All-trans retinoic acid alleviates transmissible gastroenteritis virus-induced intestinal inflammation and barrier dysfunction in weaned piglets

Background Transmissible gastroenteritis virus(TGEV)is one of the main pathogens causing severe diarrhea of pig-lets.The pathogenesis of TGEV is closely related to intestinal inflammation.All-trans retinoic acid(ATRA)is the main active metabolite of vitamin A,which has immunomodulatory and anti-inflammatory properties.However,it is unclear whether ATRA can alleviate TGEV-induced intestinal inflammation and barrier dysfunction in piglets.This study aimed to investigate the effects of ATRA on growth performance,diarrhea,intestinal inflammation and intesti-nal barrier integrity of TGEV-challenged piglets.Methods In a 19-d study,32 weaned piglets were randomly divided into 4 treatments:Control group(basal diet),TGEV group(basal diet+TGEV challenge),TGEV+ATRA5 group(basal diet+5 mg/d ATRA+TGEV challenge)and TGEV+ATRA15 group(basal diet+15 mg/d ATRA+TGEV challenge).On d 14,piglets were orally administered TGEV or the sterile medium.Results Feeding piglets with 5 and 15 mg/d ATRA alleviated the growth inhibition and diarrhea induced by TGEV(P<0.05).Feeding piglets with 5 and 15 mg/d ATRA also inhibited the increase of serum diamine oxidase(DAO)activ-ity and the decrease of occludin and claudin-1 protein levels in jejunal mucosa induced by TGEV,and maintained intestinal barrier integrity(P<0.05).Meanwhile,5 mg/d ATRA feeding increased the sucrase activity and the expres-sions of nutrient transporter related genes(GLUT2 and SLC7A1)in jejunal mucosa of TGEV-challenged piglets(P<0.05).Furthermore,5 mg/d ATRA feeding attenuated TGEV-induced intestinal inflammatory response by inhibit-ing the release of interleukin(IL)-1β,IL-8 and tumor necrosis factor-α(TNF-α),and promoting the secretion of IL-10 and secretory immunoglobulin A(sIgA)(P<0.05).Feeding 5 mg/d ATRA also down-regulated the expressions of Toll-like receptors and RIG-I like receptors signaling pathway related genes(TLR3,TLR4,RIG-I,MyD88,TRIF and MAVS)and the phosphorylation level of nuclear factor-κB-p65(NF-κB p65),and up-regulated the inhibitor kappa B alpha(IκBα)protein level in jejunal mucosa of TGEV-challenged piglets(P<0.05).Conclusions ATRA alleviated TGEV-induced intestinal barrier damage by inhibiting inflammatory response,thus improving the growth performance and inhibiting diarrhea of piglets.The mechanism was associated with the inhibi-tion of NF-κB signaling pathway mediated by TLR3,TLR4 and RIG-I.

All-trans retinoic acid regulates the expression of MMP-2 and TGF-β2 via RDH5 in retinal pigment epithelium cells

