神经干细胞和神经细胞的拉曼光谱分析

从光谱学角度研究神经干细胞（NSCs）与神经细胞（NCs）,是一个新的干细胞研究方法,能够为临床应用提供一定的实验参考依据。本文通过体外培养NSCs和NCs,应用激光共焦显微拉曼光谱仪测定两种细胞的拉曼光谱。实验结果显示,NSCs的拉曼峰646 cm-1,2917 cm-1与NCs的拉曼峰652 cm-1,2908 cm-1处存在较大差异。NCs核中B型DNA胸腺嘧啶和酪氨酸含量相对更丰,而NSCs核中具有NO2成分而NCs核中较少或没有。NSCs比NCs的DNA、酪氨酸、蛋氨酸、腺嘌呤、苯基丙氨酸、胡萝卜素、酰胺Ⅲ等含量更丰,其中以胡萝卜素和酰胺Ⅲ的相对含量最高。本文从光谱学角度分析NSCs与NCs的光谱区别,以为临床应用提供一定的实验参考依据。     

恶性骨肿瘤患者红细胞的拉曼光谱研究

通过对正常人和恶性骨肿瘤患者红细胞的拉曼光谱进行了研究.发现对于恶性骨肿瘤患者,部分谱线如1001.2,1228.8,1398.7,1372.8cm-1红移了2～4cm-1,说明这些谱线对应的基团内部构象发生微小变化.以1586cm-1为标记峰得到各峰的相对峰强比,发现恶性骨肿瘤患者红细胞的拉曼峰相对于正常人的相对强度的比值在1505,1228,1087,1001和747cm-1处显著减小,说明亚铁血红素、苯丙氨酸和脱氧核糖在红细胞内的含量减少.由统计分析可知Ⅰ1586/1228,Ⅰ1586/1001和Ⅰ1586/1169的峰-峰比的变化可为恶性骨肿瘤的早期诊断提供一定的依据.     

EC9706细胞经不同剂量X射线辐照后的拉曼光谱

为了研究不同剂量X射线对食管癌(EC9706)细胞的辐射损伤,经不同剂量X射线照射后继续培养24 h,利用拉曼光谱分析其内部蛋白质、核酸、脂类等构象和含量变化.实验结果表明,各剂量组拉曼光谱的峰强和频移与空白对照组之间存在较大的差异.对照组光谱中某些谱带在个别剂量组消失,如蛋白质主链中C-N振动1114 cm-1谱带在各剂量组中消失,脱氧核糖Dr振动谱带895 cm-1仅存在于8Gy组,937 cm-1谱带仅存在于12Gy组,膜脂中膜结合β-胡萝卜素的C=C振动谱带1523 cm-1仅存在于6Gy组.这些特征谱带在不同剂量组光谱中的变化,为进一步研究X射线辐照EC9706细胞的最佳剂量提供了一定参考.     

单个肝癌细胞的拉曼成像研究

应用拉曼成像技术检测单个人体肝癌细胞,获得了单个肝癌细胞微区的拉曼光谱图谱,同时计算出786,1450和1658 cm-1等特征峰的峰面积,这些特征峰分别归属于DNA,脂类和蛋白质,根据归一化后的数值在相应的细胞扫描位置成像,重构出这些物质的拉曼特征峰在肝癌细胞中的分布图.结果表明,应用这种方法可以很明确地看到DNA,...     

活体糖尿病小鼠中单个白细胞的拉曼光谱分析

利用拉曼光镊并结合多元统计分析方法无损地对活体糖尿病(DM)小鼠中的白细胞(WBC)进行了研究,分别检测了糖尿病和正常小鼠活体中的白细胞拉曼光谱,利用主成分分析(PCA)建立拉曼光谱诊断多元统计算法模型。结果表明,患糖尿病小鼠和正常小鼠白细胞拉曼光谱差别明显,且实验存在较好的重现性。利用PCA统计分析方法得到诊断特异性和灵敏度达到了98%。在活体糖尿病小鼠白细胞中出现较高的蛋白质的特征峰1302cm-1,表明活体糖尿病白细胞中的蛋白质浓度比正常状态高。糖尿病小鼠内的白细胞与正常相比,DNA磷酸骨架基团强度和蛋白质酰胺强度升高,表明DNA双螺旋结构、蛋白质主链和氢键体系发生变化,二级构象改变。     

