Neovascularization In Rats Research Articles

The Miao medicine Sidaxue alleviates rheumatoid arthritis in rats possibly by downregulating matrix metalloproteinases

To explore the inhibitory effect of Sidaxue, a traditional Miao herbal medicine formula, on articular bone and cartilage destruction and synovial neovascularization in rats with collagen-induced arthritis (CIA). In a SD rat model of CIA, we tested the effects of daily gavage of Sidaxue at low, moderate and high doses (10, 20, and 40 g/kg, respectively) for 21 days, with Tripterygium glycosides (GTW) as the positive control, on swelling in the hind limb plantar regions by arthritis index scoring. Pathologies in joint synovial membrane of the rats were observed with HE staining, and serum TNF-α and IL-1β levels were detected with ELISA. The expressions of NF-κB p65, matrix metalloproteinase 1 (MMP1), MMP2 and MMP9 at the mRNA and protein levels in the synovial tissues were detected using real-time PCR and Western blotting. Network pharmacology analysis was conducted to identify the important target proteins in the pathways correlated with the therapeutic effects of topical Sidaxue treatment for RA, and the core target proteins were screened by topological analysis. Treatment with GTW and Sidaxue at the 3 doses all significantly alleviated plantar swelling, lowered arthritis index scores, improved cartilage and bone damage and reduced neovascularization in CIA rats (P<0.05), and the effects of Sidaxue showed a dose dependence. Both GTW and Sidaxue treatments significantly lowered TNF-α, IL-1β, NF-κB p65, MMP1, MMP2, and MMP9 mRNA and protein expressions in the synovial tissues of CIA rats (P<0.05). Network pharmacological analysis identified MMPs as the core proteins associated with topical Sidaxue treatment of RA. Sidaxue alleviates articular bone and cartilage damages and reduces synovial neovascularization in CIA rats possibly by downregulating MMPs via the TNF-α/IL-1β/NF-κB-MMP1, 2, 9 signaling pathway, and MMPs probably plays a key role in mediating the effect of Sidaxue though the therapeutic pathways other than oral administration.

Protective effect of bone marrow mesenchymal stem cells on the survival zone of the perforator flaps in rats.

Perforator flaps have recently been used in the field of plastic surgery. Skin defects can be reconstructed to reach functional and cosmetic goals. With the development of reconstructive approaches, utilizing stem cells is a hopeful approach to enhance wound healing and tissue recovery. In this study, we assessed the effect of bone marrow mesenchymal stem cells on the perforator flap's survival in rats. Perforator flaps (2.5 × 11 cm) were transplanted into rats and focused on the thoracodorsal, intercostal, and deep circumflex arteries, which were randomly divided into three groups: control, saline, and bone marrow mesenchymal stem cells (7 × 106 ml). Seven days after the surgery, tissue edema, inflammation, and discharge were observed and photographed. Histological analyses were performed to determine flap survival. Hematoxylin and eosin staining was performed to assess levels of microvascular density determined in skin flaps. Rats in the bone marrow mesenchymal stem cells group exhibited higher average flap survival area, and higher microvascular density levels at the dynamical regions of the flaps compared with the other two groups. Subdermal injection of bone marrow mesenchymal stem cells significantly increased ischemic perforator flap survival due to stimulated neovascularization in rats. Our findings suggest the potential usefulness of bone marrow mesenchymal stem cells in preventing skin flap perforator tissue necrosis.