·AIM: To investigate the effect of all-trans retinoic acid(ATRA) on retinol dehydrogenase 5(RDH5), matrix metalloproteinase-2(MMP-2) and transforming growth factor-β2(TGF-β2) transcription levels, and the effect of RDH5 on MMP-2 and TGF-β2 in retinal pigment epithelium(RPE) cells.·METHODS: After adult RPE cell line-19(ARPE-19 cells) intervened with gradient concentrations of ATRA(0-20 μmol/L) for 24h, flow cytometry was used to detect the proliferation and apoptosis of cells in each group, and quantitative realtime polymerase chain reaction(q RT-PCR) was used to detect RDH5, MMP-2 and TGF-β2 m RNA expression. Then, after ARPE-19 cells transfected with three different si RNA targets for 48h, the RDH5 knockdown efficiency of each group and expression of MMP-2 and TGF-β2 m RNA within them was detected by q RT-PCR. ·RESULTS: Flow cytometry results showed that ATRA could inhibit the proliferation of RPE cells and promote the apoptosis of RPE cells, and the difference of apoptosis was statistically significant when the ATRA concentration exceeded 5 μmol/L and compared with the normal control group(P=0.027 and P=0.031, respectively). q RT-PCR results showed that ATRA could significantly inhibit the expression level of RDH5 m RNA(P<0.001) and promote the expression of MMP-2 and TGF-β2 m RNA(P=0.03 and P<0.001, respectively) in a dose-dependent manner, especially when treated with 5 μmol/L ATRA. The knockdown efficiency of RDH5 si RNA varies with different targets, among which RDH5 si RNA-435 had the highest knockdown efficiency, i.e., more than 50% lower than that of the negative control group(P=0.02). When RDH5 was knocked down for 48h, the results of q RT-PCR showed that the expressions of MMP-2 and TGF-β2 m RNA were significantly up-regulated(P<0.001).·CONCLUSION: ATRA inhibits the expression of RDH5 and promotes MMP-2 and TGF-β2, and further RDH5 knockdown significantly upregulates MMP-2 and TGF-β2. These findings suggest that RDH5 may be involved in an epithelial-mesenchymal transition of RPE cells mediated by ATRA.

CD71-mediated liposomal arsenic-nickel complex combined with all-trans retinoic acid for the efficacy of acute promyelocytic leukemia

Clinically,arsenic trioxide(ATO)was applied to the treatment of acute promyelocytic leukemia(APL)as a reliable and effective frontline drug.However,the administration regimen of AsⅢwas limited due to its fast clearance,short therapeutic window and toxicity as well.Based on CD71 overexpressed on APL cells,in present study,a transferrin(Tf)-modified liposome(LP)was established firstly to encapsulate AsⅢin arsenic-nickel complex by nickel acetate gradient method.The AsⅢ-loaded liposomes(AsLP)exhibited the feature of acid-sensitive release in vitro.Tf-modified AsLP(Tf-AsLP)were specifically taken up by APL cells and the acidic intracellular environment triggered liposome to release AsⅢwhich stimulated reactive oxygen species level and caspase-3 activity.Tf-AsLP prolonged half-life of AsⅢin blood circulation,lowered systemic toxicity,and promoted apoptosis and induced cell differentiation at lesion site in vivo.Considering that ATO combined with RA is usually applied as the first choice in clinic for APL treatment to improve the therapeutic effect,accordingly,a Tf-modified RA liposome(Tf-RALP)was designed to reduce the severe side effects of free RA and assist Tf-AsLP for better efficacy.As expected,the tumor inhibition rate of Tf-AsLP was improved significantly with the combination of Tf-RALP on subcutaneous tumor model.Furthermore,APL orthotopic NOD/SCID mice model was established by 60CO irradiation and HL-60 cells intravenously injection.The effect of co-administration(Tf-AsLP+Tf-RALP)was also confirmed to conspicuous decrease the number of leukemia cells in the circulatory system and prolong the survival time of APL mice by promoting the APL cells’apoptosis and differentiation in peripheral blood and bone marrow.Collectively,Tf-modified acid-sensitive AsLP could greatly reduce the systemic toxicity of free drug.Moreover,Tf-AsLP combined with Tf-RALP could achieve better efficacy.Thus,transferrinmodified AsⅢliposome would be a novel clinical strategy to improve patient compliance,with promising translation prospects.

