自体骨髓干细胞移植与归元方联用治疗急慢性肝损伤实验研究

研究中药归元方与自体骨髓干细胞移植对急慢性肝损伤的治疗作用.研究方法:用肝脏局部注射乙醇的方法复制急性局限性肝损伤模型,复合因素(CCl4、乙醇、高脂、低蛋白)刺激复制大鼠肝纤维化模型,通过定量组织学、肝功能检查、免疫组化、肝组织羟脯氨酸含量、损伤或纤维区骨髓干细胞观察等综合评价中药、自体骨髓干细胞移植及两者合用的疗效.结果:归元方与自体骨髓干细胞移植可减小肝损伤区域,改善肝功能,使纤维肝组织表达μPA增强,降低血清ALT,AST,PCⅢ,HA和肝组织羟脯氨酸的含量,改善肝组织肝纤维化评分,骨髓干细胞能在肝损伤、肝纤维化形成环境中存活、增殖,并向肝细胞分化,表达肝脏特异的角蛋白CK18.结论:归元方与自体骨髓干细胞移植对急慢性肝损伤有明确的治疗作用,两者合用可优势互补,协同增效.临床上有良好的应用前景.     

乳腺干细胞研究进展

最近的研究表明,利用干细胞抗原抗体从小鼠乳腺中分离得到的类乳腺干细胞具有干细胞的特性,体外诱导可以分化为乳腺中的各种细胞,在体内可以再生乳腺。小鼠的一个乳腺干细胞可以重新构建一个发育良好的、有功能的乳腺。因此,研究乳腺干细胞再生乳腺将对因乳腺肿瘤和癌症而切除乳腺的妇女再生功能性乳腺,为隆胸的女性提供安全可行的新材料,为健康家畜乳腺改造提供成熟的实验动物模型,为研究干细胞分化机理、再生器官的研究提供理论依据。本文综述了乳腺发生、发育,乳腺与乳腺干细胞的关系,乳腺干细胞的分类和生物学特性及其发育调控机理,有关人和啮齿类动物乳腺干细胞存在等方面的研究进展。尽管目前已经发现了许多关于血液干细胞及其子代细胞的标记性蛋白,但要找到乳腺干细胞的特异性标记性蛋白仍需花费很长的时间。     

大鼠骨髓间质干细胞的分离纯化与鉴定

采用密度梯度离心法分离骨髓间质干细胞,差速贴壁法进行纯化,应用流式细胞仪、激光共聚焦显微镜等对纯化细胞进行鉴定,结果细胞表面抗原CD２９,CD４４,CD１０５,CD１６６表达呈阳性,而CD１４,CD３４,CD４５表达呈阴性。采用RT唱PCR鉴定了三个基因：nestin,NST,Oct唱４,前两者呈阳性表达,后者弱阳性表达。这些细胞特异抗原与基因表达综合起来,表明得到的细胞具备骨髓间质干细胞的特性。密度梯度分离与差速贴壁相结合,可获得较好均一性的骨髓间质干细胞,是一种简单可靠、易于推广的骨髓干细胞获取方法,可为细胞与组织工程研究提供种子细胞。     

骨髓4℃保存方法及移植治疗晚期实体瘤的研究

对小鼠和人骨髓用TC—199营养液、复方氨基酸加ATP和生理盐水在4℃的保存效果进行了比较研究。用国产复方氨基酸加ATP保存的自体骨髓,在晚期实体瘤患者行强化疗后输回自身,有效率为75％。该方法安全、有效,且经济、方便,适合于基层医院推广应用。     

人体骨髓基质干细胞冷冻干燥的探索性实验

人体骨髓基质干细胞已成为组织工程、细胞移植和基因治疗等领域的重要研究对象,目前对它已有低温保存的研究,而通过冷冻干燥的方法对其进行保存国内外未见报导.实验尝试用冻干的方法来保存骨髓基质干细胞.文中选取海藻糖、PVP、HES等作保护剂,应用差示扫描量热仪(DSC)测量其结晶温度和玻璃化转变温度;随后对加入保护剂的细胞溶液进行冻干实验,并应用流式细胞仪对冻干样品复水后的细胞活性进行了测定,其中30%PVP 20%海藻糖对细胞的保护效果较好,细胞成活率达到16.40%.     

