细胞表面壳化的研究进展

细胞表面壳化主要是通过物理、化学等技术方法对细胞表面进行修饰,形成完整均匀的有机、无机、金属纳米粒子或者复合壳层结构,从而使不能自身壳化的生物细胞表面形成保护壳甚至赋予细胞新的功能,使细胞具备多功能性。近年来,此技术在细胞存储、细胞运输、细胞传感器、细胞芯片以及细胞治疗等方面应用广泛,发展迅速。本文综合目前的研究现状,详细介绍了可进行细胞表面壳化的细胞类型、生物表面壳化的方法以及人造细胞外壳的工程技术在生物医学以及能源环境中的应用等。

Gastrointestinal hormone abnormalities and G and D cells in functional dyspepsia patients with gastric dysmotility

AIM: To investigate the relationship between gastric dysmotility,gastrointestinal hormone abnormalities, and neuroendocrine cells in gastrointestinal mucosa in patients with functional dyspepsia (FD).METHODS: Gastric emptying was assessed with solid radiopaque markers in 54 FD patients, and the patients were divided into two groups according to the results, one with delayed gastric emptying and the other with normal gastric emptying. Seventeen healthy volunteers acted as normal controls. Fasting and postprandial plasma levels and gastroduodenal mucosal levels of gastrointestinal hormones gastrin, somatostatin (SS) and neurotensin (NT)were measured by radioimmunoassay in all the subjects.G cells (gastrin-producing cells) and D cells (SS-producing cells) in gastric antral mucosa were immunostained with rabbit anti-gastrin polyclonal antibody and rabbit anti-SS polyclonal antibody, respectively, and analyzed quantitatively by computerized image analysis.RESULTS: The postprandial plasma gastrin levels, the fasting and postprandial plasma levels and the gastric and duodenal mucosal levels of NT were significantly higher in the FD patients with delayed gastric emptying than in those with normal gastric emptying and normal controls. The number and gray value of G and D cells and the G cell/D cell number ratio did not differ significantly between normal controls and the FD patients with or without delayed gastric emptying.CONCLUSION: Our findings suggest that the abnormalities of gastrin and NT may play a role in the pathophysiology of gastric dysmotility in FD patients, and the abnormality of postprandial plasma gastrin levels in FD patients with delayed gastric emptying is not related to the changes both in the number and gray value of G cells and in the G cell/D cell number ratio in gastric antral mucosa.

Connective tissue growth factor hammerhead ribozyme attenuates human hepatic stellate cell function

AIM： To determine the effect of hammerhead ribozyme targeting connective tissue growth factor （CCN2） on human hepatic stellate cell （HSC） function.METHODS： CCN2 hammerhead ribozyme cDNA plus two self-cleaving sequences were inserted into pTriEx2 to produce pTriCCN2-Rz. Each vector was individually transfected into cultured LX-2 human HSCs, which were then stimulated by addition of transforming growth factor （TGF）-β1 to the culture medium. Semiquantitative RT-PCR was used to determine mRNA levels for CCN2 or collagen I, while protein levels of each molecule in cell /ysates and conditioned medium were measured by ELISA. Cell-cycle progression of the transfected cells was assessed by flow cytometry.RESULTS： In pTriEx2-transfected LX-2 cells, TGF-β1 treatment caused an increase in the mRNA level for CCN2 or collagen I, and an increase in produced and secreted CCN2 or extracellular collagen I protein levels, pTriCCN2-Rz-transfected LX-2 cells showed decreased basal CCN2 or collagen mRNA levels, as well as produced and secreted CCN2 or collagen I protein. Furthermore, the TGF-β1-induced increase in mRNA or protein for CCN2 or collagen I was inhibited partially in pTriCCN2-Rz-transfected LX-2 cells. Inhibition of CCN2 using hammerhead ribozyme cDNA resulted in fewer of the cells transitioning into S phase.CONCLUSION： Endogenous CCN2 is a mediator of basal or TGF-β1-induced collagen I production in human HSCs and regulates entry of the cells into S phase.