Nano transdermal system combining mitochondria-targeting cerium oxide nanoparticles with all-trans retinoic acid for psoriasis

Psoriasis is an inflammatory skin disease that is intricately linked to oxidative stress.Antioxidation and inhibition of abnormal proliferation of keratinocytes are pivotal strategies for psoriasis.Delivering drugs with these effects to the site of skin lesions is a challenge that needs to be solved.Herein,we reported a nanotransdermal delivery system composed of all-trans retinoic acid(TRA),triphenylphosphine(TPP)-modified cerium oxide(CeO2)nanoparticles,flexible nanoliposomes and gels(TCeO_(2)-TRA-FNL-Gel).The results revealed that TCeO_(2)synthesized by the anti-micelle method,with a size of approximately 5 nm,possessed excellent mitochondrial targeting ability and valence conversion capability related to scavenging reactive oxygen species(ROS).TCeO_(2)-TRA-FNL prepared by the film dispersion method,with a size of approximately 70 nm,showed high drug encapsulation efficiency(>96%).TCeO_(2)-TRA-FNL-Gel further showed sustained drug release behaviors,great transdermal permeation ability,and greater skin retention than the free TRA.The results of in vitro EGF-induced and H2O2-induced models suggested that TCeO_(2)-TRA-FNL effectively reduced the level of inflammation and alleviated oxidative stress in HaCat cells.The results of in vivo imiquimod(IMQ)-induced model indicated that TCeO_(2)-TRA-FNL-Gel could greatly alleviate the psoriasis symptoms.In summary,the transdermal drug delivery system designed in this study has shown excellent therapeutic effects on psoriasis and is prospective for the safe and accurate therapy of psoriasis.

Inhibition of all-trans retinoic acid on MDM2 gene expression in astrocytoma cell line SHG-44

Objective To investigate the impact of all-trans retinoic acid （ATRA） on MDM2 gene expression in astrocytoma cell line SHG-44, and to provide basic data for further research on the progression mechanism and gene therapy of human astrocytoma. Methods The differential expressions of MDM2 gene and protein in SHG-44 cells were detected by cDNA microarray and Western blot, respectively, before and after treatment of ATRA. The expressions of MDM2 protein in WHO grade Ⅱ and grade Ⅳ astrocytomas were determined by immunohistochemical streptavidin-peroxidase method. Some differentially expressed genes were selected randomly for Northern blot analysis. Results The intensity ratio of ATRA-treated to untreated SHG-44 cell was 0.37 in the cDNA microarray, suggesting that the expression of MDM2 gene was down-regulated in SHG-44 cells after treatment with ATRA. Some genes differentially expressed in the microarray were confirmed by Northern blot. Western blot demonstrated that the optical density ratios of MDM2 to β-actin in ATRA-treated and untreated SHG-44 were 14.02±0.35 and 21.40±0.58 （t = 24.728, P = 0.000）, respectively, suggesting that the expression of MDM2 protein was inhibited in ATRA-treated SHG-44 cells. Moreover, the percentages of MDM2-positive protein were 24.00% （6/25） and 56.52% （13/23） （x^2 = 5.298, P = 0.021） in WHO grade Ⅱ and grade Ⅳ astrocytomas, respectively, suggesting that the expression of MDM2 protein may increase along with the elevation of astrocytoma malignancy. Conclusion ATRA can inhibit MDM2 gene expression in SHG-44 cells, and MDM2 is related to astrocytoma progression.

Effect of retinoic acid on the changes of nuclear matrix-intermediate filament system in gastric carcinoma cells

AIM To explore the relationship between the configuration changes of the nuclear matrix-intermediate filament system in cancer cell induced by retinoic acid and the malignantphenotypic reversion of cancer cells.METHODS The human gastric adenocarcinomacell line MGc80-3 cells were induced with 10-6mol/ L retinoic acid and subouItured at cover slipstrip and gold grids. The cells were treated byselective extraction methOd and prepared for whole mount electron microscopy observation.The samples were examined respectively withscanning and transmission electron microscope.RESULTS The nuclear matrix filaments andintermediate filaments in MGC80-3 cells wererelatively few and scattered, not welldistributed and arranged irregularly. The nuclear lamina was ununiformly thick and compact,connected to the nuclear matrix filaments and intermediate filaments relaxedly. However, thetwo kinds of filaments were abundant and welldistributed, different in slender and thick form and interweaved into a regular network in the cells induced by 10-6 mol/ L RA. The nuclear matrix filaments and intermediate filaments were connected closely by the thin and compact fiber-Iike lamina, and interlaced into a regularnetwork throughout the whole cell region.CONCLUSION The NM-IF system in MGc80-3cells had undergone a restorational changesimilar to those of normaI cells after RAinducement. This alternation is an importantmorpholOgical and functional expression to themalignant phenotypic reversion of cancer cells.

