bcl-2 associated X (Bax) proteins. Another portion of tissues were isolated and cultured and NPCs morphology was observed under inverted microscope. Western blot analysis was used to detect collagen type Ⅱ protein expression. Then, the gene transfection experiments were launched, including 4 groups, with group A designed as degenerated NPCs only, and groups B, C, and D of degenerated NPCs transfected with IGF-1 gene lentiviral particles, PDGF gene lentiviral particles, and lentiviral particles carrying IGF-1 and PDGF double genes, respectively. At 21 days after transfection, the cell morphology of each group was observed under inverted microscope, the positive rates of IGF-1 and PDGF of each group were measured by flow cytometry, and the expression of collagen type Ⅱ protein was detected by using immunohistochemistry staining and Western blot.

Results: HE staining showed that there were a large number of notochordal cells and a small number of chondrocytes in the central NP tissue of normal group, while the NPCs in degeneration group were significantly reduced, and a large proportion of fibrocartilage tissues were found in NP tissue. Immunohistochemistry staining showed that the percentages of collagen type Ⅰ and Bax protein-positive cells in degeneration group were significantly higher than those of normal group, while the percentages of collagen type Ⅱ and Bcl-2 protein-positive cells were significantly lower than those of normal group ( P<0.05). Western blot showed that the relative expression level of collagen type Ⅱ protein in degeneration group was significantly lower than that in normal group ( t=65.493, P=0.000). At 21 days after gene transfection, compared with group A, the cell viability of groups B, C, and D increased and the morphology became more regular. Flow cytometry showed that the percentages of IGF-1-positive cells in groups B and D were significantly higher than that in group A, and the percentages of PDGF-positive cells in groups C and D were significantly higher than that in group A ( P<0.05). Immunohistochemistry staining showed that the positive stainings of collagen type Ⅱ in groups A, B, C, and D was (±), (+), (+), and (++), respectively. Western blot showed that the relative expression of collagen type Ⅱ protein in groups A, B, C, and D increased by degrees, and the differences between groups were significant ( P<0.05).

Conclusion: Both IGF-1 and PDGF can reverse the degeneration of intervertebral discs NPCs and they have synergistic effects, providing experimental basis for its application in clinical treatment approaches for degenerative disc disease.

目的: 观察比较人正常和退变髓核的细胞学和生物学差异，并研究 IFG-1 和 PDGF 对人退变髓核的修复作用。.

方法: 取患者自愿捐赠的人退变及正常髓核组织，一部分制作成组织切片，进行 HE 染色观察髓核退变前后形态变化，免疫组织化学染色观察Ⅰ型胶原、Ⅱ型胶原、B 淋巴细胞瘤 2（B-cell lymphoma 2，Bcl-2）、Bcl-2 相关 X（Bcl-2 associate X，Bax）蛋白表达；另一部分髓核组织进行细胞培养，倒置显微镜观察细胞形态，Western blot 检测Ⅱ型胶原蛋白表达。然后进行基因转染实验，分为 4 组，A 组为退变髓核细胞；B、C、D 组分别用 IGF-1 基因慢病毒颗粒、PDGF 基因慢病毒颗粒和携带 IGF-1、PDGF 双基因的慢病毒颗粒转染退变髓核细胞。转染 21 d，倒置显微镜观察各组细胞形态，流式细胞仪测定各组细胞 IGF-1 和 PDGF 阳性率，免疫组织化学染色和 Western blot 检测各组细胞Ⅱ型胶原蛋白表达。.