Influence of heme oxygenase-1 gene transfer on the viability and function of rat islets in in vitro culture

AIM. To investigate the influence of heme oxygenase-1 （HO-1） gene transfer on the viability and function of cultured rat islets in vitro. METHODS： Islets were isolated from the pancreata of Sprague-Dawley rats by intraductal collagenase digestion, and purified by discontinuous Ficoll density gradient centrifugation. Purified rat islets were transfected with adenoviral vectors containing human HO-1 gene （Ad- HO-1） or enhanced green fluorescent protein gene （Ad- EGFP）, and then cultured for seven days. Transfection was confirmed by fluorescence microscopy and Western blot. Islet viability was evaluated by acridine orange/ propidium iodide fluorescent staining. Glucose-stimulated insulin release was detected using insulin radioimmunoassay kits and was used to assess the function of islets. Stimulation index （SI） was calculated by dividing the insulin release upon high glucose stimulation by the insulin release upon low glucose stimulation. RESULTS： After seven days culture, the viability of cultured rat islets decreased significantly （92% ± 6% vs 52% ± 13%, P 〈 0.05）, and glucose-stimulated insulin release also decreased significantly （6.47 ± 0.55 mIU/ L/30IEO vs 4.57 ± 0.40 mIU/L/3OIEO., 14.93 ± 1.17 mIU/L/30IEQ vs 9.63 ± 0.71 mIU/L/30IEQ, P 〈 0.05）. Transfection of rat islets with adenoviral vectors at an 1±10 of 20 was efficient, and did not impair islet function. At 7 d post-transfection, the viability of Ad-HO-1 transfected islets was higher than that of control islets（71% ± 15% vs 52% ± 13%, P 〈 0.05）. There was no significant difference in insulin release upon low glucose stimulation （2.8 mmol/L） among Ad-HO-1 transfected group, Ad-EGFP transfected group, and control group （P 〉 0.05）, while when stimulated by high glucose （16.7 mmol/L） solution, insulin release in Ad-HO-1 transfected group was significantly higher than that in Ad-EGFP transfected group and control group, respectively （12.50 ±2.17 mIU/L/30IEQ vs 8.87 ± 0.65 mIU/L/30IEQ, 12.50 ± 2.17 mIU/L/30IEQ vs 9.63 ± 0.71 mIU/L/30IEQ, P 〈 0.05）. The SI of Ad-HO-1 transfected group was also significantly higher than that of Ad-EGFP transfected group and control group, respectively （2.21 ± 0.02 vs 2.08 ± 0.05; 2.21 ± 0.02 vs 2.11 ± 0.03, P 〈 0.05）. CONCLUSION： The viability and function of rat islets decrease over time in in vitro culture, and heine oxygenase-1 gene transfer could improve the viability and function of cultured rat islets.

Small inhibitors of ADP-ribosylation factor activation and function in mammalian cells

Small GTP-binding proteins of the ADP-ribosylation fac-tor （Arf） family control various cell functional responses including protein transport and recycling between different cellular compartments, phagocytosis, prolif-eration, cytoskeletal remodelling, and migration. The activity of Arfs is tightly regulated. GTPase-activating proteins （GAPs） inactivate Arfs by stimulating GTP hydrolysis, and guanine nucleotide exchange factors （GEFs） stimulate the conversion of inactive GDP-bound Arf to the active GTP-bound conformation. There is increasing evidence that Arf small GTPases contribute to cancer growth and invasion. Increased expression of Arf6 and of Arf-GEPs, or deregulation Arf-GAP functions have been correlated with enhanced invasive capacity of tumor cells and metastasis. The spatiotemporal spe-cifcity of Arf activation is dictated by their GEFs that integrate various signals in stimulated cells. Brefeldin A （BFA）, which inactivates a subset of Arf-GEFs, has been very useful for assessing the function of Golgi-localized Arfs. However, specifc inhibitors to investigate the individual function of BFA-sensitive and insensitive Arf-GEFs are lacking. In recent years, specifc screens have been developed, and new inhibitors with improved se-lectivity and potency to study cell functional responses regulated by BFA-sensitive and BFA-insensitive Arf pathways have been identified. These inhibitors have been instrumental for our understanding of the spa-tiotemporal activation of Arf proteins in cells and dem-onstrate the feasibility of developing small molecules interfering with Arf activation to prevent tumor invasion and metastasis.