Effects of retinoic acid on proliferation,phenotype and expression of cyclin-dependent kinase inhibitors in TGF-β1-stimulated rat hepatic stellate cells

AIM To study the molecular mechanisms ofretinoic acid(RA)on proliferation andexpression of cyclin-dependent kinase inhibitors(CKI),i.e.p16,p21 and p27 in cultured rathepatic stellate cells(HSC)stimulated withtransforming growth factor beta 1(TGF-β1).METHODS HSC were isolated from healthy ratlivers and cultured.After stimulated with1 mg/L TGF-β1,subcultured HSC were treatedwith or without 1 nmol/L RA.MTT assay,immunocytochemistry(ICC)for p16,p21,p27and α-smooth muscle actin(α-SMA)protein,insitu hybridization(ISH)for retinoic acidreceptor beta 2(RAR-β2)and p16,p21 and p27mRNA and quantitative image analysis(partially)were performed.RESULTS RA inhibited HSC proliferation(41.50%,P【0.05),decreased the protein levelof α-SMA(55.09%,P【0.05),and induced HSCto express RAR-β2 mRNA.In addition,RAincreased the protein level of p16(218.75%,P【0.05)and induced p21 protein expression;meanwhile,p27 was undetectable by ICC in bothcontrol and RA-treated HSC.However,RA hadno influence on the mRNA levels of p16,p21 orp27 as determined by ISH.CONCLISION Up-regulation of p16 and p21 on post-transcriptional level may contribule, in part to RA inhibition of TGF-β1-initiated rat HSC activation in vitro.

Synthesis and anti-tumor activity of all-trans retinoic acid derivatives

A series of retinoate and retinamide derivatives were designed, synthesized, and their anti-tumor activities were investigated in NB4 by MTT and flow cytometry assays （FCM）. All compounds showed cytotoxicity, especially compounds la and ld exhibited a higher cytotoxicity than other derivatives and all-trans rethaoic acid （ATRA）. Furthermore, compound 1d could induce NB4 cell lines differentiation efficiently. O 2009 Fei Hu Chert. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.

Expressions of cellular retinoic acid binding proteins I and retinoic acid receptor-β in the guinea pig eyes with experimental myopia

·AIM:All-trans retinoic acid (RA) is the only extrinsic biochemical candidate known to date that could act as a growth controller,the aim of this study was to investigate the expression cellular retinoic acid binding proteins I (CRABP-I) and retinoic acid receptor-β (RAR-β) in retina of the guinea pig eyes with experimental myopia.·METHODS:Ninety guinea pigs aged 14 days were equally and randomly divided into three groups:form deprivation (FD),-5D lens,and control.The diffusers for FD were white translucent hemispheres,and-5D lenses were used to introduce hyperopic defocus.Refraction was measured with streak retinoscopy after cycloplegia,and axial length was calculated with Cinescan A/B ultrasonography.Retina harvested at different time points were used to measure RA level with HPLC and expressions of cellular retinoic acid binding proteins I (CRABP-I) and RA receptor-β (RAR-β) were assayed with Western blot and Real-time PCR.SPSS13.0 software was used for statistical analysis.·RESULTS:Up-regulations of CRABP-I and RAR-β in ocular tissues correlated with changes in the refractive status and growth rate of the guinea pig eye (P <0.05).14 days of monocular form-deprivation led to-5.14D myopia and a 0.281mm axial elongation;14 days of monocular defocus produced-3.64D myopia and a 0.163 mm axial elongation.The level of retinal RA started to elevate in 7 days (P <0.05) after visual manipulation in both FD and-5D lens groups and became more prominent by 14 days (P <0.01).The expressions of CRABP-I and RAR-β increased by 14 days after visual manipulation (P <0.05),the mRNA level of RAR-β,however,increased by 7 days after visual manipulation (P <0.05),which suggested that changes of expressions of CRABP-I and RAR-β might lag behind the change of RA.·CONCLUSION:The levels of CRABP-I and RAR-β were elevated in retina of the guinea pig eye with experimental myopia.During the progression of experimental myopia,the retinal RA level increased rapidly,and there might be a positive feedback between the increase of RA and up-regulation of RAR-β.·