5α-Hydroxycostic acid inhibits choroidal neovascularization in rats through a dual signalling pathway mediated by VEGF and angiopoietin 2

Background5α-Hydroxycostic acid is a eudemane sesquiterpene that is isolated from the natural plant, Laggera alata. It exerts anti-inflammatory and anti-angiogenic effects on human breast cancer cells, but its role and underlying mechanism in choroidal neovascularization (CNV) are still unclear. We conducted a study to verify that 5α-Hydroxycostic acid can inhibit the formation and leakage of CNV, and describe the possible dual pathway by which it exerts its inhibitory effects in this process.MethodsAn in vitro model of choroidal neovascularization was established using VEGF164, while a rat model of choroidal neovascularization was established using a 532 nm laser. In both models, the effects of 5α-Hydroxycostic acid in vivo and in vitro were evaluated to determine its inhibitory effect on abnormal cell proliferation, migration and tubule formation, as well as its effect on pathological changes in choroidal tissues and the area of neovascularization leakage in rats. The levels of components in the VEGF/VEGFR and Ang2/Tie2 signaling pathways were measured in tissues and cells.ResultsIn vitro experiments have shown that 5α-Hydroxycostic acid can inhibit abnormal cell proliferation, migration and angiogenesis. Additionally, 5α-Hydroxycostic acid enhances cell adhesion by inhibiting the phosphorylation pathways of VEGFR2 and Tie2. In vivo experiments demonstrated that 5α-Hydroxycostic acid has a positive therapeutic effect on choroidal neovascularization in rats. It can effectively reduce vascular leakage, consistent with the results of the cell experiments.Conclusion5α-Hydroxycostic acid can inhibit choroidal neovascularization by interfering with the VEGF- and Ang2/Tie2-related pathways, and it may be a good candidate drug for treating CNV.

The role of erythropoietin in the regulation of cardiac tolerance to impact of ischemia and reperfusion

Acute myocardial infarction (AMI) is one of the main causes of mortality among the able-bodied population in Russia and the population of economically developed countries. In recent years, deaths from AMI in the USA and Europe have not decreased. This is due to the lack of highly effective drugs for the treatment of AMI. One of the promising drugs to improve the survival of patients with AMI is erythropoietin. We searched for full-text publications in the PubMed database and on the website of the journal Nature. In studies performed on animals, it was shown that erythropoietin (5000 U/kg) is able to increase cardiac tolerance to ischemia and reperfusion due to activation of a number of kinases (PKC, ERK1/2, Akt, JAK2, PI3K) and due to GSK-3β kinase inactivation. Erythropoietin prevents post-infarction remodeling of the heart and enhances the process of myocardial neovascularization in rats and dogs. Erythropoietin in used doses (on the average 1000 U/kg) does not affect infarct size in patients with AMI and does not have an effect on post-infarction ventricular remodeling in humans. The reason for this discrepancy between experimental and clinical data remains unclear. It is possible that the use of large doses of erythropoietin or the use of its analogues that do not affect erythropoiesis can prevent the development of post-infarction cardiac remodeling in humans.

Metabolic Characterization of Ocular Tissues in Relation to Laser-Induced Choroidal Neovascularization in Rats.

Age-related macular degeneration is a metabolic compromise disorder whose main pathological feature is choroidal neovascularization (CNV) formation. Using untargeted metabolomics analysis, we determined to assess the metabolomic alterations in a CNV rat model to provide an insight into its pathogenesis. In the CNV model, there were 24 significantly changed metabolites in the plasma and 71 in various ocular tissues. Pathway analysis showed that certain metabolic pathways changed in interrelated tissues: for instance, in terms of the altered urea cycle, arginine and proline metabolism were increased in the plasma, while spermidine and spermine biosynthesis activities were increased in the retinal pigment epithelium (RPE)/choroid. The retina and RPE/choroid shared the same changed metabolites of branched-chain amino acid metabolism. Fatty acid metabolism was found to be the significant altered metabolic pathway in the retina of this CNV model. Although the metabolism pattern of different substances is specific for each ocular tissue, there is also a certain material exchange between different tissues. Dysregulated metabolomic profiles in differential tissues may point to an interconnected pathway, oxidative stress response, which may lead to RPE cell degeneration and, ultimately, CNV development.

Mechanoresponsive Drug Loading System with Tunable Host-Guest Interactions for Ocular Disease Treatment.

