透明质酸对体外伤口收缩模型的影响

目的研究影响伤口收缩的因素,使其能有利于伤口愈合,又不致引起瘢痕的异常增生。方法建立以瘢痕成纤维细胞和I型胶原混悬液形成的胶原网架凝胶块,来模拟体内伤口收缩,观察透明质酸(HA)对其收缩的影响。结果发现HA在一定浓度范围内对伤口收缩有剂量依赖型抑制作用。结论HA有抑制瘢痕形成和异常收缩作用,为临床应用提供指导意义。     

Topical Application of Recombinant Type VII Collagen Incorporates Into the Dermal–Epidermal Junction and Promotes Wound Closure

Patients with recessive dystrophic epidermolysis bullosa (RDEB) have incurable skin fragility, blistering, and skin wounds due to mutations in the gene that codes for type VII collagen (C7) that mediates dermal–epidermal adherence in human skin. In this study, we evaluated if topically applied human recombinant C7 (rC7) could restore C7 at the dermal–epidermal junction (DEJ) and enhance wound healing. We found that rC7 applied topically onto murine skin wounds stably incorporated into the newly formed DEJ of healed wounds and accelerated wound closure by increasing re-epithelialization. Topical rC7 decreased the expression of fibrogenic transforming growth factor-β2 (TGF-β2) and increased the expression of anti-fibrogenic TGF-β3. These were accompanied by the reduced expression of connective tissue growth factor, fewer α smooth muscle actin (α-SMA)–positive myofibroblasts, and less deposition of collagen in the healed neodermis, consistent with less scar formation. In addition, using a mouse model in which skin from C7 knock out mice was grafted onto immunodeficient mice, we showed that applying rC7 onto RDEB grafts with wounds restored C7 and anchoring fibrils (AFs) at the DEJ of the grafts and corrected the dermal–epidermal separation. The topical application of rC7 may be useful for treating patients with RDEB and patients who have chronic skin wounds.     

The effect of dextrin-rhEGF on the healing of full-thickness, excisional wounds in the (db/db) diabetic mouse

Chronic wounds, such as ulceration of the lower limb, represent a significant clinical challenge in today's ageing society. With the aim of identifying improved therapeutics, we have previously described a bioresponsive, dextrin-recombinant human epidermal growth factor conjugate (dextrin-rhEGF), that (i) protects rhEGF against proteolytic degradation by human chronic wound fluid; and (ii) mediates rhEGF release by α-amylase, capable of stimulating increased proliferation/migration in normal dermal and chronic wound fibroblasts; and keratinocytes, in vitro. The aim of this study was to extend these findings, by investigating the effects of dextrin-rhEGF on wound healing in the (db/db) diabetic mouse, a widely used in vivo model of delayed wound healing. Standardised, full-thickness excisional wounds, created in the dorsal flank skin, were treated topically with succinoylated dextrin (50 μg/mL), rhEGF (10 μg/mL) or dextrin-rhEGF (1 or 10 μg/mL). Treatments were applied immediately after injury and subsequently on post-wounding, days 3 and 8. Wound healing was assessed macroscopically, in terms of initiation of neo-dermal tissue deposition and wound closure (including wound contraction and re-epithelialisation), over a 16 day period. Wound healing was assessed histologically, in terms of granulation tissue formation/maturity; cranio-caudal wound contraction and wound angiogenesis (CD31 immuno-staining), using tissues harvested at day 16. Blood samples were also analysed for α-amylase and rhEGF concentrations. In this established impaired wound healing model, the topically-applied dextrin-rhEGF significantly accelerated wound closure and neo-dermal tissue formation at the macroscopic level; and significantly increased granulation tissue deposition and angiogenesis at the histological level (p < 0.05), relative to untreated, succinoylated dextrin and rhEGF alone controls. Overall, these findings support the further development of bioresponsive polymer conjugates, for tissue repair.     