Retinoic acid receptor α promotes autophagy to alleviate liver ischemia and reperfusion injury

AIM: To study the role of autophagy and the relationship between retinoic acid receptor α(RARα) and autophagy in liver ischemia and reperfusion(IR) injury.METHODS: All-trans retinoic acid(ATRA) was administered to mice for two weeks before operation. Reverse transcription-polymerase chain reaction and Western blot were used to detect the expression levels of related factors. To demonstrate the role of RARα,LE540,a RARα inhibitor,was used to treat hepatocytes injured by H2O2 in vitro.RESULTS: ATRA pretreatment noticeably diminished levels of serum alanine aminotransferase and as-partate aminotransferase as well as the degree of histopathological changes. Apoptosis was also inhibited,whereas autophagy was promoted. In vitro,RARα was inhibited by LE540,which resulted in decreased autophagy and increased apoptosis. Similarly,the expression of Foxo3 a and p-Akt was downregulated,but Foxo1 expression was upregulated.CONCLUSION: This research provides evidence that ATRA can protect the liver from IR injury by promoting autophagy,which is dependent on Foxo3/p-Akt/Foxo1 signaling.

Retinoic acid metabolic change in retina and choroid of the guinea pig with lens-induced myopia

AIM: To investigate the role of retinoic add (RA) and retinaldehyde dehydrogenase-2 (RALDH(2)) of retina and choroid in the guinea pig lens-induced myopic eyes. METHODS: Totally 45 guinea pigs, at age of three weeks, were randomly assigned into three groups: the normal control, the lens-induced group and the recovering group. Out of focus was induced by the -6.00D concave lens on the left eye, and lasted for 15 days. All animals underwent biometric measurement (corneal radius of curvature, refraction and axial length). Subsequently, RA content in the retina and RPE/choriod complex was detected by reversed-phase high-performance liquid chromatography. RALDH(2) protein in the retina and RPE/choriod complex was evaluated by the immunohistochemical staining and Western blotting. RESULTS: After wearing -6.00D lens for 15 days, axial length of the lens-induced eye extends and myopia was formed, with RA contents increasing in both the neural retina and RPE/choroid complex. Comparing with the lens-induced group, myopic degree significantly relieved, and its RA contents in both the neural retina and RPE/choroid complex decreased in the recovering group. In the normal control, RALDH(2) protein was expressed positively in the retinal nerve fiber layer (RNFL), inner plexiform layer (IPL) and lateral border of outer nuclear layer (ONL). Retinal RALDH(2) protein increased in the lens-induced group, and was also positive in the outer plexiform layer (OPL). In the recovering group, retinal RALDH(2) protein attenuated the expression in the OPL turns to negative. RALDH(2) protein was not expressed in the choroid of any group. CONCLUSION: RA of retina and chorid participates in the regulation of the lens-induced myopia in guinea pigs, which may be related with retinal RALDH(2) protein.