Conventional administration of eye drops often requires high dosages and/or repetitive treatments to achieve therapeutic efficacy. This is inefficient and may result in side effects or even toxicity. Although many delivery systems of ophthalmic drugs have been reported, most of them work in a fixed format in which both the type and dose of the loaded drugs cannot be changed upon demand. To overcome this limitation, a hybrid double network hydrogel system composed of methacryloyl gelatin, pluronic F127 diacrylate, and β-cyclodextrin-modified oxidized dextran was developed. The hydrogels presented good mechanical strength and biocompatibility. In vitro assessments demonstrated that the hydrogels loaded with commonly used ophthalmic drugs could sustain the drug release for more than 21 days. This hydrogel system exhibited features of mechanoresponsive drug loading, and the capacity of drug loading could be significantly enhanced by macroscopically mechanical compression. Further in vivo evaluation of the drug delivery capacity showed that a dexamethasone-loaded hydrogel as a fornix insert effectively suppressed upregulation of proangiogenic factors and suture-induced corneal neovascularization in rats. This novel hydrogel system represents a promising drug delivery platform, which could potentially improve the treatments of ocular surface and other diseases.

Stromal Vascular Fraction Promotes Viability of Co-grafted Axial Skin Flaps in Rats Model.

Stromal vascular fraction (SVF) has been proved in promoting the vascularization of fascial flap through cell differentiation and paracrine effect and can be autologous transplanted without culture after isolation in vitro. We intend to establish a novel co-grafted flap model of rats to investigate the efficacy and mechanism of SVF on flaps and skinsin facilitating angiogenesis and immune regulation. 60 female Sprague Dawley rats were divided into the SVF group and the control group. A pedicled fascial flap combined with a free skin model was established, and 4×106 CM-DIl labeled SVF cells were transplanted into the fascia flap; the rats were executed on days 1, 2, 3, 7, 10 postoperatively (n=6). Flow cytometry was carried out to determine the cell proportion and surface marker of SVFs. The therapeutic effects of SVF were evaluated via Doppler blood perfusion imager, flap survival rates, histology, immunohistochemistry and immunofluorescence. The bioinformatic mechanism analysis was achieved by high-throughput RNAseq of mRNA and LncRNA. Flow cytometry confirmed SVF contains heterogeneous cellular composition, especially hematopoietic cells. Doppler blood perfusion imager showed SVF significantly improved flap survival with higher blood perfusion and survival rates. Immunohistochemistry of CD31 displayed higher level of angiogenesis in SVF-treated group, and CM-DIL-labeled SVF cells could survive and participate in revascularization, and RNA sequencing results revealed SVF promoted wound healing by facilitating intercellular adhesion, cell migration and positive immune response. SVF could reduce skin flap necrosis and activated neovascularization in rats by facilitating intercellular adhesion, cell migration and regulate positive immune response. This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266.

Vorolanib, a novel tyrosine receptor kinase receptor inhibitor with potent preclinical anti-angiogenic and anti-tumor activity

Vorolanib (CM082) is a multi-targeted tyrosine kinase receptor inhibitor with a short half-life and limited tissue accumulation that has been shown to reduce choroidal neovascularization in rats. In this preclinical study, vorolanib demonstrated competitive binding and inhibitory activities with KDR, PDGFRβ, FLT3, and C-Kit, and inhibited RET and AMPKα1 more weakly than sunitinib, indicating more stringent kinase selectivity. Vorolanib inhibited vascular endothelial growth factor (VEGF)-induced proliferation of human umbilical vein endothelial cells (HUVECs) and HUVEC tube formation in vitro. In mouse xenograft models, vorolanib inhibited tumor growth of MV-4-11, A549, 786-O, HT-29, BxPC-3, and A375 cells in a dose-dependent fashion. Complete tumor regression was achieved in the MV-4-11 xenograft model. No significant toxicities were observed in vorolanib groups, whereas a significant negative impact on body weights was observed in the sunitinib group at a dose of 40 mg/kg qd. Overall, vorolanib is a novel multi-kinase receptor inhibitor with potent preclinical anti-angiogenic and anti-tumor activity that is potentially less toxic than other similar kinase inhibitors.