结果: HE 染色示，正常组中央髓核组织可见大量脊索细胞和少量类软骨细胞；而退变组髓核细胞明显减少，且髓核组织内可见大量纤维软骨组织。免疫组织化学染色示，退变组Ⅰ型胶原、Bax 蛋白阳性细胞百分比显著高于正常组，Ⅱ型胶原、Bcl-2 蛋白阳性细胞百分比显著低于正常组，差异均有统计学意义（ P<0.05）。Western blot 检测示，退变组Ⅱ型胶原蛋白相对表达量显著低于正常组（ t=65.493， P=0.000）。基因转染后 21 d，与 A 组比较，B、C、D 组细胞活性增加，形态变得较为规则。流式细胞仪检测示 B、D 组 IGF-1 阳性率较 A 组明显增高，C、D 组PDGF 阳性率较 A 组明显增高（ P<0.05）。免疫组织化学染色观察示，A、B、C、D 组Ⅱ型胶原蛋白阳性染色情况依次为（±）、（+）、（+）、（++）。Western blot 检测示，A、B、C、D 组Ⅱ型胶原蛋白相对表达量依次增加，各组间比较差异均有统计学意义（ P<0.05）。.

结论: IGF-1 与 PDGF 均能逆转椎间盘髓核细胞的退变且二者具有协同作用，为其今后应用于临床治疗椎间盘退变性疾病提供了实验依据。.

Keywords: Intervertebral disc degeneration; gene transfection; insulin-like growth factor 1; nucleus pulposus; platelet derived growth factor.

The specific cytokines that regulate pediatric acute respiratory distress syndrome (PARDS) pathophysiology remains unclear. Here, we evaluated the respiratory cytokine profile in PARDS to identify the molecular signatures associated with severe disease. A multiplex suspension immunoassay was used to profile 45 cytokines, chemokines and growth factors. Cytokine concentrations were compared between severe and non-severe PARDS, and correlated with oxygenation index (OI). Partial least squares regression modelling and regression coefficient plots were used to identify a composite of key mediators that differentially segregated severe from non-severe disease. The mean (standard deviation) age and OI of this cohort was 5.2 (4.9) years and 17.8 (11.3), respectively. Early PARDS patients with severe disease exhibited a cytokine signature that was up-regulated for IL-12p70, IL-17A, MCP-1, IL-4, IL-1β, IL-6, MIP-1β, SCF, EGF and HGF. In particular, pro-inflammatory cytokines (IL-6, MCP-1, IP-10, IL-17A, IL-12p70) positively correlated with OI early in the disease. Whereas late PARDS was characterized by a differential lung cytokine signature consisting of both up-regulated (IL-8, IL-12p70, VEGF-D, IL-4, GM-CSF) and down-regulated (IL-1β, EGF, Eotaxin, IL-1RA, and PDGF-BB) profiles segregating non-severe and severe groups. This cytokine signature was associated with increased transcription, T cell activation and proliferation as well as activation of mitogen-activated protein kinase pathway that underpin PARDS severity.

BACKGROUND

Acute rejection is the single most important risk factor for the development of subsequent chronic rejection. Platelet-derived growth factor (PDGF) is a major mitogen that mediates mesenchymal cell proliferation in chronic rejection. Therefore, we investigated whether PDGF ligands and receptors are induced during acute renal allograft rejection in rat.

METHODS

Kidney transplantations were performed from Dark Agouti (DA) to Wistar-Furth (WF) rats, and syngenic controls were performed from DA to DA rats. Allografts were immunosuppressed with cyclosporine (CsA) 1.5 mg/kg/d subcutaneously or left untreated. Grafts were harvested at 1, 3, 5, and 7 days for histology and immunohistochemistry.

RESULTS

In syngenic grafts, no histological signs of acute rejection were seen and the expression of PDGF ligands and receptors remained almost nonexistent. In nontreated allografts, intense rejection resulted in graft necrosis in 7 days. Acute rejection was associated with the induction of all PDGF ligands and receptors (P<0.05 compared to syngenic controls). The expression of PDGF ligands and receptors was located mainly to graft-infiltrating macrophages but also to capillary endothelium and arteriolar smooth muscle cells. CsA significantly ameliorated acute rejection but failed to inhibit the induction of PDGF and its receptors in CsA-treated allografts.

CONCLUSIONS

Our results demonstrate that PDGF ligands and receptors are induced during acute rejection. PDGF may be induced directly as a reparative response to graft injury in acute rejection or indirectly by various inflammatory mediators released by graft-infiltrating inflammatory cells. This study indicates that PDGF ligands and receptors are already induced in acute rejection, which suggests a link between acute rejection and the subsequent development of chronic rejection.