糖尿病肾病中肾小管上皮细胞-肌成纤维细胞转分化的意义

目的:通过糖尿病肾病(DN)患者肾脏标本及DN大鼠模型,探讨肾小管上皮细胞-肌成纤维细胞转分化(EMT)在DN中的意义。方法:肾穿刺活检取DN患者肾脏组织,通过Western blot检测E-cadherin及α-SMA蛋白的表达,免疫组化观察E-cadherin及α-SMA的表达,Real time-PCR检测ColⅠRNA基因的表达,并检测空腹血糖、血清肌酐及24h尿蛋白水平。注射链脲佐菌素建立大鼠DN模型,分别于建模后1、2、4、8、12及24周处死动物并取肾脏组织,并用上述同样方法检测相关指标。结果:DN患者中E-cadherin表达下降,α-SMA及ColⅠ mRNA表达上调。DN组大鼠肾组织α-SMA蛋白及ColⅠ mRNA水平明显高于正常对照组(P<0.05),E-cadherin蛋白表达显著低于正常对照组(P<0.05);相关分析结果提示,DN组大鼠α-SMA、ColⅠ的变化趋势与血糖、24h尿道白及肌酐的变化呈正相关;E-cadherin蛋白表达与上述生化指标的变化呈负相关。结论:EMT能够导致DN患者肾功能的下降,在DN进展中起重要作用。     

The effects of cross‐linking a collagen‐elastin dermal template on scaffold bio‐stability and degradation

MatriDerm is a collagen‐elastin dermal template that promotes regeneration in full‐thickness wound repair. Due to its noncross‐linked status, MatriDerm biodegrades quickly in a wound. Facilitating vascularization and dermal repair, it is desirable for MatriDerm to remain present until the wound healing process is complete, optimizing tissue regeneration and reducing wound contraction. The aim of this study was to investigate the effect of cross‐linking MatriDerm on its mechanical and biological properties and to enhance its regenerative functionality. MatriDerm was chemically cross‐linked and characterized in comparison with noncross‐linked MatriDerm. Scaffold properties including surface morphology, protein release and mechanical strength were assessed. Cell‐scaffold interaction, cell proliferation and migration were examined using human dermal fibroblasts. Scaffold biodegradation and its impact on wound healing and contraction were studied in a mouse model. Results showed that cross‐linked MatriDerm displayed a small reduction in pore size, significantly less protein loss and a threefold increase in tensile strength. A significant increase in fibroblast proliferation and migration was observed in cross‐linked MatriDerm with reduced scaffold contraction in vitro. In the mouse model, noncross‐linked MatriDerm was almost completely biodegraded after 14 days whereas cross‐linked MatriDerm remained intact. No significant difference in wound contraction was found between scaffolds. In conclusion, cross‐linked MatriDerm showed a significant increase in stability and strength, enhancing its durability and cell‐scaffold interaction. in vivo analysis showed cross‐linked MatriDerm had a reduced biodegradation rate with a similar host response. The extended structural integrity of cross‐linked MatriDerm could potentially facilitate improved skin tissue regeneration, promoting the formation of a more pliable scar.     

抗纤灵对5/6肾切除诱导慢性肾脏纤维化小鼠模型细胞外基质的影响

目的探讨抗纤灵对5/6肾切除诱导的慢性肾脏纤维化小鼠模型肾组织细胞外基质(ECM)的影响。方法C57BL/6J雄性小鼠,采用5/6肾切除制备慢性肾脏纤维化模型,2周后根据肌酐水平把手术组小鼠分为模型组,雷帕霉素和抗纤灵组,同时设立假手术组,每组10只,雷帕霉素,抗纤灵组分别给予雷帕霉素(0.00 016 g/kg),抗纤灵(0.4 g/(kg·d)灌胃治疗,模型组和假手术组等量蒸馏水,连续用药4周后处死小鼠,免疫荧光法测定肾组织I型胶原(ColⅠ),Ⅲ型胶原(Col Ⅲ),α-平滑肌肌动蛋白(α-SMA)纤维化面积;RT-PCR法检测肾组中,Co Ⅰ,Col Ⅲ,α-SMAmRNA的表达。结果与假手组比较,模型组Co Ⅰ,Col Ⅲ,α-SMA胶原纤维化面积比值明显增加,Co Ⅰ,Col Ⅲ,α-SMAmRNA显著增高(P < 0.01),与模型组比较,雷帕霉素,抗纤灵组胶原纤维面积比值,mRNA表达水平均有所降低(P < 0.01)。雷帕霉素与抗纤灵组比较,两者之间无显著性差异。结论抗纤灵具有雷帕霉素类似的作用,能够通过抑制小鼠肾组织Co Ⅰ,Col Ⅲ,α-SMAmRNA的表达,减少Co Ⅰ,Col Ⅲ,α-SMA纤维化面积,对肾纤维化有一定的改善作用。      