The effect pathway of retinoic acid through regulation of retinoic acid receptor α in gastric cancer cells

AIM To evaluate the role of RARα gone in mediating the growth inhibitory effect of all-trans retinoic acid(ATRA) on gastric cancer cells. METHODS The expression levels of retinoic acid receptors(PARs)in gastric cancer cells were detected by Northern blot.Transient transfection and chlorophenicol acetyl transferase(CAT)assay were used to show the transcriptional activity of β retinoic acid response element (βPARE)and AP-1 activity.Cell growth inhibition was determined by MTT assay and anchorage-independent growth assay,respectively.Stable transfection was performed by the method of Lipofectamine,and the cells were screened by G418. RESULTS ATRA could induce expression level of RARα in MGC80-3,BGC-823 and SGC-7901 cells obviously, resulting in growth inhibition of these cell lines.After sense RARα gone was transfected into MKN-45 cells that expressed rather low level of RARα and could not be induced by ATPA,the cell growth was inhibited by ATPA markedly.In contrast,when antisense RARα gone was transfected into BGC-823 cells,a little inhibitory effect by ATPA was seen,compared with the parallel BGC-823 cells.In transient transfection assay,ATPA effectively induced transcriptional activity of βRARE in MGC80-3, BGC-823,SGC-7902 and MKN/RARα cell lines,but not in MKN-45 and BGC/aRARα cell lines.Similar results were observed in measuring anti-AP-1 activity by ATPA in these cancer cell lines. CONCLUSION ATRA inhibits the growth of gastric cancer cells by up-regulating the level of RARα; RARα is the major mediator of ATRA action in gastric cancer cells;and adequate level of RARα is required for ATRA effect on gastric cancer cells.

Study on the Structure of Supramolecular Inclusion Complex of β-Cyclodextrin with Retinoic Acid

Inclusion compound of retinoic acid with (-cyclodextrin was prepared by coprecipitating method, the structure of resulting product was studied by elemental analysis, differential scanning caloriemetry(DSC) analysis, FT-IR spectroscopy and X-ray diffractometry, and the formed supramolecule self-assembles in aqueous solution according to molar ratio 2:1 of host-guest.

Mechanism of Retinoic Acid and Mitogen-activated Protein Kinases Regulating Hyperoxia Lung Injury

To investigate the protective effect of retinoic acid （RA） on hyperoxic lung injury and the role of RA as a modulator on mitogen-activated protein kinases （MAPKs）, gastation 21 d Sprague- Dawley （SD） fetuses （term = 22 d） were delivered by hysterotomy. Within 12-24 h of birth, premature rat pups were randomly divided into 4 groups （n= 12 each） ： air-exposed control group （group Ⅰ ） ; hyperoxia-exposed group （ group Ⅱ ）, air-exposed plus RA group （group Ⅲ ）, hyperoxia-exposed plus RA group （group Ⅳ）. Group Ⅰ , Ⅲ were kept in room air, and group Ⅱ , Ⅳ were placed in 85 % oxygen. The pups in groups Ⅲ and Ⅳ were intraperitoneally injected with RA （500 μg/kg every day）. All lung tissues of premature rat pups were collected at the 4th day after birth. Terminal transferase d-UTP nick end labeling （TUNEL） staining was used for the detection of cell apoptosis. The expression of PCNA was immunohistochemically detected. Western blot analysis was employed for the determination of phosphorylated and total nonphosphorylated ERKs, JNKs or p38. Our results showed that lungs from the pups exposed to hyperoxia for 4 d exhibited TUNEL-positive nuclei increased markedly throughout the parenchyma （P〈0.01）, and decreased significantly after RA treatment （P〈0.01）. The index of PCNA-positive cells was significantly decreased （P〈0.01）, and was significantly increased by RA treatment （P〈0.01）. The air-space size was significantly enlarged, secondary crests were markedly decreased in hyperoxia-exposed animals. RA treatment improved lung air spaces and secondary crests in air-exposed pups, hut had no effect on hyperoxia-exposure pups. Western blotting showed that the amounts of JNK, p38 and ERK proteins in hyperoxia-exposure or RA-treated lung tissues were same as those in untreated lung tissues （P〈0.05）, whereas activation of these MAPKs was markedly altered by hyperoxia and RA. After hyperoxia exposure, p-ERK1/2, p-JNK1/2 and p-p38 were dramatically increased （P〈0.01）, whereas p-JNK1/2 and p-p38 were markedly declined and p-ERK1/2 was further elevated by RA treatment （P〈0.01）. It is concluded that RA could decrease cell apoptosis and stimulate cell proliferation under hyperoxic condition. The protection Of RA on hyperoxia-induced lung injury was related＇to the regulation of MAP kinase activation.