OBJECTIVE

The aim of the study was to examine the effects of flufenamic acid on proliferation, clonogenic activity, migratory ability, cell-cycle distribution, and p44/42-mitogen-activated protein kinase (MAPK) expression on serum-stimulated human aortic smooth muscle cells (haSMCs) in vitro.

METHODS

HaSMCs were treated with flufenamic acid in three different doses (40 micromol/L, 200 micromol/L, 400 micromol/L) for 4 days, and then flufenamic-acid-free culture medium was supplemented every 4 days until day 20 after initial treatment. The growth kinetics were assessed. Cell-cycle analysis was performed by flow cytometry. The clonogenic activity was evaluated with use of colony formation assays. The migratory ability was investigated by stimulation with platelet derived growth factor (PDGF-BB) in 24 well plates with 8-microm pore membrane inserts. The p44/42 MAPK was detected by Western blot technique.

RESULTS

Flufenamic acid inhibited the proliferation (400 micromol/L treatment over 4 d; 179,700 +/- 49,800 vs 747,900 +/- 144,000; P <.001), clonogenic activity (400 micromol/L treatment over 4 d; 1 +/- 0.3 vs 50 +/- 1.4; P <.001) and migratory ability (400 micromol/L treatment over 4 d; 8 cells +/- 2 vs 48 cells +/- 15; P <.001) of haSMCs in a dose-dependent manner. Cell-cycle analysis revealed a G2/M-phase block (400 micromol/L treatment over 4 d; 28.9 +/- 1.5 vs 9.5 +/- 3.2; P <.001). The expression of p44/42 MAPK was reduced for a treatment with 400 micromol/L flufenamic acid (controls, 427 BLU +/- 0.305 vs treatment group, 190 BLU +/- 106; P <.05)

CONCLUSIONS

Flufenamic acid inhibits the proliferation and migration of haSMCs. Further experiments with animal models concerning stenosis and restenosis are necessary to evaluate the potential of this promising drug.

The IPLB-LdFB cell line from the fat body of the insect Lymantria dispar shows the presence of immunoreactive, platelet-derived growth factor (PDGF)-AB and transforming growth factor (TGF)-beta1 molecules, as well as the corresponding plasma membrane-like receptors, i.e. PDGFR-alpha, PDGFR-beta and TGFR-beta type II. Cytofluorimetric and morphological studies reveal that the reducing sugar 2-deoxy-D-ribose (dRib), an apoptotic agent for human cells, induces apoptosis in a concentration- and time-dependent manner even in IPLB-LdFB cells. PDGF-AB and TGF-beta1 partially counteract the effect of dRib, indicating a survival role of these factors in this apoptotic model of insect cells.

A plasminogen activator (PA) system is involved in ovulation, implantation, tumor invasion and metastasis. In order to clarify the regulation of this PA system in endometrial cells, we examined which agent affecting cellular function altered tissue-type plasminogen activator (t-PA) secretion by endometrial carcinoma cell line (KLE cells) in vitro. Triiodothyronine, retinoic acid, insulin, 8-bromo-cAMP, PDGF, IGF-I, basic FGF or TNF-alpha did not alter t-PA secretion while the activator of protein kinase C, phorbol myristate acetate (PMA) stimulated t-PA secretion in a dose-dependent fashion (10(-10)-10(-8) M). The time required to give a statistically significant increase in t-PA over control was 3 hours, and the maximal increase was seen after 24 hours of exposure. Another active phorbol ester, PDD also stimulated t-PA secretion while inactive forms of phorbol ester, 4 alpha-PDD and phorbol did not alter it. Cholera toxin or 8-bromo-cAMP did not affect t-PA secretion, but enhanced PMA-stimulated t-PA secretion. Cycloheximide and actinomycin D completely abolished PMA-stimulated t-PA secretion. These results suggest that (1) t-PA secretion in the endometrial carcinoma cell is modulated by a protein kinase C system, (2) This effect is through new RNA production and protein synthesis. (3) There is a complicated relationship between protein the kinase C and protein kinase A system as to the regulation of t-PA secretion. This would be a suitable model to clarify the PA system in endometrial cells.