GSK-3beta in mouse fibroblasts controls wound healing and fibrosis through an endothelin-1-dependent mechanism

Glycogen synthase kinase–3 (GSK-3) is a widely expressed and highly conserved serine/threonine protein kinase encoded by 2 genes, GSK3A and GSK3B. GSK-3 is thought to be involved in tissue repair and fibrogenesis, but its role in these processes is currently unknown. To investigate the function of GSK-3β in fibroblasts, we generated mice harboring a fibroblast-specific deletion of Gsk3b and evaluated their wound-healing and fibrogenic responses. We have shown that Gsk3b-conditional-KO mice (Gsk3b-CKO mice) exhibited accelerated wound closure, increased fibrogenesis, and excessive scarring compared with control mice. In addition, Gsk3b-CKO mice showed elevated collagen production, decreased cell apoptosis, elevated levels of profibrotic α-SMA, and increased myofibroblast formation during wound healing. In cultured Gsk3b-CKO fibroblasts, adhesion, spreading, migration, and contraction were enhanced. Both Gsk3b-CKO mice and fibroblasts showed elevated expression and production of endothelin-1 (ET-1) compared with control mice and cells. Antagonizing ET-1 reversed the phenotype of Gsk3b-CKO fibroblasts and mice. Thus, GSK-3β appears to control the progression of wound healing and fibrosis by modulating ET-1 levels. These results suggest that targeting the GSK-3β pathway or ET-1 may be of benefit in controlling tissue repair and fibrogenic responses in vivo.     

Human corneal fibroblast migration and extracellular matrix synthesis during stromal repair: Role played by platelet‐derived growth factor‐BB,basic fibroblast growth factor,and transforming growth factor‐β1

The development of treatments that modulate corneal wound healing to avoid fibrosis during tissue repair is important for the restoration of corneal transparency after an injury. To date, few studies have studied the influence of growth factors (GFs) on human corneal fibroblast (HCF) expression of extracellular matrix (ECM) proteins such as collagen types I and III, proteoglycans such as perlecan, or proteins implicated in cellular migration such as α5β1‐integrin and syndecan‐4. Using in vitro HCFs, a mechanical wound model was developed to study the influence of the GFs basic fibroblast GF (bFGF), platelet‐derived GF (PDGF‐BB) and transforming GF‐β1 (TGFβ1) on ECM protein production and cellular migration. Our results show that mechanical wounding provokes the autocrine release of bFGF and TGFβ1 at different time points during the wound closure. The HCF response to PDGF‐BB was a rapid closure due to fast cellular migration associated with a high focal adhesion replacement and a high expression of collagen and proteoglycans, producing nonfibrotic healing. bFGF stimulated nonfibrotic ECM production and limited the migration process. Finally, TGFβ1 induced expression of the fibrotic markers collagen type III and α5β1 integrin, and it inhibited cellular migration due to the formation of focal adhesions with a low turnover rate. The novel in vitro HCF mechanical wound model can be used to understand the role played by GFs in human corneal repair. The model can also be used to test the effects of different treatments aimed at improving the healing process. Copyright © 2016 John Wiley & Sons, Ltd.     

碱性成纤维细胞生长因子对增生性瘢痕与正常皮肤成纤维细胞胶原、纤维连接蛋白表达的影响

背景:碱性成纤维细胞生长因子能促进愈合伤口产生胶原蛋白、纤维连接蛋白和基质酶的基质成分.然而,细胞增殖、细胞外基质及新生血管的形成或伤口基质重塑过程失调,会导致瘢痕组织过度增殖.目的:观察碱性成纤维细胞生长因子在正常皮肤创面愈合和增生性瘢痕形成中的作用.方法:从5例进行瘢痕修复手术患者身上同时取正常皮肤和增生性瘢痕组织,分离培养正常人皮肤成纤维细胞和增生性瘢痕成纤维细胞.应用RT-PCR和酶联免疫吸附法检测两种成纤维细胞胶原、纤维连接蛋白基因表达和蛋白合成.采用JC-1染色和流式细胞术测定成纤维细胞线粒体膜电位改变,采用化学发光法检测细胞内ATP水平改变.观察碱性成纤维细胞生长因子对两种细胞的上述指标的影响.结果与结论:不同浓度碱性成纤维细胞生长因子可减慢增生性瘢痕成纤维细胞生长,抑制增生性瘢痕成纤维细胞Ⅰ型胶原表达和合成(P<0.05).碱性成纤维细胞生长因子对正常皮肤和增生性瘢痕成纤维细胞Ⅲ型胶原表达和合成均无影响.然而可上调正常皮肤成纤维细胞表达纤维连接蛋白(P<0.05).此外,10,100 μg/L碱性成纤维细胞生长因子处理后增生性瘢痕成纤维细胞线粒体膜电位呈去极化趋势,正常皮肤成纤维细胞中ATP水平显著增高(P<0.05).结果表明,碱性成纤维细胞生长因子在正常皮肤创面愈合和增生性瘢痕形成中可能有不同的作用和机制.     

