The biological function of type I receptors of bone morphogenetic protein in bone

Bone morphogenetic proteins （BMPs） have multiple roles in skeletal development, homeostasis and regeneration. BMPs signal via type I and type II serine/threonine kinase receptors （BMPRI and BMPRII）. In recent decades, genetic studies in humans and mice have demonstrated that perturbations in BMP signaling via BMPRI resulted in various diseases in bone, cartilage, and muscles. In this review, we focus on all three types of BMPRI, which consist of activin-like kinase 2 （ALK2, also called type IA activin receptor）, activin- llke kinase 3 （ALK3, also called BMPRIA）, and activin-like kinase 6 （ALK6, also called BMPRIB）. The research areas covered include the current progress regarding the roles of these receptors during myogenesis, chondrogenesis, and osteogenesis. Understanding the physiological and pathological functions of these receptors at the cellular and molecular levels will advance drug development and tissue regeneration for treating musculoskeletal diseases and bone defects in the future.

Influence of bone morphogenetic protein type IA receptor conditional knockout in lens on expression of bone morphogenetic protein 4 in lens

AIM: To investigate the influence of bone morphogenetic protein type IA receptor [BMPR-IA(ALK3)] conditional knockout in lens on expression of bone morphogenetic protein 4(BMP4) in lens during the development of the vertebrate eye.METHODS: Cre-positive mice were mated with Crenegative mice to generate 50% Cre-positive(conditional knockout, CKO) 4 embryos, 8 eyes and 50% Cre-negative offspring(wild type, WT) 4 embryos, 8 eyes. The embryos were fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned to a thickness of 4 μm.Removal of paraffin wax and dehydrating for sections,and then the procedure of in situ hybridization was processed, BMP4 MK1784-m(BOSTER) was used, and observed the expression of BMP4 in the lens in experimental group and control group. We selected SPSS11.0 software for statistical analysis, P【0.05 showed that the difference was statistically significant.· RESULTS: Four embryos of each genotype were examined, totally we had 8 embryos, 16 eyes. We got the uniform outcomes in all the embryos. We found ALK3 was required during lens growing, but was not essential for the formation of lens. We observed that the expression of BMP4 in the lens was significantly reduced in all 8 ALK3 CKO lens, BMP4 expression was normal in all the 8 WT lens, P 【0.01. This phenomenon became increasingly visible in accordance with embryo development. The most apparent alteration was present at stage E15.5.CONCLUSION: ALK3 is essential for lens growth. The influence of ALK3 on the expression of BMP4 is present during the development of mice lens.

Establishment and identification of fibroblast clones expressing human bone morphogenetic protein 2

Objective:To establish fibroblasts stably expressing human bone morphogenetic protein 2(hBMP2). Methods:Eukaryonic expression vector(pcDNA3.1-B2) was transduced into NIH3T3 cells using SofastTM, a new generation cationic polymer gene transfection reagent. The positive cell clones were selected with G418. The stable transfection and expression of BMP2 in the NIH3T3 cells were determined by RT-PCR and immunohistochemical stain. Results: BMP2 mRNA was transcripted and expressed in the transfected NIH3T3 cells. Conclusion: With positive compound transfection, outside human BMP2 gene can be successfully transducted into NIH3T3 cells, which is the key step to induce periodontal cells to osseous phenotypes.

Expression of Vascular Endothelial Growth Factor in Bone Morphogenetic Protein-2 Induced Osteogenesis

An experimental model of femoral muscular pouch in 20 mice was adopted. The expression of VEGF was examined by in situ hybridization method and immunohistochemical method in bone morphogenetic protein- 2 induced osteogenesis . The experimental results demonstrated that the expression signals of VEGF mRNA and VEGF appeared in cytoplasm during condensation of mesenehymal cell. As the mesenchymal cells differentiated into precartilage, the expression signals decreased in mesenehymal cells, but increased in chondrocytes and kept getting denser in the process of cartilage maturity. The peak expression of VEGF mRNA and VEGF in the experimental group appeared on the 14 th day, accompanied by numerons hypertrophic chondrocytes. When mature cartilage calcified and neu, bone trabecula formed, the expression of VEGF mRNA and VEGF decreased in chondrocytes, but still expressed moderately in the osteoblasts and osteocytes. Signals of VEGF mRNA and VEGF can not be detected in the control groups.