In vivo evaluation of laser-induced choroidal neovascularization in rats simultaneously using optical coherence tomography and photoacoustic microscopy

Determination of the precise location and the degree of the Choroidal neovascularization (CNV) lesion is essential for diagnosation Neovascular age-related macular degeneration (AMD) and evaluation the efficacy of treatment. Noninvasive imaging techniques with specific contrast for CNV evaluation are demanded. In this paper, two noninvasive imaging techniques, namely Optical coherence tomography (OCT) and Photoacoustic microscopy (PAM), are combined to provide specific detection of CNV for their complimentary contrast mechanisms. In vivo time-serial evaluation of Laser-induced CNV in rats is present at days 1, 3, 5, 7, 14, 21 after laser photocoagulation is applied to the rat fundus. Both OCT and PAM show that the CNV increases to its maximum at day 7 and decreases at day 14. Quantification of CNV area and CNV thickness is given. The dual-modal information of CNV is consistent with the histologic evaluation by hematoxylin and eosin (H&amp;E) staining.

Evaluation of the effect of Carvacrol on retinal neovascularization in rats

Objectives: To compare the effect of intraperitoneal administered Carvacrol with bevacizumab in an oxygen-induced retinopathy (OIR) model in rats.Materials and Methods: Twenty-eight newborn rats were included in the study and the OIR model was created with the 50/10% oxygen style. The study consisted of 4 groups and each rat in the groups received an intraperitoneal injection of 0.01 mL once on the postnatal 14th day. While the OIR model was not created in Group I (control group), it was created for Groups II, III, and IV. Groups I and II were injected with 0.9% NaCl solution, Group III with bevacizumab, and Group IV with carvacrol. The rats were sacrificed on the postnatal 18th day. Results: Histopathological and immunohistochemical studies showed that the number of retinal vascular endothelial cells (RVECs) and nuclear factor (NF)-κB levels decreased similarly in Group III and Group IV compared to Group II. RVECs values for Group I, Group II, Group III and Group IV were measured as 1.26±0.80, 27.10±3.63, 7.54±1.38, and 6.22±1.22, respectively and it differed significantly between groups (p&amp;lt;0.001). Likewise, NF-κB levels were recorded as 0.61 ± 0.30, 4.36±0.65, 2.68±0.44, and 2.85±0.58, respectively and it differed significantly between groups (p&amp;lt;0.001). On the other hand, RVECs and NF-κB levels were similar between Group III, and Group IV (p values were 0.58 and 0.91, respectively).Conclusions: The study demonstrated that carvacrol significantly reduced retinal pathological neovascularizations, RVECs, and NF-κB levels. Moreover, the observed effects were comparable to those of bevacizumab.

Laser-Induced Choroidal Neovascularization in Rats.

The laser-induced choroidal neovascularization (CNV) model has been widely used for research on wet age-related macular degeneration (wet-AMD) and other ocular neovascular diseases. In this model, the Bruch membrane is perforated by laser injury, resulting in neovascularization formed from the choroidal capillaries. It has become a standard method to evaluate the effect of different treatments on CNV progression in preclinical studies. This protocol can be used in various species, including rat, mouse, pig, and monkey. The rodent laser-induced CNV model is the most commonly used because of the advantages in both cost- and time-efficiency. It takes only 10-15min to complete the whole laser procedure after adequate training and practicing the technique. Peak CNV formation occurs at approximately 2weeks after laser application. The entire protocol may require up to 3weeks to complete the treatment, fundus image acquisition, and tissue collection for histologic analysis. This chapter describes the detailed procedures, protocols, and useful notes on how to induce CNV by laser.