Expression of the retinoic acid-metabolizing enzymes RALDH2 and CYP26b1 during mouse postnatal testis development

Aim： To study the expression pattern of the retinoic acid metabolizing enzymes RALDH2 and CYP26bl during mouse postnatal testis development at both mRNA and protein levels. Methods： Real-time polymerase chain reaction and Western blot analysis were performed to determine the relative quantity of RALDH2 and CYP26bl at both mRNA and protein levels at postnatal day 1, 5, 10, 20, and in adult mice （70 days testes）. Testicular localization of RALDH2 and CYP26b 1 during mouse postnatal development was examined using immunohistochemistry assay. Results： Aldhla2 transcripts and its protein RALDH2 began to increase at postnatal day 10, and remained at a high level through postnatal day 20 to adulthood. Cyp2661 transcripts and CYP26bl protein did not change significantly during mouse postnatal testis development. RALDH2 was undetectable in the postnatal 1, 5 and 10 day testes using immunohis- tochemistry assay. At postnatal day 20 it was detected in pachytene spermatocytes. Robust expression of RALDH2 was restricted in round spermatids in the adult mouse testis. In the developing and adult testis, CYP26bl protein was confined to the peritubular myoepithelial cells. Conclusion： Our results indicate that following birth, the level of retinoic acid in the seminiferous tubules might begin to increase at postnatal day 10, and maintain a high level through postnatal day 20 to adulthood.

Inhibition of matrix metalloproteinases expression in human dental pulp cells by all-trans retinoic acid

All-trans retinoic acid（ATRA） inhibits matrix metalloproteinase（MMP）-2 and MMP-9 in synovial fibroblasts, skin fibroblasts,bronchoalveolar lavage cells and cancer cells, but activates MMP-9 in neuroblast and leukemia cells. Very little is known regarding whether ATRA can activate or inhibit MMPs in human dental pulp cells（HDPCs）. The purpose of this study was to determine the effects of ATRA on the production and secretion of MMP-2 and-9 in HDPCs. The productions and messenger RNA（mRNA） expressions of MMP-2 and-9 were accessed by gelatin zymography and real-time polymerase chain reaction（PCR）, respectively. ATRA was found to decrease MMP-2 level in a dose-dependent manner. Significant reduction in MMP-2 mRNA expression was also observed in HDPCs treated with 25 mmol？L21ATRA. However, HDPCs treated with ATRA had no effect on the pattern of MMP-9 produced or secreted in either cell extracts or conditioned medium fractions. Taken together, ATRA had an inhibitory effect on MMP-2 expression in HDPCs,which suggests that ATRA could be a candidate as a medicament which could control the inflammation of pulp tissue in vital pulp therapy and regenerative endodontics.

Role of miR-124 in the regulation of retinoic acid-induced Neuro-2A cell differentiation

Retinoic acid can cause many types of cells,including mouse neuroblastoma Neuro-2 A cells,to differentiate into neurons.However,it is still unknown whether microRNAs(miRNAs)play a role in this neuronal differentiation.To address this issue,real-time polymerase chain reaction assays were used to detect the expression of several differentiation-related miRNAs during the differentiation of retinoic acid-treated Neuro-2 A cells.The results revealed that miR-124 and miR-9 were upregulated,while miR-125 b was downregulated in retinoic acid-treated Neuro-2 A cells.To identify the miRNA that may play a key role,miR-124 expression was regulated by transfection of miRNA mimics or inhibitors.Morphological analysis results showed that inhibition of miR-124 expression reversed the effects of retinoic acid on neurite outgrowth.Moreover,miR-124 overexpression alone caused Neuro-2 A cells to differentiate into neurons,and its inhibitor could block this effect.These results suggest that miR-124 plays an important role in retinoic acid-induced differentiation of Neuro-2 A cells.