Roles and regulation of bone morphogenetic protein-7 in kidney development and diseases

The gene encoding bone morphogenetic protein-7(BMP7) is expressed in the developing kidney in embryos and also in the mature organ in adults. During kidney development, expression of BMP7 is essential to determine the final number of nephrons in and proper size of the organ. The secreted BMP7 acts on the nephron progenitor cells to exert its dual functions: To maintain and expand the progenitor population and to provide them with competence to respond to differentiation cues, each relying on distinct signaling pathways. Intriguingly, in the adult organ, BMP7 has been implicated in protection against and regeneration from injury. Exogenous administration of recombinant BMP7 to animal models of kidney diseases has shown promising effects in counteracting inflammation, apoptosis and fibrosis evoked upon injury. Although the expression pattern of BMP7 has been well described, the mechanisms by which it is regulated have remained elusive and the processes by which the secretion sites of BMP7 impinge upon its functions in kidney development and diseases have not yet been assessed. Understanding the regulatory mechanisms will pave the way towards gaining better insight into the roles of BMP7, and to achieving desired control of the gene expression as a therapeutic strategy for kidney diseases.

Construction of Adeno-associated Virus System for Human Bone Morphogenetic Protein 7 Gene

To construct the recombinant adeno-associated virus （rAAV） vector with human bone morphogenetic protein 7 （BMP7） and observe the BMP7 mRNA expression in vitro, BMP7 CDS sequence was cloned into expression plasmid pAAV-MCS of AAV Helper Free System. The recombinant plasmid was identified with enzyme digestion and sequencing. The recombinant plasmid, pAAV-RC, pHelper were co-transfected into AAV-293 cells according to the calcium phosphate-based protocol. The viral stock was collected by 4 rounds of freeze/thaw. After purified and concentrated, the recombinant virus titer was determined by dot-blot assay. HEK293 cells were transfected with the recombinant virus at different MOI, and the expression of BMP7 mRNA was detected by RT-PCR. The results showed rAAV-BMP7 was constructed and packaged successfully. The physical particle titer was 2.5×10^11 vector genomes/mL. There was different expression level of BMP7 mRNA after transfecton. These data suggested that recombinant AAV mediated a stable expression of hBMP7 mRNA in 293 cells. The AAV production method may pave the way of an effective strategy for the jaw bone defection around dental implants.

Bone Morphogenetic Protein 15 as a Candidate Gene for Prolificacy of Jining Grey Goat

On the basis of the ovine bone morphogenetic protein 15(BMP15)gene,two pairs of primers(PI and P2)were designed to amplify exons 1 and 2 of the BMP15 gene in five randomly selected does of both Angora and Jining Grey goats.The sequences of BMP15 exon 1(P1 amplification)of Angora and Jining Grey goats were identical.There was a 3-nucleotide(CTT)insertion in positions 268 to 270 of goat BMP 15 exon1 compared with that of sheep(GenBank accession number AF236078),which caused a leucine insertion in the 12th position of amino acid sequence.Sequence length of goat BMP 15 exon 2(P2 amplification)was identical with that of sheep(AF236079),but there were seven nucleotide and four amino acid changes between goat and sheep.The nucleotide in the 963rd position of BMP15 exon 2 was A for Angora goat and sheep,and G for Jining Grey goat.Based on this A963G mutation,primer pair P3 was designed to detect single nucleotide polymorphism of BMP15 exon 2 in breeds of high prolificacy(Jining Grey),moderate prolificacy(Boer)and low prolificacy(Angora and Inner Mongolia Cashmere)by polymerase chain reactionsingle strand conformation polymorphism(PCR-SSCP).Three genotypes(AA,AG and GG)were detected in Jining Grey goats,two genotypes(AG and GG)in Boer,and only the AA genotype in Angora and Inner Mongolia Cashmere goats.Sequencing revealed one mutation(A963G)in genotype GG compared with genotype AA,and this mutation resulted in an amino acid change of serine→glycine(S300G).In Jining Grey goats,frequencies of AA,AG and GG genotypes were 0.008,0.059 and 0.933,respectively.Genotypic distributions of the BMP 15 gene were significantly different(P<0.05 or P<0.001)between Jining Grey and Boer,Angora,and Inner Mongolia Cashmere goats.In Jining Grey goats,the does with the GG genotype had 0.71(P<0.05)or 1.57(P<0.05)additional kids than did those with AG or AA genotypes,and does with the AG genotype had 0.86(P<0.05)more kids than did those with the AA genotype.These results tentatively indicate that the BMP15 gene is either a major gene that affects prolificacy in Jining Grey goats,or may be a molecular marker in close linkage with such a gene.