Sunitinib malate-loaded biodegradable microspheres for the prevention of corneal neovascularization in rats

Corneal neovascularization (NV) predisposes patients to compromised corneal transparency and visional acuity. Sunitinib malate (Sunb-malate) targeting against multiple receptor tyrosine kinases, exerts potent antiangiogenesis. However, the rapid clearance of Sunb-malate eye drops administered through topical instillation limits its therapeutic efficacy and poses a challenge for potential patient compliance. Sunb-malate, the water-soluble form of sunitinib, was shown to have higher intraocular penetration through transscleral diffusion following subconjunctival (SCT) injection in comparison to its sunitinib free base formulation. However, it is difficult to load highly water-soluble drugs and achieve sustained drug release. We developed Sunb-malate loaded poly(D,L-lactic-co-glycolic acid) (PLGA) microspheres (Sunb-malate MS) with a particle size of approximately 15 μm and a drug loading of 7 wt%. Sunb-malate MS sustained the drug release for 30 days under the in vitro infinite sink condition. Subconjunctival (SCT) injection of Sunb-malate MS provided a prolonged ocular drug retention and did not cause ocular toxicity at a dose of 150 μg of active agent. Sunb-malate MS following SCT injection more effectively suppressed the suture-induced corneal NV than either Sunb-malate free drug or the placebo MS. Local sustained release of Sunb-malate through the SCT injection of Sunb-malate MS mitigated the proliferation of vascular endothelial cells and the recruitment of mural cells into the cornea. Moreover, the gene upregulation of proangiogenic factors induced by the pathological process was greatly neutralized by SCT injection of Sunb-malate MS. Our findings provide a sustained release platform for local delivery of tyrosine kinase inhibitors to treat corneal NV.

Therapeutic Effects against Tissue Necrosis of Remote Ischemic Preconditioning Combined with Human Adipose-Derived Stem Cells in Random-Pattern Skin Flap Rat Models

Introduction Remote ischemic preconditioning (rIPC) is a preventive strategy against ischemia–reperfusion injury. To reduce ischemia–reperfusion injury of random-pattern skin flaps, we investigated the therapeutic effects of rIPC combined with human adipose-derived stem cells (hADSCs) in a rat model. Material and Methods In total, 24 female Sprague Dawley rats were divided into four groups (n = 6 each): control (skin flap only), rIPC, hADSCs, and rIPC + hADSCs. rIPC was performed in the hind limb of the rats over three cycles of 5 min of occlusion and 5 min of reperfusion, using a tourniquet. A rectangular (3 × 9 cm) dorsal skin flap was used. hADSCs (5 × 105 cells/100 µL) labeled with fluorescent dye were transplanted into the normal subcutaneous tissue at the skin flap boundary. After 14 days, the therapeutic effects of rIPC and hADSCs were evaluated via analysis of the necrotic flap area, histopathologic assessment, and immunohistochemistry (von Willebrand Factor (vWF) and CD31). Results The necrotic area of the skin flap significantly decreased in the rIPC + hADSCs group (32.75 ± 1.43%) compared with the control (40.60 ± 3.27%, P < 0.01) and rIPC groups (38.84 ± 0.77%, P < 0.05). Dye-labeled hADSCs migrated to the skin flap from the injection site. In the rIPC + hADSCs group, the epithelial tissue and skin appendage had regenerated, and the smooth muscle and subcutaneous fat layers were preserved. Many more vWF- and CD31-positive vessels were observed in the rIPC + hADSCs group compared with the other groups. Conclusions The rIPC + hADSCs treatment appeared to reduce skin flap necrosis and activated neovascularization in rats. Therefore, it may be a good strategy for clinical treatment of ischemia–reperfusion injury.

Deferoxamine Ameliorates Compressed Spinal Cord Injury by Promoting Neovascularization in Rats.