花鲈胚胎干细胞移植及嵌合体的构建

旨在通过花鲈胚胎干细胞(LJES1)移植构建嵌合体,证实LJES1细胞的体内分化能力和发育全能性.采用脂质体介导法将线性化pCMV-EGFP质粒导入到长期培养的LJES1细胞内,细胞绿色荧光蛋白(GFP)的表达率为5%～10%,以此为供体,通过显微注射方法把20～50个LJES1细胞移植到花鲈囊胚中,共移植囊胚478枚,获得了20枚表达GFP的嵌合体胚胎,其中的15枚胚胎发育成鱼苗.荧光显微观察和PCR检测显示了LJES1细胞可在宿主胚胎内片状嵌合和单细胞分散嵌合,嵌合部位可分布于胚体的三个胚层;同时也证明GFP是一种优良的遗传标记.本实验结果为证实LJES1细胞的发育全能性提供了有力证据.     

疾病危险度-共患病指数在单倍型造血干细胞移植中的应用

本文的研究目的是基于疾病危险度指数(disease risk index, DRI)和造血干细胞移植共患病指数(hematopoietic cell transplantation-specific comorbidity index, HCT-CI),提出适合单倍型造血干细胞移植(haploidentical hem...     

造血干细胞分离纯化的研究进展

造血干细胞分离纯化的研究进展王立生，吴祖泽，贺福初（军事医学科学院放射医学研究所北京１００８５０）早在１９３８年，Ｄｏｗｎｅｙ就提出造血组织中存在可生成各类血细胞的造血干细胞，但直到１９６１年Ｔｉｌｌ和ＭｃＣｕｌｌｏｃｈ［１］建立脾结节实验技术，造血...     

矿化柞蚕丝胶膜表面粗糙度的调控及其对骨髓间充质干细胞生长行为的影响

生物材料表面的粗糙度是影响细胞行为的重要因素之一。为了调控丝蛋白生物材料表面的粗糙度,并评价材料表面粗糙度对细胞生长行为的影响,首先,通过湿化学共沉淀法,以柞蚕丝胶(AS)溶液为模板,诱导了羟基磷灰石(HAp)晶体成核,进而调控了AS膜表面的粗糙度。然后,采用SEM、粗糙仪、FTIR及EDX等对HAp/AS复合膜表面形貌、粗糙度及成分进行了表征。最后,通过SEM和CellTiter 96?AQueous单溶液细胞增殖检测试剂盒(MTS)检测了骨髓间充质干细胞(BMSCs)在HAp/AS复合膜表面的形貌及增殖率。结果表明:纯AS膜的表面粗糙度为0.15μm,矿化1、8及24h后,表面粗糙度分别为0.38、0.46和1.20μm;矿化24h后,在HAp/AS复合膜表面可观察到直径为30~80nm的球状复合物,生成的矿化物为HAp;HAp/AS复合膜具有良好的细胞相容性,表面粗糙度为1.20μm的复合膜能够显著促进BMSCs的增殖,粗糙度对BMSCs在HAp/AS复合膜表面的粘附和形貌有着重要的影响。因此,可通过矿化的方法在生物大分子表面诱导HAp晶体的成核与生长,从而调控材料的表面粗糙度,研究材料界面上的细胞行为。     

骨髓,胎肝和外周血造血干细胞的低温生物效应

用生物降温仪、低温显微镜和细胞化学、细胞培养等技术研究证明,低温保存液的相变温度随降温速率的加快而增高,以1℃/min降温时,其相交点在—13.9℃,骨髓细胞浓度对相变温度无影响。液氮(—196℃)冻存后,外用血淋巴细胞体积无明显变化,其糖原和非特异性酯酶含量明显降低。胎肝细胞冻存1098天染色体正常,冻存930天后氧耗量与新鲜值无明显差别,冻存7天后超微弱发光强度为新鲜值的92％。小鼠骨髓和皮肤冻存后,移植免疫排斥反应降低。人骨髓冻存3～300天,胎肝冻存1～1257天,外周血冻存6～53天,其CFU—GM回收率分别为68.9％、62.0％和77.3％。造血细胞活力不随保存时间延长而降低。     

体外壳聚糖-明胶-生长因子缓释支架对骨髓间充质干细胞增殖的影响

制备具有缓释功能的壳聚糖-明胶-碱性成纤维细胞生长因子(bFGF)-肝细胞生长因子(HGF)三维大孔支架,探讨两种生长因子在体外对骨髓间充质干细胞(BMSCs)增殖的协同作用。方法:采用冷冻干燥法,将不同比例的壳聚糖、明胶、bFGF和HGF依次混合,使其成为具有一定孔径的三维缓释支架。取SD大鼠乳鼠股骨和胫骨骨髓,分离、培养BMSCs。取生长状态良好的第三代BMSCs,将其接种于96孔板后,加入支架混合培养,5d后进行MTT细胞增殖测定。结果:在与含1μg/mL的bFGF支架混合培养,以及与同时含1μg/mL的bFGF、1μg/mL的HGF支架混合培养后,BMSCs增殖明显(P<0.05),但这二组间无统计学差异(P>0.05)。分别与含0.1μg/mL的bFGF支架、0.01μg/mL的bFGF支架、1μg/mL的HGF支架混合培养后,细胞增殖无统计学意义(P>0.05)。结论:壳聚糖-明胶可作为生长因子缓释支架材料;bFGF具有促进BMSCs增殖的作用,促进作用的大小与加入bFGF的量有关;HGF对BMSCs不具有增殖作用;在实验浓度范围内bFGF和HGF体外促进BMSCs增殖上不具有协同性。     