Cloning, expression profiling and promoter functional analysis of bone morphogenetic protein 2 in the tongue sole(Cynoglossus semilaevis)

BMP2 plays crucial roles in vertebrate developmental process and acts as a bone inducer during osteogenesis. We present here the molecular cloning of bmp2 cDNA from the marine flatfish Cynoglossus semilaevis, and the analysis of bmp2 expression profiling and promoter function. The full length of bmp2 cDNA sequence is 2 048 bp,which encodes a protein of 422 amino acids. Tissue expression distribution of bmp2 was examined in 14 tissues of mature individuals by quantitative real time PCR（qRT-PCR）. The results revealed that bmp2 was expressed ubiquitously, and the highest expression level was detected in the spinal cord. Moreover, bmp2 expression levels were detected at 15 sampling time points of early developmental stages（egg, larva, juvenile and fingerling stages）.The highest expression level of bmp2 was observed at the gastrula stage, which was about ten times higher than those at the other three embryo stages. Whole-mount in situ hybridization showed that the bmp2 signal was strongly detected at the location of the crown-like larval fin, heart and liver, and slightly expressed in the notochord at one day post hatch（dph）; then the expression of bmp2 started to be concentrated in notochord at three dph. Subsequently, we characterized the 5′-flanking region of bmp2 by testing the promoter activity by Luciferase reporter assays. Positive regulatory region was detected at the location of –179 to ＋109. The predicted transcription factor binding sites（E-box binding factors, zinc finger transcription factor, etc.） in this region might participate in the transcriptional regulation of the bmp2 gene.

Molecular Cloning and Sequence Analysis of FullLength cDNA Encoding Human Bone Morphogenetic Protein—7

Objective:To clone the full-length human bane morphogenetic protein-7 (BMP-7 ) gene and analyse its sequence, to aid in investigation of its function and structure. Methods : Total RNA was isolated from Chinese fetal kidney by the acid gmnidinium thiocyanate phenol-chloroform method. Two overlapping segments of human BMP- 1 cDNA were obtained by reverse transcription (RT)-PCR. Following application, the two segments were ligated to each other and subcloned into POEM-T easy vector to form PEGM-T easy/hBMP-7 recombinant plasmid. Sanger dideoxy chain-termination method was used to sequence the cDNA. Results. There was 750 bp fragment obtained RT-PCR using #2 primer from 5' end of BMP-7 gene (PCR by using # 2 and # 1) ,and 540 bp fragment from 3' end was generated by KT-PCR using # 4 primer (PCR using # 3 and # 4). Full-length cDNA encoding BMP-7 was obtained by religation of two segments. When compared with hBMP-7 sequence in Gene bank (XM30619) ,our full-length BMP-7 cDNA has a G instead of a T at nucleotide 862. This change results in valine substituting for phenylalanine in the protein. Conclusion. This is the first time that BMP-7 cDNA was successfully cloned from Chinese fetal kidney. BMP-7 cDNA plays an important role in healing injuries of the osteo-articular system. This makes BMP-7 is an attractive target far various clinical applications.