The therapeutic effect of deferoxamine (DFO) for spinal cord injury (SCI) has been demonstrated in previous studies; however, the exact mechanism of action is still unclear. Here, we hypothesized that DFO ameliorates spinal cord compression by promoting neovascularization. Using an SCI model of moderate compression, rats were intraperitoneally injected with 30mg/kg or 100mg/kg DFO for 1-2weeks, and significant neovascularization was found in the injured spinal cord, showing overexpression of hypoxia inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF), and an increase in the number of new blood vessels. In addition, SCI in rats was significantly ameliorated after treatment with DFO, with less motor dysfunction, increased spared neural tissue, and improved electrophysiological conduction. By contrast, the ameliorative effect of DFO on SCI was suppressed when DFO-induced neovascularization was blocked by lenvatinib, a vascular endothelial growth factor receptor inhibitor, further suggesting that the primary pharmacological effect of DFO in SCI is the promotion of neovascularization. Therefore, we concluded that DFO effectively alleviated SCI by promoting neovascularization in the injured spinal cord. Considering that DFO is an FDA-approved free radical scavenger and iron chelator, it may represent a promising alternative strategy for SCI therapy in the future.

Complement factor B knockdown by short hairpin RNA inhibits laser-induced choroidal neovascularization in rats.

To evaluate whether recombinant complement factor B (CFB) short hairpin RNA (shRNA) reduces laser-induced choroidal neovascularization (CNV) in rats. Laser-induced rat CNV model was established, and then the animals underwent fundus fluorescence angiography (FFA) and hematoxylin and eosin (HE) staining. On day 3 and 7 after photocoagulation, the expression of CFB and membrane attack complex (MAC) was detected by immunhischemistry. A recombinant CFB-shRNA plasmid was constructed. CFB and scrambled shRNA plasmids were intravenous injected into rats via the tail vein on the day of laser treatment, respectively. On day 7, the incidence of CNV was determined by FFA, and the expression of CFB and vascular endothelial growth factor (VEGF) in retinal pigment epithelium (RPE)/choroidal tissues was detected by immunhischemistry, Western blot and/or semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) in CFB and scrambled shRNA groups. The possible adverse effects of CFB-shRNA injection were assessed by transmission electron microscopy and electroretinography. FFA and HE results indicated that a laser-induced rat CNV model was successfully established on day 7 after photocoagulation. The expression of CFB and MAC was extremely weak in normal retina and choroid, and increased on day 3 after photocoagulation. However, it started to reduce on day 7. CFB shRNA plasmid was successfully constructed and induced CFB knockdown in the retinal and choroidal tissues. FFA showed CFB knockdown significantly inhibited incidence of CNV in rats. Moreover, CFB knockdown significantly inhibited the expression of VEGF in RPE/choroidal tissues. CFB shRNA caused no obvious side effects in eyes. CFB knockdown significantly inhibits the formation and development of CNV in vivo through reducing the expression of VEGF, which is a potential therapy target. The alternative pathway of complement activation plays an important role in CNV formation.

Dimethyloxallyl Glycine-Incorporated Borosilicate Bioactive Glass Scaffolds for Improving Angiogenesis and Osteogenesis in Critical-Sized Calvarial Defects

In the field of bone tissue engineering, there has been an increasing interest in biomedical materials with both high angiogenic ability and osteogenic ability. Among various osteogenesis materials, bioactive borosilicate and borate glass scaffolds possess suitable degradation rate and mechanical strength, thus drawing many scholars' interests and attention. In this study, we fabricated bioactive glass scaffolds composed of borosilicate 2B6Sr using the Template-Method and incorporated Dimethyloxalylglycine (DMOG), a small-molecule angiogenic drug possessing good angiogenic ability, to improve bone regeneration. The in-vitro studies showed that porous borosilicate bioactive glass scaffolds released slowly, a steady amount of DMOG and stimulated the proliferation and osteogenic differentiation of human bone marrow stromal cells hBMSCs. In-vivo studies showed that the borosilicate bioactive glass scaffolds could significantly promote new bone formation and neovascularization in rats' calvarial bone defects. These results indicated that DMOG-incorporated bioactive glass scaffold is a successful compound with excellent angiogenesis-osteogenesis ability, which has favorable clinical prospects.

Transscleral sustained ranibizumab delivery using an episcleral implantable device: Suppression of laser-induced choroidal neovascularization in rats.