Promising effects of exosomes isolated from menstrual blood‐derived mesenchymal stem cell on wound‐healing process in diabetic mouse model

Wound healing is a complicated process that contains a number of overlapping and consecutive phases, disruption in each of which can cause chronic nonhealing wounds. In the current study, we investigated the effects of exosomes as paracrine factors released from menstrual blood‐derived mesenchymal stem cells (MenSCs) on wound‐healing process in diabetic mice. The exosomes were isolated from MenSCs conditioned media using ultracentrifugation and were characterized by scanning electron microscope and western blotting assay. A full thickness excisional wound was created on the dorsal skin of each streptozotocin‐induced diabetic mouse. The mice were divided into three groups as follows: phosphate buffered saline, exosomes, and MenSC groups. We found that MenSC‐derived exosomes can resolve inflammation via induced M1–M2 macrophage polarization. It was observed that exosomes enhance neoangiogenesis through vascular endothelial growth factor A upregulation. Re‐epithelialization accelerated in the exosome‐treated mice, most likely through NF‐κB p65 subunit upregulation and activation of the NF‐κB signaling pathway. The results demonstrated that exosomes possibly cause less scar formation through decreased Col1:Col3 ratio. These notable results showed that the MenSC‐derived exosomes effectively ameliorated cutaneous nonhealing wounds. We suggest that exosomes can be employed in regenerative medicine for skin repair in difficult‐to‐heal conditions such as diabetic foot ulcer.     

α3(V) collagen is critical for glucose homeostasis in mice due to effects in pancreatic islets and peripheral tissues

Collagen V, broadly expressed as α1(V)2 α2(V) heterotrimers that regulate collagen fibril geometry and strength, also occurs in some tissues, such as white adipose tissue (WAT), pancreatic islets, and skeletal muscle, as the poorly characterized α1(V) α2(V) α3(V) heterotrimer. Here, we investigate the role of α3(V) collagen chains by generating mice with a null allele of the α3(V) gene Col5a3 (Col5a3–/– mice). Female Col5a3–/– mice had reduced dermal fat and were resistant to high-fat diet–induced weight gain. Male and female mutant mice were glucose intolerant, insulin-resistant, and hyperglycemic, and these metabolic defects worsened with age. Col5a3–/– mice demonstrated decreased numbers of pancreatic islets, which were more susceptible to streptozotocin-induced apoptosis, and islets isolated from mutant mice displayed blunted glucose-stimulated insulin secretion. Moreover, Col5a3–/– WAT and skeletal muscle were defective in glucose uptake and mobilization of intracellular GLUT4 glucose transporter to the plasma membrane in response to insulin. Our results underscore the emerging view of the importance of ECM to the microenvironments that inform proper development/functioning of specialized cells, such as adipocytes, β cells, and skeletal muscle.     

Effect of povidone-iodine on wound healing in control, diabetic and steroid depressed rats

A comparative study was made on the effect of povidone-iodine on wound healing in normal, diabetic and steroid depressed states in the excision wound model in rats. Healing was assessed by the rate of contraction of wounds and epithelialization after three weeks of topical application. Normal and diabetic groups were comparable (P < 0.02) concerning the above-mentioned parameters as well as collagen formation. The steroid group showed significant retardation in healing time (P < 0.001), epithelialization (P < 0.001) and collagen formation (P < 0.001) showing that povidone-iodine did not overcome the steroid effect.     