Construction of bicistronic green fluorescent protein labeled pSELECT GFPzeo human bone morphogenetic protein 2 eukaryotic expression vector

Objective To construct green fluorescent protein (GFP)-labeled pSELECT-GFP zeohBMP2 eukaryotic expression vector.Methods The encoding fragment of hBMP2 gene was obtained from a recombinant plasmid pcDNA3.1/CT-hBMP2 by using polymerase

Expression of mature peptide of human bone morphogenetic protein-2 in Escherichia coli

To express die mature peptide of human bone morphogenetic protein-2 in Escherichia coil. Methods: TheDNA fragment encoding the mature peptide of human bone morphogenetic protein-2 (hBMP-2m) was inserted into expression vectorpDH in which foreign gene was controlled by PRPL promoters. E. coli DH5a transformed with recombinant plasmid pDHB2m wasinduced at 42℃to express the target protein. The expressed product was partially purified and refolded, and then implanted intorat thigh muscles to assay its bone inductive activity. Results: After induction, a protein band on SDS-PAGE gel with an apparentmol. wt. of 13kD was observed to anticipate in the strain carrying pDHB2m, but not in the control. The expressed hBMP-2m accounted for 45%-60% of the total bacterial protein. The expressed product existed in a form of inclusion body. After partially purified and refolded, rhBMP-2m could induce the formation of cartilage and bone tissue heterotopically. Conclusion: The maturepeptide of human bone morphogenetic protein-2 has ben successfully expressed in E. coli and the product has ectopic bone inductive activity.

CO-TRANSFECTION OF RAT BONE MARROW MESENCHYMAL STEM CELLS WITH HUMAN BMP2 AND VEGF165 GENES

Objective To explore the feasibility and efficacy of lentivirus-mediated co-transfection of rat bone marrow mesenchymal stem cells (MSCs) with human vascular endothelial growth factor 165 (hVEGF165) gene and human bone morphogenetic protein 2 (hBMP2) gene. Methods The hVEGF165 and hBMP2 cDNAs were obtained from human osteosarcoma cell line MG63 and cloned into lentiviral expression vectors designed to co-express the copepod green fluorescent protein (copGFP). The expression lentivector and packaging Plasmid Mix were co-transferred to 293TN cells, which produced the lentivirus carrying hVEGF165 (Lv-VEGF) or hBMP2 (Lv-BMP), respectively. MSCs of Wistar rats were co-transfected with Lv-BMP and Lv-VEGF (BMP+VEGF group), or each alone (BMP group and VEGF group), or with no virus (Control group). The mRNA and protein expressions of hVEGF165 and hBMP2 genes in each group were detected by real-time PCR and enzyme linked immunosorbent assay (ELISA). Results Lentiviral expression vectors carrying hVEGF165 or hBMP2 were correctly constructed and confirmed by restriction endonucleses analysis and DNA sequencing analysis. A transfer efficiency up to 90% was archieved in all the transfected groups detected by the fraction of fluorescent cells using fluorescent microscopy. From the results generated by real-time PCR and ELISA, VEGF165 and BMP2 genes were co-expressed in BMP+VEGF group. No significant difference of BMP2 expression was detected between BMP+VEGF and BMP groups (P>0.05). Similarly, there was no significant difference of VEGF165 expression between BMP+VEGF and VEGF groups (P>0.05). Conclusion VEGF165 and BMP2 genes were successfully co-expressed in MSCs by lentivirus-mediated co-transfection, which provided a further foundation for the combined gene therapy of bone regeneration.

Expression of BMP-7 and its receptors in renal tubolo-interstitial fibrosis induced with unilateral ureteral obstruction in rats

Objective:To study the changes of the expression of bone morphogenetic protein-7(BMP-7) and its receptors(BMPR-Ⅱ,ALK2,ALK3 and ALK6) in the renal tubulointerstitial fibrosis induced with unilateral ureteral obstruction in rats. Methods: Sixty Wistar male rats were divided randomly into the normal control,sham-operation and unilateral ureteral obstruction(UUO) groups and the rats were killed on the 1st,3rd,7th and 14th postoperative days respectively.The mRNA level of BMP-7,BMPR-Ⅱ,ALK2,ALK3 and ALK6 was determined with RT-PCR.The site and level of protein expression of BMP-7 were observed with immunohistochemical staining. Results: The mRNA level of BMP-7,BMPR-Ⅱ,ALK2 and ALK3 was significantly decreased in the rats of UUO group than in those of the sham-operation group but the mRNA level of ALK6 showed no obvious changes in all the rats.Immunohistochemical staining revealed that the protein of BMP-7 was mainly expressed in the renal tubules and interstitial tissue of the kidneys in normal rats but it was decreased gradually along with the unilateral ureteral obstruction. Conclusion: It is found that the loss of BMP-7 and its receptors including BMPR-Ⅱ,ALK2 and ALK3 occurs in the early phase of renal tubulointerstitial fibrosis before the appearance of other pathological changes in the kidney and may play an important role in the occurrence and progress of renal tubulointerstitial fibrosis.