Deconstructed Microfluidic Bone Marrow On‐A‐Chip to Study Normal and Malignant Hemopoietic Cell–Niche Interactions

Human hematopoietic niches are complex specialized microenvironments that maintain and regulate hematopoietic stem and progenitor cells (HSPC). Thus far, most of the studies performed investigating alterations of HSPC‐niche dynamic interactions are conducted in animal models. Herein, organ microengineering with microfluidics is combined to develop a human bone marrow (BM)‐on‐a‐chip with an integrated recirculating perfusion system that consolidates a variety of important parameters such as 3D architecture, cell–cell/cell–matrix interactions, and circulation, allowing a better mimicry of in vivo conditions. The complex BM environment is deconvoluted to 4 major distinct, but integrated, tissue‐engineered 3D niche constructs housed within a single, closed, recirculating microfluidic device system, and equipped with cell tracking technology. It is shown that this technology successfully enables the identification and quantification of preferential interactions—homing and retention—of circulating normal and malignant HSPC with distinct niches.     

Nanosensors for Continuous and Noninvasive Monitoring of Mesenchymal Stem Cell Osteogenic Differentiation

Assessing mesenchymal stem cell (MSC) differentiation status is crucial to verify therapeutic efficacy and optimize treatment procedures. Currently, this involves destructive methods including antibody‐based protein detection and polymerase chain reaction gene analysis, or laborious and technically challenging genetic reporters. Development of noninvasive methods for real‐time differentiation status assessment can greatly benefit MSC‐based therapies. This report introduces a nanoparticle‐based sensing platform that encapsulates two molecular beacon (MB) probes within the same biodegradable polymeric nanoparticles. One MB targets housekeeping gene glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) as an internal reference, while another detects alkaline phosphatase (ALP), a functional biomarker. Following internalization, MBs are gradually released as the nanoparticle degrades. GAPDH MBs provide a stable reference signal throughout the monitoring period (18 days) regardless of differentiation induction. Meanwhile, ALP mRNA undergoes well‐defined dynamics with peak expression observed during early stages of osteogenic differentiation. By normalizing ALP‐MB signal with GAPDH‐MB, changes in ALP expression can be monitored, to noninvasively validate osteogenic differentiation. As proof‐of‐concept, a dual‐colored nanosensor is applied to validate MSC osteogenesis on 2D culture and polycaprolactone films containing osteo‐inductive tricalcium phospate.     

Personalized Hydrogels for Engineering Diverse Fully Autologous Tissue Implants

Despite incremental improvements in the field of tissue engineering, no technology is currently available for producing completely autologous implants where both the cells and the scaffolding material are generated from the patient, and thus do not provoke an immune response that may lead to implant rejection. Here, a new approach is introduced to efficiently engineer any tissue type, which its differentiation cues are known, from one small tissue biopsy. Pieces of omental tissues are extracted from patients and, while the cells are reprogrammed to become induced pluripotent stem cells, the extracellular matrix is processed into an immunologically matching, thermoresponsive hydrogel. Efficient cell differentiation within a large 3D hydrogel is reported, and, as a proof of concept, the generation of functional cardiac, cortical, spinal cord, and adipogenic tissue implants is demonstrated. This versatile bioengineering approach may assist to regenerate any tissue and organ with a minimal risk for immune rejection.     

Applications of Nanomaterials in Cell Stem Therapies and the Onset of Nanomedicine

Stem cells hold enormous potential in the treatment of diseases such as diabetes, arthritis, cirrhosis, spinal cord injury, and Alzheimer's disease, due to their unique ability to differentiate into various cell lines and tissues and integrate seamlessly into damaged or diseased tissue. The use of nanoparticles as bioactive molecules is still considered a nascent science, but their unique physical and chemical properties hold great hopes for drug delivery, cancer targeting, and bioimaging. There is active worldwide ongoing research to generate advanced therapeutic compounds for incurable diseases, combining the unique properties of nanomaterials and stem cells. The present review will cover emerging areas of nanotechnology applications in stem cell therapy, one of the next frontiers of medical science.