Successful treatment of age-related macular diseases requires an effective controlled drug release system with less invasive route of administration in the eye to reduce the burden of frequent intravitreal injections for patients. In this study, we developed an episcleral implantable device for sustained release of ranibizumab, and evaluated its efficacy on suppression of laser-induced choroidal neovascularization (CNV) in rats. We tested both biodegradable and non-biodegradable sheet-type devices consisting of crosslinked gelatin/chitosan (Gel/CS) and photopolymerized poly(ethyleneglycol) dimethacrylate that incorporated collagen microparticles (PEGDM/COL). In vitro release studies of FITC-labeled albumin showed a constant release from PEGDM/COL sheets compared to Gel/CS sheets. The Gel/CS sheets gradually biodegraded in the sclera during the 24-week implantation; however, the PEGDM/COL sheets did not degrade. FITC-albumin was detected in the retina during 18 weeks implantation in the PEGDM/COL sheet-treated group, and was detected in the Gel/CS sheet-treated group during 6 weeks implantation. CNV was suppressed 18 weeks after application of ranibizumab-loaded PEGDM/COL sheets compared to a placebo PEGDM/COL sheet-treated group, and to the intravitreal ranibizumab-injected group. In conclusion, the PEGDM/COL sheet device suppressed CNV via a transscleral administration route for 18 weeks, indicating that prolonged sustained ranibizumab release could reduce the burden of repeated intravitreal injections.

Inhibitory effect of bromfenac sodium hydrate ophthalmic solution on alkali burn induced-corneal neovascularization in rats

Objective To investigate the inhibitory effect of bromfenac sodium hydrate ophthalmic solution on corneal neovascularization (CNV) induced by alkali burn. Methods A total of 192 specific pathogen free (SPF) degree adult male Sprague-Dawley (SD) rats were used in this study.One hundred and seventy-two rats were chosen to establish CNV model with alkali burn in the right eyes.Following alkali burn, rats were randomly divided into CNV group, model control group, bromfenac sodium group and fluorometholone group, with 43 rats (43 eyes) in each group.Another 20 rats (40 eyes) served as normal control group.One day after modeling, the model control group, bromfenac sodium group and fluorometholone group received phosphate buffer saline (PBS), bromfenac sodium hydrate ophthalmic solution and 0.1% fluorometholone eye drops, respectively.The state of cornea and anterior chamber and the growth of CNV of rats in each group were observed by slit-lamp microscope every day after modeling.At 1, 3, 7, 14, 21 and 28 days after modeling, the anterior segment photos of the experimental eyes were captured, and the percent of cornea areas covered by CNV was calculated.At 7, 14 and 28 days after modeling, the eye tissue sections were stained with hematoxylin and eosin staining and immunohistochemistry staining to evaluate the expressions of CD45 and VEGF-A.Real-time quantitative PCR and enzyme-linked immunosorbent assay (ELISA) were used to detect the expression of COX-2 and VEGF mRNA and protein level.The use and care of the animals complied with the Statement of the Association for Research in Vision and Ophthalmology(ARVO). Results Each model group showed corneal edema and opacification 1 day after modeling.The corneal edema was aggravated 7 days after modeling.On the 14th day after modeling, the degree of corneal opacity and edema decreased gradually.On the 28th day after modeling, leucoma was observed in CNV group and model control group, and nebula was observed in bromfenac sodium group and fluorometholone group.At 7, 14, 21 and 28 days after modeling, the percentages of CNV areas in bromfenac sodium group and fluorometholone group were significantly lower than those in CNV group and model control group (all at P 0.05). On the 7th day after modeling, the thinning of corneal epithelial layer, edema and arrangement disorder of stroma layer were observed, and the expression of VEGF-A was positive in all model groups; a small amount of CD45 positive inflammatory cell infiltrations were observed in CNV group and model control group.On the 14th and 28th day after modeling, CNV was seen in the center of cornea in CNV group and model control group; the epithelial keratosis and reduction of corneal edema were seen in each group, and no inflammatory cell infiltration was observed in each group.On the 7th day after modeling, the expressions of COX-2 and VEGF mRNA in CNV group and model control group were significantly higher than those in normal control group, bromfenac sodium group and fluorometholone group (all at P<0.05), the expressions of COX-2 and VEGF protein in bromfenac sodium group were significantly lower than those in CNV group (all at P<0.05). The corneal perforation rate in model control group and bromfenac sodium group was 10% (1 case in 10 rats). The corneal perforation rate in fluorometholone group was 30% (3 cases in 10 rats). In each model group, 10% to 30% rats had hyphema. Conclusions Bromfenac sodium hydrate ophthalmic solution can inhibit the formation and growth of CNV after alkali burn in rats.This effect may be mediated by regulating COX-2 expression, reducing inflammation and inhibiting VEGF production. Key words: Corneal neovascularization; Alkali burn; Bromfenac sodium; Cyclooxygenase-2