绵羊骨形态发生蛋白受体-1B基因研究进展

绵羊繁殖性能在绵羊产业中有着重要意义,除通过一些技术手段(如同期发情、人工授精、超数排卵等)之外,在目前已知的关于能够提高绵羊繁殖力的16个基因共20个突变体当中,以骨形态发生蛋白受体-1B(bone morphogenetic protein receptor type-1B,BMPR-1B)基因对绵羊繁殖力的影响最大。BMPR-1B基因是世界上第一个被发现的多羔主效基因,其编码区的A746G突变导致蛋白质序列中第249位的谷氨酰胺被置换为精氨酸(Q249R),最终能够引起绵羊排卵数和产羔数增加。作者介绍了绵羊多羔主效基因BMPR-1B及其突变体FecB(A746G)的发现与结构,简述了该基因分子方面的作用机理,对BMP/Smad信号通路的调控以及与绵羊繁殖之间的联系,并简单分析了FecB突变后对绵羊卵巢、卵泡等组织细胞功能,激素调节和相关基因表达的影响。进一步加深对BMPR-1B基因的了解,为研究人员探明该基因诱使绵羊等动物提高产羔数的调控机制,相关配体、调控因子和上下游信号蛋白的影响以及加快哺乳动物高效育种繁殖、扩大种群规模和多胎品系的建立,增加养殖人员的经济收入等提供一些参考和帮助。

BMP亚家族成员中影响绵羊繁殖性能的基因研究进展

排卵数和产羔数是反映绵羊繁殖性状的关键性指标,也是绵羊重要的经济性状,二者属于中低遗传力性状,受基因型、环境和营养状况等多重因素共同影响。由BMP、GDF和AMH等组成的BMP亚家族是TGF-β超家族的重要组成部分,BMPR1B、BMP15和GDF9是目前与绵羊繁殖性能相关性研究最为深入的基因。该文对BMP亚家族成员中提升绵羊排卵数及产羔数的基因的SNP位点进行归纳,总结了这些基因影响繁殖性能的SNP位点的发现,品种、定位、表型,以及对排卵数和产羔率的影响机制,旨在为绵羊多胎品种的选育和高效繁殖提供理论参考。

骨形态发生蛋白/Wnt信号通路调控成骨:揭示骨骼形成和重塑的分子机制

背景:成骨细胞是负责骨骼形成和重塑的主要细胞类型,其功能的正常发挥受到多种信号通路的精密调控,其中,骨形态发生蛋白和Wnt信号通路在成骨过程中起着关键作用。目的:综述了骨形态发生蛋白/Wnt信号通路在调控成骨细胞功能中的作用,并分析其在不同生理和病理条件下的变化,以期进一步揭示骨骼形成和重塑的分子机制。方法:以中文检索词“BMP信号通路,Wnt信号通路,成骨”及英文检索词“BMP signaling pathway,Wnt signaling pathway,osteogenesis”在中国知网、万方和PubMed数据库中检索研究原著,检索时限为各数据库建库至2023年6月的相关文献,最终筛选61篇文献进行分析总结。采用文献综述的方法,对骨形态发生蛋白/Wnt信号通路调控成骨作用的研究进行梳理和分析。结果与结论:①骨形态发生蛋白和Wnt信号通路在成骨细胞的分化、增殖和成熟过程中发挥重要作用。骨形态发生蛋白信号通路主要通过激活Smad蛋白来调控成骨相关基因的表达,Smad蛋白被激活后进入细胞核,调控与成骨相关的基因表达,与骨形态发生蛋白不同,Wnt信号通路主要依赖于β-catenin的激活来发挥生物学效应。②不同生理和病理状态下,骨形态发生蛋白/Wnt信号通路的调控作用会受到多种因素的影响。生长因子及激素及机械应力等会在一定程度上影响骨形态发生蛋白/Wnt信号通路的活性。③骨形态发生蛋白/Wnt信号通路在调控成骨过程中与其他信号通路相互作用,共同构成复杂的调控网络。④中药和天然化合物可以通过调控信号通路来促进骨骼健康,为中药治疗骨骼疾病提供了新的可能性。⑤未来研究可进一步探讨骨形态发生蛋白/Wnt信号通路与其他信号通路的相互作用以及其在不同生理和病理条件下的变化,解析此复杂网络中的关键节点和调控机制,为骨骼相关疾病的治疗提供更精确的靶点,也为揭示骨骼形成和重塑的分子机制提供新的思路。