Lesions of D-galactosamine (D-GalN)-induced hepatotoxicity resemble those of human acute viral hepatitis. This study investigated hepatic mesenchymal cells including hepatic stellate cells (HSCs) and myofibroblasts in D-GalN-induced hepatotoxicity. Rats, injected with D-GalN (800 mg/kg body weight, once, intraperitoneally) were examined on post single injection (PSI) at 8 hours and days 1 to 5. Lesions consisting of hepatocyte necrosis and reparative fibrosis were present diffusely or focally within the hepatic lobules on PSI days 1 and 2, and then the injury recovered on PSI days 3 and 5. Myofibroblasts expressing vimentin, desmin, and α-smooth muscle actin (α-SMA) were present in the lesions. Double immunofluorescence showed that myofibroblasts reacted simultaneously to vimentin/α-SMA, desmin/α-SMA, and desmin/vimentin; furthermore, myofibroblasts reacting to vimentin, desmin, and α-SMA also co-expressed glial fibrillary acidic protein (GFAP), a marker of HSCs. Additionally, GFAP-expressing myofibroblasts reacted to nestin and A3 (both are markers of immature mesenchymal cells). Cells reacting to Thy-1, a marker for immature mesenchymal cells, also appeared in fibrotic lesions. In agreement with the myofibroblastic appearance, mRNAs of fibrosis-related factors (TGF-β1, PDGF-β, TNF-α, Timp2, and Mmp2) increased mainly on PSI days 1 and 2. Myofibroblasts with expression of various cytoskeletal proteins were present in diffuse or focal hepatic lesions, and they might be derived partly from immature HSCs and from immature mesenchymal cells.

Keywords: D-GalN; hepatic stellate cells; liver fibrosis; myofibroblasts; rats; toxicologic pathology.

OBJECTIVE

To investigate the effect of local delivery of delta12-prostaglandinJ2-loaded poly (lactic-co-glycolic acid) (Δ(12)-PGJ2-NC) on growth factors expression and bone formation.

METHODS

Δ(12)-PGJ2-NC was prepared by the emulsion solvent diffusion method. The physical and chemical properties of the nanoparticles were evaluated by particle size analysis, transmission electron microscopy, drug-loading ratio and the in vitro release study. Then standardized transcortical defect (5.0 mm × 1.5 mm) was conducted in the femur of 48 male Wistar rats which were randomly divided into four groups (n = 12), S, K, F, and N. Thirty microliter of saline (S), unloaded nanoparticles (K), Δ(12)-PGJ2 (F) and Δ(12)-PGJ2-NC(N) in a collagen vehicle were delivered inside a titanium chamber fixed over the defect. Then, four subgroups were randomly divided in each group named as D3, D7, D14, and D28 (n = 3) according to the days 3, 7, 14, and 28 after the surgery. At days 3, 7, 14, and 28, the mRNA expression of the bone morphogenetic protein-6 (BMP-6), platelet-derived growth factor-B (PDGF-B) in defect aera was analyzed by real time quantitive-polymerase blotting. HE staining was employed to reveal new bone formation in weeks 2 and 4.

RESULTS

Δ(12)-PGJ2-NC appeared opalescent white and remained relatively stable, with an average particle size of (135.2 ± 0.85) nm. The images from transmission electron microscopy showed that Δ(12)-PGJ2-NC was spherical in shape and homogeneously distributed. The encapsulation efficiency of Δ(12)-PGJ2 with the poly (lactic-co-glycolic acid) (PLGA) nanocapsules was about 92%. The in vitro release of Δ(12)-PGJ2-NC at 37 °C showed a sustained fashion and the average accumulated amount was 30%, 52%, 77%, 91%, and 98% respectively, at 0.5, 1, 2, 4 and 6 h. Compared with the animals treated with saline, after dose of 100 mg/L Δ(12)-PGJ2 and Δ(12)-PGJ2-NC apllication, the mRNA expression level of BMP-6, PDGF-B increased significantly (P < 0.05, P < 0.001). The protein expression of BMP-6, Ephrin-B2 also was up-regulated. Histomorphometry revealed that new bone formation increased at the same dose of 100 mg/L. But the unloaded nanoparticles did not have the same effect (P>> 0.05).