Роль натрийуретических пептидов и эритропоэтина в регуляции толерантности сердца к действию ишемии и реперфузии. Анализ экспериментальных и клинических данных

Experimental studies show that natriuretic peptides have an infarct-limiting effect both in vitro and in vivo. In studies performed on animals, it was shown that erythropoietin is able to prevent ischemic and reperfusion damage of the heart both in vitro and in vivo by activating a number of kinases and by inactivating GSK-3β kinase. These data suggest that the cardioprotective effect of erythropoietin and natriuretic peptides appears to be associated with the activation of cardiac receptors. Erythropoietin had an anti-apoptotic effect both in vitro and in vivo. In the infarct-limiting effect of the atrial natriuretic peptide, NO synthase, protein kinase C, and the mitoKATP channel are involved. It was found that the in vivo infarct-reducing effect of erythropoietin is associated with the activation of PI3 kinase and ERK1/2 kinase. Brain natriuretic peptide prevents post-infarction cardiac remodeling and increases the survival rate of rats with experimental myocardial infarction. Erythropoietin inhibits post-infarction cardiac remodeling and enhances the process of myocardial neovascularization in rats and dogs. Natriuretic peptides have an infarct-reducing effect in patients with acute myocardial infarction (AMI). Atrial natriuretic peptide prevents post-infarction cardiac remodeling in patients with AMI. Erythropoietin does not affect infarct size in patients with AMI. A single injection of erythropoietin does not affect the post-infarction remodeling of the heart.

MicroRNA-9 inhibits retinal neovascularization in rats with diabetic retinopathy by targeting vascular endothelial growth factor A.

Diabetic retinopathy (DR) is a leading cause of adult visual impairment and loss. This study aims to explore the effects of microRNA-9 (miR-9) on retinal neovascularization during DR by targeting the vascular endothelial growth factor A (VEGFA). DR rat models were successfully established. Retinal microvascular endothelial cells (RMECs) of DR rats were isolated and treated with miR-9 mimic, miR-9 inhibitor or small interfering RNA (siRNA)-VEGFA. The expressions of miR-9, VEGFA, and cluster of differentiation 31 (CD31) of the rats' tissues and cells were examined. The targeting relationship between miR-9 and VEGFA was testified. The tubule formation, the cell proliferation and the periodic distribution and apoptosis were evaluated after transfection. In the retinal tissues of DR rats, miR-9 expression decreased while the expression of VEGFA and CD31 increased. Notably, miR-9 targeted and inhibited VEGFA expression. In response to the treatment of miR-9 mimic and siRNA-VEGFA, a reduction was identified in CD31 expression, tubule formation, and proliferation of RMECs and cell ratio in the S phase, but an increase was observed in apoptosis rate of RMECs. The treatment of miR-9 inhibitor reversed the manifestations. Our study demonstrated that miR-9 could inhibit retinal neovascularization of DR and tubule formation, and promote apoptosis in RMECs by targeting VEGFA.