绝经后2型糖尿病LRP5基因多态性与骨量异常的研究

目的分析低密度脂蛋白受体相关蛋白5(low density lipoprotein receptor related protein 5,LRP5)rs556442、rs312778位点基因多态性及突变在绝经后女性2型糖尿病(type 2 diabetes mellitus,T2DM)患者中骨量异常的意义。方法收集2021年5月至2023年5月新疆石河子地区的142例绝经后女性资料进行回顾性分析,分为正常对照组(A组,n=29)、T2DM组(B组,n=30)、骨量降低组(C组,n=28)、骨量降低+T2DM组(D组,n=55)。收集记录相关基线资料,运用全自动生化测定仪测定受试者血糖、血脂及骨代谢指标。通过双能X线骨密度仪测定骨密度(bone mineral density BMD),LRP5基因位点多态性采用飞行时间质谱法(MALDI-TOF-MS)测定,采用SNPscan技术对上述SNP位点进行基因分型。结果①四组基线资料比较,绝经年限、年龄及腰臀比(waist hip ratio,WHR)比较差异具有统计学意义(P<0.05);②与A组相比,B组的空腹血糖(FPG)、糖化血红蛋白(HbA1c)升高(P<0.05);D组的FPG、HbA1c的血清学水平升高,甘油三酯(TG)血清学水平降低(P<0.05);③rs556442位点:与A组相比,D组基因型分布(AA/AG/GG基因型)有统计学意义(P<0.05);rs312778位点组间基因型(CC/CT/TT基因型)及等位基因分布频率均无统计学意义(P>0.05);④rs556442位点:与AA基因型相比,B、C组AG/GG基因型的股骨颈BMD水平降低;D组AG/GG型的HDL血清学水平降低(P<0.05)。rs312778位点:与CC基因型相比,A组HbA1C、FPG血清学水平较CT/TT基因型低(P<0.05);D组CT/TT基因型的低密度脂蛋白(LDL)血清学水平升高(P<0.05);⑤多元线性回归分析:rs556442位点,TG增加是腰L_(1~4)BMD降低的危险因素(P<0.05);rs312778位点,体质量指数(body mass index,BMI)降低是腰L_(1~4)及股骨颈BMD降低的危险因素,TG增加是腰L_(1~4)BMD降低的危险因素(P<0.05);⑥在rs556442、rs312778位点骨量异常与基因型分布均无统计学意义(P>0.05)。结论新疆石河子地区绝经后T2DM女性患者LRP5基因rs556442、rs312778基因位点的突变可能与骨量降低有关。