单个肝癌细胞的拉曼光谱分析研究

本工作利用激光镊子拉曼光谱系统研究了人正常肝细胞株(LO2)和肝癌细胞株(SMMC-7721)的单细胞拉曼光谱,对于每个细胞在不同部位测三个点.实验结果显示:正常细胞和癌细胞的平均拉曼光谱存在显著差异;癌细胞谱线强度整体变弱;正常细胞的1658cm-1处峰和1450cm-1处峰的强度比值为0.63,癌细胞的为0.99;癌细胞的核酸、蛋白质、脂类等重要生物分子在结构或含量上都发生了不同的改变.用PCA主成分对单个细胞的平均拉曼光谱进行分析,结果发现PCA可以正确区分出正常细胞和癌细胞.这些结果表明,激光镊子拉曼光谱是判断正常肝细胞和肝癌细胞的有效方法.     

飞秒激光辐照ZnO晶体

利用飞秒激光对ZnO晶体进行辐照,对辐照前后的晶体样品进行发光光谱及拉曼光谱检测.辐照后发光光谱的某些发光峰强度有明显增强,但未产生新的发光峰,表明没有新的缺陷结构产生,但晶体内锌空位、间隙位锌、间隙位缺陷浓度增加.拉曼光谱结果表明,辐照后ZnO晶体未产生相变,但随着辐照激光功率的增大,拉曼峰327 cm-1,437 cm-1强度明显减弱,表明在飞秒激光辐照作用下氧化锌的结晶程度下降.但574 cm-1峰值却随着辐照功率的增大而变大,分析表明该拉曼峰很可能是由于晶体内间隙位缺陷所致.同时实验过程中观察到飞秒激光倍频光产生.     

甲状腺癌组织与正常组织红外光谱的研究

应用傅立叶变换红外光谱法( FTlR)对甲状腺癌组织及正常组织进行了分析。实验结 果表明:正常组织红外光谱分别位于1165cm- 1和1745cm- 1附近的吸收峰较为明显,而在癌组织中近乎消失;癌组织中970cm- 1附近的吸收峰强度较正常组织在相对强度上有所增加;正常组织中1084cm- 1附近的吸收峰在癌组织中向低波数移动3cm- 1且强度增加;甲状腺癌组织中酰胺Ⅰ带、Ⅱ带的吸收峰强度高于正常组织,酰胺Ⅱ带吸收峰向低波数移动2cm- 1 ,酰胺Ⅰ带、Ⅱ带的相对峰高比( I1654 / I1543 )高于正常组织。研究表明: FTIR可以从分子水平上揭示甲状腺正常组织和癌组织的特异性,能够对甲状腺肿瘤组织的良、恶性鉴别提供可靠的信息。     

碳源浓度影响微生物PHB合成代谢的单细胞拉曼光谱分析

应用激光镊子拉曼光谱实时监测杀虫贪铜菌(Cupriavidus necator)H16菌株在特定氮源、不同果糖浓度下生物塑料聚β-羟基丁酸(PHB)的合成过程及其生化动态,以期获知碳源浓度对PHB合成的影响机制。结果显示:1)2.0%果糖浓度(质量分数)下的PHB产量获得最佳发酵效果;2)发酵启动及发酵前期胞内物质的合成速度基本相同,但果糖浓度影响对数生长期的长短;3)782、1574、1656和1732 cm-1等峰信号强度变化动态显示,高碳源浓度下核酸物质的拉曼信号长时间维持在较高的水平,这可能是影响细胞合成PHB的重要因素;4)单个细胞在1732 cm-1峰处的平均强度与发酵液PHB总含量线性相关,可用于定量监测发酵过程中PHB总量动态。发现了C.necator H16菌株在不同碳源浓度下PHB合成代谢过程中胞内主要生物大分子的变化动态,为PHB发酵提供全新的光谱学信息,也为基于单细胞拉曼光谱快速分选高产优良菌株提供实验依据。     

天然与处理翡翠的光谱学研究

文章通过对天然翡翠和漂白充填处理翡翠的共焦显微拉曼光谱(514nm)测定,得到表征翡翠硬玉结构的拉曼位移为1036cm-1,696cm-1和374cm-1,显微放大能观察到翡翠经酸洗产生的裂纹及其充填的有机物,经红外光谱的测定证实该充填物为环氧树脂,且表征环氧树脂的拉曼峰为777cm-1,1123cm-1,1611cm-1,2930cm-1和3065cm-1;通过相同测试条件下,并在极短的积分时间内对天然翡翠的绿色、白色部位和染绿色翡翠进行拉曼光谱(785nm)测定,并结合绿色翡翠紫外-可见光-近红外吸收光谱的测定,得到吸收光谱中的吸收峰637nm,658nm和691nm归属于翡翠晶体中的Cr3 ,Cr3 的大量存在将使翡翠的拉曼光谱产生极强的荧光,而染色翡翠中染料产生的荧光相对较弱.     