The X-linked mental retardation gene PHF8 is a histone demethylase involved in neuronal differentiation

Recent studies have identified mutations in PHF8, an X-linked gene encoding a JmjC domain-containing protein, as a causal factor for X-linked mental retardation （XLMR） and cleft lip/cleft palate. However, the underlying mechanism is unknown. Here we show that PHF8 is a histone demethylase and coactivator for retinoic acid receptor （RAR）. Although activities for both H3K4me3/2/1 and H3K9me2/1 demethylation were detected in cellularbased assays, reeombinant PHF8 exhibited only H3K9me2/1 demethylase activity in vitro, suggesting that PHF8 is an H3K9me2/1 demethylase whose specificity may be modulated in vivo. Importantly, a mutant PHF8 （phenylalanine at position 279 to serine） identified in the XLMR patients is defective in enzymatie activity, indicating that the loss of histone demethylase activity is causally linked with the onset of disease. In addition, we show that PHF8 binds specifically to H3K4me3/2 peptides via an N-terminal PHD finger domain. Consistent with a role for PHF8 in neuronal differentiation, knockdown of PHF8 in mouse embryonic carcinoma P19 cells impairs RA-induced neuronal differentiation, whereas overexpression of the wild-type but not the F279S mutant PHF8 drives PI9 cells toward neuronal differentiation. Furthermore, we show that PHF8 interacts with RAR~ and functions as a coactivator for RARa. Taken together, our results suggest that histone methylation modulated by PHF8 plays a critical role in neuronal differentiation.

The influence of neoadjuvant chemotherapy on immunity function in elderly patients with the stages of Ⅱ and Ⅲ esophageal cancer

Objective: We aimed to study the influence of neoadjuvant chemotherapy on immunity function in elderly patients with the stages of II and III esophageal cancer. Methods: Thirty-seven elderly patients (age ranged from 60 to 75 years) with the stages of II and III esophageal cancer underwent 2 cycles chemotherapy preoperatively with single-drug regimen (docetaxel, 35 mg/m2 once a week, on days 1, 8 and 15, at interval of 2 weeks for one cycle). Surgery were performed three weeks later. Blood samples were drawn separately on the day of admission, 1 day before operation, 7 day and 1 month after operation, and we conducted the Flow Cytometry to detect the levels of CD3+, CD4+, CD8+,CD4+/CD8+ and NK cells. Results: There were no significant differences in the levels of CD3+, CD4+, CD8+, CD4+/CD8+ and NK cells between before and after chemotherapy (P > 0.05). On day 7 after operation, the levels of CD3+, CD4+, CD4+/CD8+ and NK cells were degraded and CD8+ increased significantly (P < 0.05). One month after operation, the levels of CD3, CD4+, CD4+/CD8 and NK cells were higher than normal, and CD8 was depressed significantly (P < 0.05). Conclusion: Neoadjuvant chemotherapy has no significant impact on cellular immune function in elderly patients with the stages of II and III esophageal cancer, it is an effective and safe treatment.

Structure and Function of the Digestive System of Solen Grandis Dunker

Structure and function of the digestive system of a bivalve mollusc, Solen grandis, were studied using light microscopy and histochemical methods. The wall of digestive tube consists of four layers: the mucosal epithelium, connective tissue, muscular and fibrosa or serosa (only in the portion of rectum) from the inner to the outer. The ciliated columnar epithelial cells, dispersed by cup-shaped mucous cells, rest on a thin base membrane. There are abundant blood spaces in connective tissue layer. The digestive diverticula are composed of multi-branched duct and digestive tubules. The digestive tubules are lined with digestive and basophilic secretory cells, and surrounded by a layer of smooth muscle fibers and connective tissues. Activities of acid and alkaline phosphatases, esterase and lipase are detected in the digestive cells, and the epithelia of stomach and intestine, suggesting that these cells are capable of intracellular digesting of food materials and absorbing.Besides, acid phosphatase and esterase activities are present in the posterior portion of esophagus. Phagocytes are abundant in blood spaces and the lumens of stomach and intestine, containing brown granules derived from the engulfed food materials. The present work indicates that phagocytes play important roles in ingestion and digestion of food materials, which is supported as well by the activities of acid phosphatase, esterase and lipase detected in blood spaces.