血清BMP及PPARγ水平与中轴型脊柱关节炎的骶髂关节脂肪化生的相关性研究

目的本研究旨在调查中轴型脊柱关节炎(axSpA)患者血清中骨形态发生蛋白(BMP)和过氧化物酶体增殖激活受体γ(PPARγ)的水平与脂肪化生的相关性,进而评估脂肪化生严重程度并预测早期结构进展。方法纳入30名axSpA病例和34名健康者作为对照组,采用Logistic回归分析同期临床、影像资料及血清学指标。结果病例组的血清BMP4,BMP9和PPARγ水平高于健康组,差异均具有统计学意义(均P<0.001)。axSpA患者的脂肪化生程度与血清BMP4(r=0.581,P<0.001)和PPARγ(r=0.507,P=0.004)均具呈正相关关系。重度脂肪化生组的血清BMP4(17.290±7.880 pg/mL)高于无、轻度脂肪化生组(9.131±4.087 pg/mL),差异具有统计学意义(P=0.002);PPARγ在重度脂肪化生组(104.561±49.764 pg/mL)亦高于无、轻度脂肪化生组(64.392±27.007 pg/mL),差异具有统计学意义(P=0.045)。BMP4水平的升高可能是axSpA患者重度脂肪化生的危险因素(OR=1.367,95%CI:[1.026,1.822],P=0.033)。结论血清BMP4和PPARγ水平有助于评估axSpA患者的脂肪化生程度。

过表达lncRNA HAGLR促胫骨骨折大鼠骨髓间充质干细胞成骨分化的机制研究

目的研究骨质疏松(OP)-胫骨骨折(TF)大鼠长链非编码RNA同源盒D基因簇反义生长相关长链非编码RNA(lncRNA HAGLR)与其下游靶基因的表达情况,并探讨lncRNA HAGLR对大鼠骨髓间充质干细胞(MSC)成骨分化的作用与机制。方法30只SD雌性大鼠随机分为sham组、OP组、OP-TF组,每组10只。ELISA法检测大鼠血清碱性磷酸酶(ALP)和抗酒石酸酸性磷酸酶(TRAP)的水平。对大鼠MSC细胞系R7500使用成骨分化诱导培养基进行诱导,并分为MSC组和成骨诱导组(Osteogenic-MSC组)。分别转染pcDNA-HAGLR、pcDNA-NC、miR-19a-3p的模拟物(miR-19a-3p mimic)、mimic的阴性对照(NC mimic)、miR-19a-3p的抑制剂(miR-19a-3p inhibitor)、miR-19a-3p inhibitor的阴性对照(NC inhibitor)至R7500后进行相应分组。双荧光素酶报告基因实验验证lncRNA HAGLR和miR-19a-3p以及骨形态发生蛋白2(BMP2)和miR-19a-3p的靶向关系。qRT-PCR检测各组lncRNA HAGLR和miR-19a-3p的表达。Western blot检测BMP2、ALP、胶原蛋白I(COL-I)、骨钙素(OCN)、骨桥蛋白(OPN)的表达。ALP染色和AR染色检测MSC的成骨分化能力。结果OP组和OP-TF组的血清ALP和TRAP水平均高于sham组,差异有统计学意义(P<0.05)。而OP组与sham组胫骨组织中lncRNA HAGLR、miR-19a-3p、BMP2的表达水平比较差异均无统计学意义(P>0.05),而OP-TF组胫骨组织中lncRNA HAGLR、BMP2的表达水平均明显低于sham组和OP组(P<0.05),OP-TF组胫骨组织中miR-19a-3p的表达水平高于sham组和OP组(P<0.05)。与MSC组相比,Osteogenic-MSC组的lncRNA HAGLR表达水平明显升高(P<0.05),而miR-19a-3p的表达降低(P<0.05)。双荧光素酶报告基因实验表明lncRNA HAGLR与miR-19a-3p具有靶向关系,miR-19a-3p与BMP2具有靶向关系。pcDNA-HAGLR组的miR-19a-3p的表达水平低于pcDNA-NC组(P<0.05)。miR-19a-3p mimic组与NC mimic组的lncRNA HAGLR的表达水平比较,差异无统计学意义(P>0.05)。与NC mimic组相比,miR-19a-3p mimic组BMP2的表达水平降低(P<0.05),miR-19a-3p表达水平升高(P<0.05)。pcDNA-HAGLR组细胞较pcDNA-NC组具有更强的成骨分化能力和更高的ALP活性(P<0.05)。miR-19a-3p inhibitor组细胞较NC inhibitor组具有更强的成骨分化能力和更高的ALP活性(P<0.05)。结论胫骨骨折大鼠lncRNA HAGLR和BMP2表达降低,miR-19a-3p表达增高。过表达lncRNA HAGLR通过靶向调控miR-19a-3p/BMP2轴促进大鼠MSC的成骨分化。