内皮抑素通过Notch1/血小板源生长因子信号通路对小鼠肺纤维化的抑制作用

目的研究内皮抑素通过Notch1/血小板源生长因子（Notch1/PDGF）信号通路对小鼠肺纤维化的抑制作用。 方法将30只小鼠分为对照组、模型组和内皮抑素组,每组各10只。模型组和内皮抑素组小鼠给予气管内注射博莱霉素（5 mg/kg）以建立肺纤维化模型,对照组小鼠仅注射等量等渗NaCl溶液;内皮抑素组小鼠每天腹腔注射内皮抑素（2.3 mg/kg）,对照组和模型组小鼠每天腹腔注射等量等渗NaCl溶液,所有小鼠均持续注射21 d。21 d后处死所有小鼠,取左肺组织行病理学染色;取右肺组织,应用Western-blotting法检测Collagen I,Notch1/PDGF信号通路相关蛋白转化生长因子β1（TGF-β1）、Hes1、Hey1、PDGF-B、PDGF受体β（PDGFR-β）及周细胞相关蛋白Desmin、神经元-胶质细胞抗原2（NG2）及α-平滑肌肌动蛋白（α-SMA）的表达水平。 结果光镜下,可见对照组小鼠肺泡结构完整,未见异常胶原蛋白;模型组小鼠肺泡结构被破坏,有大量胶原蛋白沉积;内皮抑素组小鼠可见肺组织结构相对完整,而胶原蛋白明显减少。3组小鼠间Collagen I、TGF-β1、Hes1、Hey1、PDGF-B、PDGFR-β、Desmin、NG2及α-SMA蛋白表达水平的比较,差异均有统计学意义（F = 12.068、30.603、29.757、35.451、16.059、16.420、24.512、19.084、28.102,P均<0.001）。进一步两两比较发现,内皮抑素组小鼠的Collagen I、TGF-β1、Hes1、Hey1、PDGF-B、PDGFR-β及α-SMA蛋白表达水平均显著低于模型组小鼠,Desmin及NG2蛋白表达水平均显著高于模型组小鼠（P均<0.05）;而内皮抑素组与对照组小鼠间Collagen I、TGF-β1、Hes1、Hey1、PDGF-B、PDGFR-β、Desmin、NG2及α-SMA蛋白表达水平的比较,差异均无统计学意义（P均>0.05）。 结论内皮抑素可通过Notch1/PDGF信号通路抑制周细胞向肌成纤维细胞转化,从而影响小鼠肺纤维化。     

烧伤后不同类型胶原在增生性瘢痕发生中的作用研究

目的观察烧伤后瘢痕形成过程中Ⅰ、Ⅲ型胶原纤维的动态变化。方法采用苦味酸天狼猩红染色法对不同时期的瘢痕组织进行Ⅰ、Ⅲ型胶原纤维观察,RIA法对瘢痕组织中的Ⅰ、Ⅲ型前胶原进行检测。结果偏光观察正常皮肤中以Ⅲ型纤维为主,1月以后Ⅰ型纤维明显增加,1年以后的瘢痕组织中几乎全部为粗大的Ⅰ型纤维,极少Ⅲ型纤维。放免结果Ⅰ/Ⅲ型前胶原比例逐渐增加,但Ⅲ型前胶原的含量仍然较高;晚期瘢痕中Ⅰ、Ⅲ型前胶原含量均显著降低。结论烧伤后瘢痕形成过程中Ⅰ型胶原逐渐增加,Ⅲ型胶原逐渐减少,后期几乎完全为Ⅰ型胶原纤维取代,前胶原的变化与胶原纤维的变化不吻合,不能反映胶原纤维的实际情况。     

Effects of relaxin on cardiac fibrosis,apoptosis, and tachyarrhythmia in rats with myocardial infarction