Arrayed rGOSH/PMASH Microcapsule Platform Integrating Surface Topography,Chemical Cues,and Electrical Stimulation for Three‐Dimensional Neuron‐Like Cell Growth and Neurite Sprouting

The biocompatible thiol‐functionalized rGO_SH/PMA_SH microcapsules encapsulating nerve growth factor (NGF) are arrayed onto a transparent and conductive substrate, i.e., indium tin oxide (ITO), to integrate electrically stimulated cellular differentiation, electrically controlled NGF release, and topographically rough nano‐surfaces into a 3‐D platform for nerve regeneration. The rGO_SH/PMA_SH microcapsules with microscale topography function not only as an adhesive coating to promote the adhesion of PC12 cells but also as electroactive NGF‐releasing electrodes that stimulate NGF release and accelerate the differentiation of PC12 cells during electrical stimulation. Once electrical treatment is applied, NGF release and electrically enhanced cellular differentiation lead to an obvious increase both in the percentage of cells with neurites and in the neurite length. This length can reach nearly 90 μm within 2 days of cell culture. The average neurite length is significantly increased (four‐fold) after culture on the rGO_SH/PMA_SH microcapsule substrate for 2 days compared with culture on a substrate without an rGO_SH/PMA_SH coating. These multifunctional rGO_SH/PMA_SH microcapsules may be used as potential 3‐D patterned substrates for neural regeneration and neural prosthetics in tissue engineering applications.     

Hyaluronic Acid Catechol: A Biopolymer Exhibiting a pH‐Dependent Adhesive or Cohesive Property for Human Neural Stem Cell Engineering

Nature has developed materials that are integrated and effective at controlling their properties of adhesiveness and cohesiveness; the chemistry of these materials has been optimized during evolution. For example, a catechol moiety found in the adhesive proteins of marine mussels regulates its properties between adhesion and cohesion, rapidly adapting to environmental conditions. However, in synthetic materials chemistry, introduced chemical moieties are usually monofunctional, either being adhesive or cohesive; typically, this is not effective compared to natural materials. Herein, it is demonstrated that hyaluronic acid‐catechol (HA‐catechol) conjugates can exhibit either adhesiveness, functionalizing the surface of materials, or cohesiveness, building 3D hydrogels. Up to now, catechol‐conjugated polymers have shown to be useful in one of these two functions. The usefulness of the polymer in stem cell engineering is demonstrated. A platform for neural stem cell culture may be prepared, utilizing the adhesive property of HA‐catechol, and hydrogels are fabricated to encapsulate the neural stem cells, utilizing the cohesive property of the HA conjugate. Moreover, the HA‐catechol hydrogels are highly neural stem cell compatible, showing better viability compared to existing methods based on HA hydrogels.     

3D Printed Stem‐Cell Derived Neural Progenitors Generate Spinal Cord Scaffolds

A bioengineered spinal cord is fabricated via extrusion‐based multimaterial 3D bioprinting, in which clusters of induced pluripotent stem cell (iPSC)‐derived spinal neuronal progenitor cells (sNPCs) and oligodendrocyte progenitor cells (OPCs) are placed in precise positions within 3D printed biocompatible scaffolds during assembly. The location of a cluster of cells, of a single type or multiple types, is controlled using a point‐dispensing printing method with a 200 µm center‐to‐center spacing within 150 µm wide channels. The bioprinted sNPCs differentiate and extend axons throughout microscale scaffold channels, and the activity of these neuronal networks is confirmed by physiological spontaneous calcium flux studies. Successful bioprinting of OPCs in combination with sNPCs demonstrates a multicellular neural tissue engineering approach, where the ability to direct the patterning and combination of transplanted neuronal and glial cells can be beneficial in rebuilding functional axonal connections across areas of central nervous system (CNS) tissue damage. This platform can be used to prepare novel biomimetic, hydrogel‐based scaffolds modeling complex CNS tissue architecture in vitro and harnessed to develop new clinical approaches to treat neurological diseases, including spinal cord injury.     