All-trans retinoic acid upregulates VEGF expression in glioma cells in vitro

All-trans retinoid acid （ATRA） is one of the most potent and most thoroughly studied differentiation inducers that induce the differentiation and apoptosis of glioma cells. However, the effect of ATRA on angiogenesis of glioma re- mains poorly understood. We examined the effect of ATRA on the expression of vascular endothelial growth fac- tor （VEGF） in different glioma cell lines and investigated the underlying mechanism, intending to partially reveal the effects of ATRA on angiogenesis of glioma. Glioma cells were treated by ATRA at 5 and 10 μmol/L. The VEGF mRNA transcript levels were determined by real-time RT-PCR and the protein levels of VEGF in glioma cells were evaluated by Western blotting assays. Moreover, hypoxia-inducible factor-1α （HIF-la） mRNA expression was analyzed by using real-time RT-PCR. After treatment with 5 and 10 μmol/L ATRA, the VEGF mRNA tran- script levels in glioma cells increased remarkably, compared with that in the control group, and the relative protein expression of VEGF was also up-regulated. Meanwhile, the HIF-la mRNA expression also increased. ATRA in- creases the expression of VEGF in glioma cells at both transcriptional and translational levels.

Effect of All-trans Retinoic Acid on Liver Fibrosis Induced by Common Bile Duct Ligation in Rats

The aim of this study was to investigate the effect and possible mechanism of all-trans retinoic acid （ATRA） on liver fibrosis induced by common bile duct ligation （CBDL） in rats. Fifty-three female Wistar rats were randomly divided into 5 groups： sham operation group （group J, 5 animals） and groups A, B, C and D （12 animals in each group）. The rats in groups A, B, C and D were subjected to CBDL to induce liver fibrosis, while those in group J to sham operation. From the 3rd week the rats in groups B, C and D respectively received daily administration of ATRA via gas- tric tube at three different doses [0.1, 1.5 and 7.5 mg/kg body weight （BW）]. Animals were sacrificed at 6th week. Rats＇ liver tissues were observed for pathologic changes under a light microscope. The protein levels of type Ⅰ collagen （COLⅠ）, matrix metalloproteinase-2 （MMP2）, MMP13 and tissue inhibitors of metalloproteinase-1 （TIMP-1） in liver tissues were determined by immunohistochemical techniques. The expression levels of TGF-β1 and CTGF mRNA in liver tissues were detected by semi-quantitative reverse transcription-polymerase chain reaction （RT-PCR）. The results showed that loss of normal hepatic architecture and formation of obvious fibrosis were observed in group A, while ATRA treatment for 4 weeks notably alleviated the pathological changes of hepatocytes. The expres- sion of COL Ⅰ and TIMP-1 proteins in group A was increased, while decreased in ATRA-treated CBDL groups （P〈0.05）. ATRA （1.5 and 7.5 mg/kg BW） reduced the expression levels of COLⅠ protein more greatly than that of 0.1 mg/kg BW （P〈0.05）. ATRA treatment increased the protein levels of MMP2 and MMP13. The expression levels of TGF-β1 and CTGF mRNA in group A were increased. In comparison with group A, the mRNA levels of TGF-β1 and CTGF in ATRA-treated CBDL groups were significantly decreased （P〈0.05）. It was concluded that ATRA could inhibit CBDL-induced liver fibrosis in rats by suppressing the expression of TGF-β1 and CTGF so as to di- minish the inhibition of TIMP-1 on MMP2 and MMP13 and increase the activity of MMP2 and MMP13.