Concomitant chemoradiation using gemcitabine in locally advanced hepatocellular carcinoma

Objective:Hepatocellular carcinoma (HCC) is among the most common and rapidly increasing cancers in Pakistan.There is currently no standard management for advanced HCC.The aim of the study was to assess response rate and toxicity of concomitant gemcitabine and external radiation therapy (ERT) in locally advanced HCC.Methods:Sixty patients were enrolled.Gemcitabine,70 mg/m2 was given weekly during ERT.ERT was delivered with 60Co beam up to 30.6 Gy,1.8 Gy/fraction.Tumor response was assessed by computed tomography (CT) at eight weeks.Complete and partial response (CR and PR),progressive and stable disease (PD and SD) were assessed.Hematological,gastrointestinal and hepatic toxicities were assessed weekly.Results:No CR was seen.PR,SD and PD were seen in 33%,40% and 27% of patients respectively.Grade 3/4 toxicity for total leukocyte count and platelets was observed in 7% and 27% patients.Grade 3/4 toxicity for liver failure,bilirubin,aspartate aminotransferase,alanine aminotransferase and alkaline phosphatase was seen in 35%,28%,38%,24% and 43% patients respectively.Conclusion:The study showed that concomitant gemcitabine and ERT is a feasible option with moderate toxicity in advanced HCC.

Stem cells modified by brain-derived neurotrophic factor to promote stem cells differentiation into neurons and enhance neuromotor function after brain injury

Objective： To promote stem cells differentiation into neurons and enhance neuromotor function after brain injury through brain-derived neurotrophic factor （BDNF） induction. Methods： Recombinant adenovirus vector was applied to the transfection of BDNF into human-derived umbilical cord mesenchymal stem cells （UCMSCs）. Enzyme linked immunosorbent assay （ELISA） was used to determine the secretion phase of BDNF. The brain injury model of athymic mice induced by hydraulic pressure percussion was established for transplantation of stem cells into the edge of injury site. Nerve function scores were obtained, and the expression level of transfected and non-transfected BDNF, proportion of neuron specific enolase （NSE） and glial fibrillary acidic protein （GFAP）, and the number of apoptosis cells were compared respectively. Results： The BDNF expression achieved its stabilization at a high level 72 hours after gene transfection. The mouse obtained a better score of nerve function, and the proportion of the NSE-positive cells increased significantly （P〈0.05）, but GFAP-positive cells decreased in BDNF- UCMSCs group compared with the other two groups （P〈0.05）. At the site of high expression of BDNF, the number of apoptosis cells decreased markedly. Conclusion： BDNF gene can promote the differentiation of the stem cells into neurons rather than glial cells, and enhance neuromotor function after brain injury.

Electrochemically and DNA-triggered cell release from ferrocene/β-cyclodextrin and aptamer modified dual- functionalized graphene substrate

Here we report a dual-functionalized electrochemical substrate to trigger cancer cells release based on the supramolecular interaction between β-cyclodextrin （β-CD） and Fc on clinical trial II aptamer AS1411 functionalized graphene platform. On one hand, the host-guest interaction can be reversible electrochemically controlled to realize cancer cells capture/release, and 1-adamantylamine binding can further amplify this surface change by competing interaction with β-CD. On the other hand, the AS1411 aptamer and its complementary DNA （cDNA） also can be used as a switchable anchor for cell adhesion. Our work gives an example for label-free, multi-functionalized triggered cell release based on aptamer and β-CD/graphene-modified surface and this multi-ways for cell catch-and-release on graphene modified surface also provides their potential biomedical application.

MicroRNAs in tumor immunity:functional regulation in tumor-associated macrophages

Tumor-associated macrophages(TAMs)are the most abundant immune cells in the tumor microenvironment(TME)and are critical for cancer initiation and progression.MicroRNAs(miRNAs)could notably influence the phenotype of TAMs through various targets and signal pathways during cancer progression due to their posttranscriptional regulation.In this review,we discuss mainly the regulatory function of miRNAs on macrophage differentiation,functional polarization,and cellular crosstalk.Firstly,during the generation process,miRNAs take part in the differentiation from myeloid cells to mature macrophages,and this maturation process directly influences their recruitment into the TME,attracted by tumor cells.Secondly,macrophages in the TME can be either tumor-promoting or tumor-suppressing,depending on their functional polarization.Large numbers of miRNAs can influence the polarization of macrophages,which is crucial for tumor progression,including tumor cell invasion,intravasation,extravasation,and premetastatic site formation.Thirdly,crosstalk between tumor cells and macrophages is essential for TME formation and tumor progression,and miRNAs can be the mediator of communication in different forms,especially when encapsulated in microvesicles or exosomes.We also assess the potential value of certain macrophage-related miRNAs(MRMs)as diagnostic and prognostic markers,and discuss the possible development of MRM-based therapies.