CONCLUSIONS

A stable Δ(12)-PGJ2 loaded nanoparticle was successfully prepared. Δ(12)-PGJ2-NC may upregulate the expression of BMP-6, PDGF-B and Ephrin-B2, and promote new bone formation in bone defect area.

Platelet-derived growth factor (PDGF) and basic fibroblast growth factor (bFGF) have been implicated in myointimal proliferative arteriopathy, a lesion seen in monocrotaline-induced pulmonary hypertension (MIPH). The purpose of this study was to examine the expression of PDGF and bFGF in the lungs of rats given monocrotaline and to examine the effects of amrinone on the hearts and lungs of these rats. Twenty-four 26-day-old rats were randomized to receive either monocrotaline (approximately 3.6 mg/kg/d) or no monocrotaline and concomitantly to receive either amrinone (100 mg/kg/d) or no amrinone for 21 days. Lungs were examined for immunohistochemical evidence of PDGF and bFGF, and hearts were examined for effects of pulmonary hypertension and amrinone. Immunohistochemical staining of lungs showed no evidence of PDGF except in bronchioles. bFGF staining was similar between groups (no monocrotaline 25%, monocrotaline 27%, monocrotaline and amrinone 22%), and the staining was confined to the arterial walls. Rats given monocrotaline showed significantly greater right ventricular (RV) weight (0.13 +/- 0.02 g versus 0.23 +/- 0.04 g [mean +/- SD], P < 0.001), right ventricular/left ventricular (RV/LV) weight ratio (0.29 +/- 0.06 versus 0.59 +/- 0.1, P < 0.001), and lung/body weight ratio (0.006 +/- 0.001 versus 0.01 +/- 0.003, P < 0.05) than controls. Rats given monocrotaline and amrinone were not significantly different from rats given only monocrotaline with regard to RV weight, RV/LV weight ratio, or lung/body weight ratio. We conclude that the vasculopathy seen in MIPH is not associated with the presence of PDGF or bFGF, suggesting that other growth factors may mediate this process. The course of MIPH is not altered by amrinone.

OBJECTIVE

To study the effect of maternal vitamin D deficiency on lung morphogenesis and platelet-derived growth factor-A (PDGF-A) expression in rat offspring.

METHODS

Sprague-Dawley (SD) female rats were randomly divided into two groups: normal control and vitamin D deficiency, with 6 rats in each group. The vitamin D deficiecy group was kept away from light and fed with the forage without vitamin D. After 2 weeks, the rats were mated with normal SD male rats. The morphological changes of fetal rat lungs on day 20 of gestation and 1-day-old neonatal rat lungs were observed by light microscope and electronic microscope. The levels of PDGF-A mRNA and protein in fetal and neonatal rat lungs were measured by reverse transcriptase-polymerase chain reaction (RT-PCR) technique and Western blot method respectively.

RESULTS

Under the light microscope, smaller alveolar space, smaller diameter of the respiratory membrane and thicker alveolus mesenchyma were observed in lungs of fetal and neonatal rats from the vitamin D deficiency group compared with the controls (P<0.05). Under the electronic microscope, fewer lamellar bodies but more glycogen deposition in intracytoplasm were observed in the lungs of fetal rats from the vitamin D deficiency group compared with the controls. There was an increased number of empty lamellar bodies in neonatal rats from the vitamin D deficiency group. The levels of PDGF-A mRNA and protein in lungs of fetal and neonatal rats from the vitamin D deficiency group were significantly lower than the controls (P<0.05).

CONCLUSIONS

Maternal vitamin D deficiency during pregnancy may inhibit the development of lung morphogenesis and PDGF-A expression in late fetal and neonatal rats. The low expression of PDGF-A may be involved in the inhibitory effect of vitamin D deficiency on the lung development.