A Selective,Hydrogel-Based Prodrug Delivery System Efficiently Activates a Suicide Gene to Remove Undifferentiated Human Stem Cells Within Neural Grafts

The directed differentiation of human pluripotent stem cells (hPSCs) into defined populations has advanced regenerative medicine, especially for Parkinson's disease where clinical trials are underway. Despite this, tumorigenic risks associated with incompletely patterned and/or quiescent proliferative cells within grafts remain. Addressing this, donor stem cells carrying the suicide gene, thymidine kinase (activated by the prodrug ganciclovir, GCV), are employed to enable the programmed ablation of proliferative cells within neural grafts. However, coinciding the short half-life of GCV with the short S-phase of neural progenitors is a key challenge. To overcome this, a smart hydrogel delivery matrix is fabricatedto prolong GCV presentation. Following matrix embedment, GCV retains its functionality, demonstrated by ablation of hPSCs and proliferating neural progenitors in vitro. A prolonged GCV release is measured by mass spectrometry following the injection of a GCV-functionalized hydrogel into mouse brains. Compared to suboptimal, daily systemic GCV injections, the intracerebral delivery of the functionalized hydrogel, as a “one-off treatment”, reduce proliferative cells in both hPSC-derived teratomas and neural grafts, without affecting the graft's functional unit (i.e., neurons). It is demonstrated that a functionalized biomaterial can enhance prodrug delivery and address safety concerns associated with the use of hPSCs for brain repair.     

Nanoscale Optoregulation of Neural Stem Cell Differentiation by Intracellular Alteration of Redox Balance

Regulation of stem cell (SC) fate, a decision between self‐renewal and differentiation, is of immense importance in regenerative medicine and has been proven to be a powerful stimulus regulating many cell functions influencing the SC fate. This study uses triphenylphosphonium‐functionalized gold nanoparticles (TPP‐AuNPs) to explore the interplay of intracellular electromagnetic (EM) exposure and the SC fate. Localized EM waves are generated inside neural stem cells (NSCs) to stimulate TPP‐AuNPs (AuNPs), targeting the mitochondria through inducing reactive oxygen species and differentiating these cells into neurons. Following laser irradiation of TPP‐AuNPs‐transfected NSCs, their differentiation to neurons is monitored by tracing the relevant markers both at the genetic and protein levels. The electrophysiology technique is further used to examine the functionality of neurons. The results confirm that TPP‐AuNPs subjected to electromotive forces have the potential to regulate cellular fate, although further investigations are still required to shed light on the mechanisms underlying the interaction of EM‐stimulated TPP‐AuNPs on cellular fate to design highly adjustable cell differentiation and reprogramming methods.     

Multifunctional Hydroxyapatite Nanobelt Haystacks Integrated Neural Stem Cell Spheroid for Rapid Spinal Cord Injury Repair

Due to the unwarranted lifespan and differentiation, applying neural stem cells (NSCs) in spinal cord injury (SCI) remains challenging. In this study, 3D bioactive hydroxyapatite (HAp) nanobelt haystack-mouse NSC (mNSC) hybrid spheroids are customized in which the specific nanobelt haystack framework provided the structural function of hypoxia alleviation in the spherical core and biological process of neural differentiation promotion. Commodified with superparamagnetic ferroferric oxide (Fe₃O₄) nanoparticles and a polydopamine (PDA) coating, the HAp nanobelts are endowed with magnetic field-driven properties and enhanced cell-nanobelt adhesion. The engineered bioresponsive 3D nanobelt haystack-mNSC hybrid spheroids effectively repair SCI in vivo, showing new potential for stem cell therapy by incorporating nanomaterials in 3D culture based on cell-material interactions.     

Surface Topography and Electrical Signaling: Single and Synergistic Effects on Neural Differentiation of Stem Cells

Incomplete regeneration and restoration of function in damaged nerves is a major clinical challenge. In this regard, stem cells hold much promise in nerve tissue engineering, with advantages such as prevention of scar‐tissue ingrowth and guidance of axonal regrowth. Engineering 3D and patterned microenvironments using biomaterials with chemical and mechanical characteristics close to those of normal nervous tissue has enabled new approaches for guided differentiation of various stem cells toward neural cells and possible treatment of neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's diseases. Differentiation of stem cells in a neurogenic lineage is largely affected by signals from the surrounding microenvironment (niche). The stem cell niche refers to a specific microenvironment around the stem cells, which provides specific biochemical (soluble factors) and biophysical signals (topography, electrical, and mechanical). This specified niche regulates the stem cells' behavior and fate. While the role of chemical cues in neural differentiation is well appreciated, recently, the cues presented by the physical microenvironment are increasingly documented to be important regulators of nerve cell differentiation. The single and synergistic effects of surface topography and electrical signals on neural differentiation of stem cells are reviewed.