Relaxin is safe and efficient to use for treating acute heart failure. However, the electrophysiological and arrhythmogenic effects of relaxin in an experimental healing infarction model remain unknown. In this study, a rat model with myocardial infarction (MI) received relaxin (0.5 mg/kg per day) or vehicle (sodium acetate) infusion via implantable mini-pumps for 2 weeks. Thereafter, hemodynamic measurement, electrophysiological study, histological examination, and immunofluorescence labeling were performed. Relaxin treatment significantly attenuated tachyarrhythmia inducibility and cardiac dysfunction in healing infarcted heart. Epicardial monophasic action potentials showed that relaxin significantly reduced the dispersion of action potential duration in postinfarcted hearts. Histological study revealed that relaxin significantly reduced myocardial apoptosis and cardiac fibrotic collagen deposition. Western blot revealed that relaxin treatment significantly suppressed the protein expression levels of TGFβ1, α-SMA, and type I collagen. Furthermore, abnormal alterations of Connexin 43, including reduction and lateralization, were significantly attenuated by relaxin treatment at the infarcted border zone. This study provides strong evidence that continuous relaxin intervention ameliorates cardiac fibrosis and apoptosis, attenuates remodeling of gap junction and focal heterogeneity of repolarization, and reduces vulnerability to tachyarrhythmias.     

Osteocyte and osteoblast apoptosis and excessive bone deposition accompany failure of collagenase cleavage of collagen

Mice carrying a targeted mutation (r) in Col1a1, encoding a collagenase-resistant form of type I collagen, have altered skeletal remodeling. In hematoxylin and eosin-stained paraffin sections, we detect empty lacunae in osteocytes in calvariae from Col1a1(r/r) mice at age 2 weeks, increasing through age 10-12 months. Empty lacunae appear to result from osteocyte apoptosis, since staining of osteocytes/periosteal osteoblasts with terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling is increased in Col1a1(r/r) relative to wild-type bones. Osteocyte perilacunar matrices stained with Ab that recognizes collagenase collagen alpha1(I) chain cleavage ends in wild-type but not Col1a1(r/r) calvariae. Increased calvarial periosteal and tibial/femoral endosteal bone deposition was found in Col1a1(r/r) mice from ages 3-12 months. Calcein labeling of calvarial surfaces was increased in Col1a1(r/r) relative to wild-type mice. Daily injections of synthetic parathyroid hormone for 30 days increased calcein-surface labeling in wild-type but caused no further increase in the already high calcein staining of Col1a1(r/r) bones. Thus, failure of collagenase cleavage of type I collagen in Col1a1(r/r) mice is associated with osteocyte/osteoblast death but increases bone deposition in a manner that mimics the parathyroid hormone-induced bone surface activation seen in wild-type mice.     

Wound Healing Activities of Rafflesia Hasseltii Extract in Rats

The effects of topical application of Rafflesia hasseltii buds and flowers extract on the rate of wound healing and histology of healed wound were assessed. Four groups of adult male Sprague Dawley rats were experimentally wounded in the posterior neck area. A thin layer of blank placebo was applied topically to wounds of Group 1 rats. Wounds of experimental animals (Group 2 and 3) were treated with placebo containing 5% and 10% R. hasseltii buds extract, respectively. A thin layer of Intrasite gel was applied topically to wounds of Group 4 animals as reference. Macroscopically, wounds treated with placebo containing 5% and 10% R. hasseltii buds extract or Intrasite gel have been significantly accelerated the rate of wound healing compared to placebo-treated wounds. Histological analysis of healed wounds has confirmed this effect. Wounds treated with placebo containing 5%, 10% R. hasseltii buds extract or Intrasite gel showed markedly less scar width at wound enclosure and granulating tissue contained markedly more collagen and proliferating fibroblasts, but with the absence of inflammatory cells compared to wounds treated with blank placebo. In conclusion, the findings of increased rate of wound closure and contraction together with the histological findingssuggest that Rafflesia hasseltii buds extract is very effective in accelerating the wound healing process.     

LCB 03-0110, a novel pan-discoidin domain receptor/c-Src family tyrosine kinase inhibitor, suppresses scar formation by inhibiting fibroblast and macrophage activation