Enhanced in vitro anticancer activity of quercetin mediated by functionalized CdTe QDs

We report in this study the effects of red-emitting CdTe QDs capped with cysteamine(Cys-CdTe) on the in vitro anticancer activity of the well-known flavenoid quercetin(Qu). Various techniques, including the methylthiazolyldiphenyl-tetrazolium bromide assay, the real-time cell electronic sensing system, the optical and fluorescence imaging, and electrochemical methods have been utilized to study the potential interactions of Cys-CdTe QDs with Qu. The observations demonstrate that the safe-dosage Cys-CdTe QDs can greatly improve the drug uptake and enhance the inhibition efficiency of Qu towards the proliferation of cancer cells such as HepG2 cells. This study implies that Cys-CdTe QDs may be used for cancer therapy and that they exert a synergic anticancer effect when bound to drug molecules.

Effects of the Aidi Dripping Pills on Immune Functions of the Tumor-bearing Mouse

Objective： To study the effects of Aidi Dripping Pills on immune functions of the tumor-bearing mouse on the basis of the previous experimental studies on its tumor-inhibiting and life-prolonging effects. Methods： By using the transplantation tumor mouse models, the effects of Aidi Dripping Pills on the lymphocyte transformation rate and the hemolysin formation in the Sis0 tumor-bearing mice, and on the phagocytic function of macrophages in the abdominal cavity of H22 tumor-bearing mice were investigated. Results： In the 2.25 g/kg and 1.125 g/kg Aidi Dripping Pills groups, the lymphocyte transformation rates in the Sis0 tumor-bearing mice were significantly higher than that of the control group （P〈0.01）. In all the Aidi Dripping Pills groups, HC50 significantly increased （P〈0.01 or P〈0.05）, carbon granular clearance significantly raised, and both the phagocytic index and phagocytic coefficient were significantly higher than those in the control group （P〈0.01 or P〈0.05）. Conclusion： The Aidi Dripping Pills can significantly increase the cellular immune function, the humoral immune function and the phagocytic function of the mononuclear- macrphages, so it may show anti-tumor effects by enhancing the function of the reticuloendothelial system.

STIM1 and Orai1:novel targets for vascular diseases?

The past five years have witnessed the discovery of the endoplasmic reticulum calcium(Ca2+) sensor STIM1 and the plasma membrane Ca2+channel Orai1 as the bona fide molecular components of the store-operated Ca2+ entry(SOCE) and the Ca2+ release-activated Ca2+current(I CRAC) .It has been known for two decades that SOCE and ICRAC are required for lymphocyte activation as evidenced by severe immunodeficient phenotypes in patients lacking ICRAC.In recent years however,studies have uncovered expression of STIM1 and Orai1 proteins in various tissues and described additional roles for these proteins in physiological functions and pathophysiological conditions.Here,we will summarize novel findings pertaining to the role of STIM1 and Orai1 in the vascular system and discuss their potential use as targets in the therapy of vascular disease.

Post-transcriptional gene regulation by RNA-binding proteins in vascular endothelial dysfunction

Endothelial cell dysfunction is a term which implies the dysregulation of normal endothelial cell functions,including impairment of the barrier functions,control of vascular tone,disturbance of proliferative and migratory capacity of endothelial cells,as well as control of leukocyte trafficking.Endothelial dysfunction is an early step in vascular inflammatory diseases such as atherosclerosis,diabetic vascular complications,sepsis-induced or severe virus infection-induced organ injuries.The expressions of inflammatory cytokines and vascular adhesion molecules induced by various stimuli,such as modified lipids,smoking,advanced glycation end products and bacteria toxin,significantly contribute to the development of endothelial dysfunction.The transcriptional regulation of inflammatory cytokines and vascular adhesion molecules has been well-studied.However,the regulation of those gene expressions at post-transcriptional level is emerging.RNA-binding proteins have emerged as critical regulators of gene expression acting predominantly at the post-transcriptional level in microRNA-dependent or independent manners.This review summarizes the latest insights into the roles of RNA-binding proteins in controlling vascular endothelial cell functions and their contribution to the pathogenesis of vascular inflammatory diseases.