Wound healing generally induces an inflammatory response associated with tissue fibrosis in which activated macrophage and myofibroblast cells are primarily involved. Although this is known to be the underlying mechanism for scarring and various fibrotic pathologies, no effective intervention is currently available. We identified (3-(2-(3-(morpholinomethyl)phenyl)thieno[3,2-b]pyridin-7-ylamino)phenol (LCB 03-0110), a thienopyridine derivative, as a potent inhibitor of discoidin domain receptor family tyrosine kinases and discovered that this compound strongly inhibits several tyrosine kinases, including the c-Src family, spleen tyrosine kinase, Bruton's tyrosine kinase, and vascular endothelial growth factor receptor 2, which are important for immune cell signaling and inflammatory reactions. LCB 03-0110 suppressed the proliferation and migration of primary dermal fibroblasts induced by transforming growth factor β1 and type I collagen, and this result correlated with the inhibition ability of the compound against enhanced expression of α-smooth muscle actin and activation of Akt1 and focal adhesion kinase. In J774A.1 macrophage cells activated by lipopolysaccharide LCB 03-0110 inhibited cell migration and nitric oxide, inducible nitric-oxide synthase, cyclooxygenase 2, and tumor necrosis factor-α synthesis. LCB 03-0110 applied topically to full excisional wounds on rabbit ears suppressed the accumulation of myofibroblast and macrophage cells in the healing wound and reduced hypertrophic scar formation after wound closing, without delaying the wound closing process. Taken together, the pharmacological activities of LCB 03-0110 suggest that it could be an effective agent for suppressing fibroinflammation by simultaneously targeting activated fibroblasts and macrophages.     

骨形成蛋白-7对晚期糖基化终产物诱导大鼠腹膜间皮细胞上皮-间叶转化的影响

目的探讨骨形成蛋白-7(BMP-7)对晚期糖基化终产物诱导大鼠腹膜间皮细胞上皮-间叶转化(EMT)的影响。方法晚期糖基化终产物诱导发生上皮-间叶转化的体外培养大鼠腹膜间皮细胞,分别经含5 ng/mL及80 mmol/L晚期糖基化终产物的M199培养基和含10 ng/mL BMP-7及80 mmol/L晚期糖基化终产物的M199培养基培养48 h,以含80 mmol/L晚期糖基化终产物的M199培养基为对照,应用实时定量PCR法检测间皮细胞E-cadherin、α平滑肌肌动蛋白(α-SMA)、Ⅰ型胶原(Collagen I)、转化生长因子β1(TGF-β1)、血管内皮生长因子(VEGF)mRNA的表达;应用Western印迹法检测E-cadherin、α-SMA蛋白的表达;应用ELISA法检测间皮细胞TGF-β1、VEGF蛋白表达水平。结果 BMP-7作用后,上皮-间叶转化的大鼠腹膜间皮细胞E-cadherin mRNA和E-cadherin蛋白表达水平显著增加(P<0.05)。BMP-7作用后,上皮-间叶转化的大鼠腹膜间皮α-SMA、Collagen I、TGF-β1、VEGFmRNA和α-SMA、TGF-β、VEGF蛋白表达水平显著下降(P<0.05)。结论 BMP-7能上调上皮-间叶转化的大鼠腹膜间皮细胞E-cadherin表达和下调α-SMA、Collagen I、TGF-β、VEGF表达,BMP-7能逆转晚期糖基化终产物诱导大鼠腹膜间皮细胞EMT。     

Transgenic expression of human matrix metalloproteinase-1 attenuates pulmonary arterial hypertension in mice

PAH (pulmonary arterial hypertension) is a debilitating and life-threatening disease, often affecting young people. We specifically expressed human MMP-1 (matrix metalloproteinase-1) in mouse macrophages and examined its effects in attenuating the decompensating features of MCT (monocrotaline)-induced PAH. Measurement of RV (right ventricular) pressure revealed a 2.5-fold increase after treatment with MCT, which was reduced to 1.5-fold in MMP-1 transgenic mice. There was conspicuous pulmonary inflammation with chronic infiltration of mononuclear cells after the administration of MCT, which was significantly diminished in transgenic mice. Furthermore, transgenic mice showed decreased collagen deposition compared with WT (wild-type). Staining for Mac-3 (macrophage-3) and α-SMA (α-smooth muscle actin) revealed extensive infiltration of macrophages and medial hypertrophy of large pulmonary vessels with complete occlusion of small arteries respectively. These changes were markedly reduced in MMP-1 transgenic mice compared with WT. Western blotting for molecules involved in cell multiplication and proliferation depicted a significant decrease in the lung tissue of transgenic mice after the treatment with MCT. In conclusion, the present study demonstrated that transgenic expression of human MMP-1 decreased proliferation of smooth muscle cells and prevented excessive deposition of collagen in the pulmonary arterial tree. Our results indicate that up-regulation of MMP-1 could attenuate the debilitation of human PAH and provide an option for therapeutic intervention.