Targeting stem cell signaling pathways for drug discovery:advances in the Notch and Wnt pathways

Signaling pathways transduce extracellular stimuli into cells through molecular cascades to regulate cellular functions.In stem cells,a small number of pathways,notably those of TGF-?/BMP,Hedgehog,Notch,and Wnt,are responsible for the regulation of pluripotency and differentiation.During embryonic development,these pathways govern cell fate specifications as well as the formation of tissues and organs.In adulthood,their normal functions are important for tissue homeostasis and regeneration,whereas aberrations result in diseases,such as cancer and degenerative disorders.In complex biological systems,stem cell signaling pathways work in concert as a network and exhibit crosstalk,such as the negative crosstalk between Wnt and Notch.Over the past decade,genetic and genomic studies have identified a number of potential drug targets that are involved in stem cell signaling pathways.Indeed,discovery of new targets and drugs for these pathways has become one of the most active areas in both the research community and pharmaceutical industry.Remarkable progress has been made and several promising drug candidates have entered into clinical trials.This review focuses on recent advances in the discovery of novel drugs which target the Notch and Wnt pathways.

Membrane nanotubes:Novel communication between distant cells

The many kinds of cell structures involved in cell-cell communication include tight junction,adherens junction and gap junction,but almost all are between adjacent cells.Recently,a general and dynamic membrane tether,termed tunneling nanotubes or membrane nanotubes(MNTs),was discovered to be involved in communication between distant cells.By facilitating intercellular communication,MNTs contribute to many biological functions and pathologic changes in cells.Many works have revealed the structure,formation and functional properties of MNTs.However,as novel structures,further research is needed.

Aqueous self-assembly and surface-functionalized nanodots for live cell imaging and labeling

Nanoparticles have enormous potential for bioimaging and biolabeling applications, in which conventional organically based fluorescent labels degrade and fail to provide long-term tracking. Thus, the development of approaches to make fluorescent probes water soluble and label cells efficient is desirable for most biological applications. Here, we report on the fabrication and charac- terization of self-assembled nanodots （SANDs） from 3-aminopropyltriethoxysilane （APTES） as a probe for protein labeling. We show that fluorescent SAND probes exhibit both bright photoluminescence and biocompatibility in an aqueous environment. Selective in vitro imaging using protein and carbohydrate labeling of hepatoma cell lines are demonstrated using biocompatible SANDs conjugated with avidin and galactose, respectively. Cytotoxicity tests show that conjugated SAND particles have negligible effects on cell proliferation. Unlike other synthetic systems that require multistep treatments to achieve robust surface functionalization and to develop flexible bioconjugation strategies, our results demonstrate the versatility of this one-step SAND fabrication method for creating multicolor fluorescent probes with the tailored functionalities, effident emission, as well as excellent biocompatibility, required for broad biological use.

Oligonucleotide delivery by chitosan-functionalized porous silicon nanoparticles

Porous silicon nanoparficles （pSiNPs） are a promising nanocarrier system for drug delivery owing to their biocompatibility, biodegradability, and non-inflammatory nature. Here, we investigate the fabrication and characterization of thermally hydrocarbonized pSiNPs （THCpSiNPs） and chitosan-coated THCpSiNPs for therapeutic oligonucleotide delivery. Chitosan coating after oligonucleotide loading significantly improves sustained oligonucleotide release and suppresses burst release effects. Moreover, cellular uptake, endocytosis, and cytotoxicity of oligonucleotide-loaded THCpSiNPs have been evaluated in vitro. Standard cell viability assays demonstrate that cells incubated with the NPs at a concentration of 0.1 mg/mL are 95% viable. In addition, chitosan coating significantly enhances the uptake of oligonucleotide-loaded THCpSiNPs across the cell membrane. Moreover, histopathological analysis of liver, kidney, spleen, and skin tissue collected from mice receiving NPs further demonstrates the biocompatible and non-inflammatory properties of the NPs as a gene delivery vehicle for intravenous and subcutaneous administration in vivo. Taken together, these results suggest that THCpSiNPs provide a versatile platform that could be used as efficient vehicles for the intracellular delivery of oligonucleotides for gene therapy.