One Step Quick Detection of Cancer Cell Surface Marker by Integrated NiFe-based Magnetic Biosensing Cell Cultural Chip

RGD peptides has been used to detect cell surface integrin and direct clinical effective therapeutic drug selection. Herein we report that a quick one step detection of cell surface marker that was realized by a specially designed NiF e-based magnetic biosensing cell chip combined with functionalized magnetic nanoparticles. Magnetic nanoparticles with 20-30 nm in diameter were prepared by coprecipitation and modified with RGD-4C, and the resultant RGD-functionalized magnetic nanoparticles were used for targeting cancer cells cultured on the NiF e-based magnetic biosensing chip and distinguish the amount of cell surface receptor-integrin.Cell lines such as Calu3, Hela, A549, CaF br, HEK293 and HUVEC exhibiting different integrin expression were chosen as test samples. Calu3, Hela, HEK293 and HUVEC cells were successfully identified. This approach has advantages in the qualitative screening test. Compared with traditional method, it is fast, sensitive, low cost,easy-operative, and needs very little human intervention. The novel method has great potential in applications such as fast clinical cell surface marker detection, and diagnosis of early cancer, and can be easily extended to other biomedical applications based on molecular recognition.

Current status and challenges for cell-cultured milk technology: a systematic review

Cellular agriculture is an innovative technology for manufacturing sustainable agricultural products as an alternative to traditional agriculture.While most cellular agriculture is predominantly centered on the production of cultured meat,there is a growing demand for an understanding of the production techniques involved in dairy products within cellular agriculture.This review focuses on the current status of cellular agriculture in the dairy sector and technical challenges for cell-cultured milk production.Cellular agriculture technology in the dairy sector has been classified into fermentation-based and animal cell culture-based cellular agriculture.Currently,various companies synthesize milk components through precision fermentation technology.Nevertheless,several startup companies are pursuing animal cell-based technology,driven by public concerns regarding genetically modified organisms in precision fermentation technology.Hence,this review offers an up-to-date exploration of animal cell-based cellular agriculture to produce milk components,specifically emphasizing the structural,functional,and productive aspects of mammary epithelial cells,providing new information for industry and academia.

MACS-W:A modified optical clearing agent for imaging 3D cell cultures

Three-dimensional(3D)cell cultures have contributed to a variety of biological research fields by filling the gap between monolayers and animal models.The modern optical sectioning microscopic methods make it possible to probe the complexity of 3D cell cultures but are limited by the inherent opaqueness.While tissue optical clearing methods have emerged as powerful tools for investigating whole-mount tissues in 3D,they often have limitations,such as being too harsh for fragile 3D cell cultures,requiring complex handling protocols,or inducing tissue deformation with shrinkage or expansion.To address this issue,we proposed a modified optical clearing method for 3D cell cultures,called MACS-W,which is simple,highly efficient,and morphology-preserving.In our evaluation of MACS-W,we found that it exhibits excellent clearing capability in just 10 min,with minimal deformation,and helps drug evaluation on tumor spheroids.In summary,MACS-W is a fast,minimally-deformative and fluorescence compatible clearing method that has the potential to be widely used in the studies of 3D cell cultures.

Bioengineering breakthroughs:The impact of stem cell models on advanced therapy medicinal product development

The burgeoning field of bioengineering has witnessed significant strides due to the advent of stem cell models,particularly in their application in advanced therapy medicinal products(ATMPs).In this review,we examine the multifaceted impact of these developments,emphasizing the potential of stem cell models to enhance the sophistication of ATMPs and to offer alternatives to animal testing.Stem cell-derived tissues are particularly promising because they can reshape the preclinical landscape by providing more physiologically relevant and ethically sound platforms for drug screening and disease modelling.We also discuss the critical challenges of reproducibility and accuracy in measurements to ensure the integrity and utility of stem cell models in research and application.Moreover,this review highlights the imperative of stem cell models to align with regulatory standards,ensuring using stem cells in ATMPs translates into safe and effective clinical therapies.With regulatory approval serving as a gateway to clinical adoption,the collaborative efforts between scientists and regulators are vital for the progression of stem cell applications from bench to bedside.We advocate for a balanced approach that nurtures innovation within the framework of rigorous validation and regulatory compliance,ensuring that stem cell-base solutions are maximized to promote public trust and patient health in ATMPs.

3D Collagen Gels:A Promising Platform for Dendritic Cell Culture in Biomaterials Research

The three-dimensional(3D)cell culture system has garnered significant attention in recent years as a means of studying cell behavior and tissue development,as opposed to traditional two-dimensional cultures.These systems can induce specific cell reactions,promote specific tissue functions,and serve as valuable tools for research in tissue engineering,regenerative medicine,and drug discovery.This paper discusses current developments in the field of three-dimensional cell culture and the potential applications of 3D type 1 collagen gels to enhance the growth and maturation of dendritic cells.

Biotransformation of Gastrodin by Cell Suspension Cultures of Catharanthus roseus

应用长春花 (Catharanthusroseus (L .)G .Don)悬浮细胞培养体系对天麻素进行了生物转化反应研究。经过8d培养形成一个转化产物 ,应用光谱方法鉴定转化产物的结构为对羟基苯甲醇 ,为天麻素水解后形成的甙元。

Productions of Taxol and Related Taxanes by Cell Suspension Cultures of Taxus yunnanensis

A high taxol yield cell line of Taxus yunnanensis Cheng et L. K. Fu keeps a high taxol_producing level after successive subcultures for more than eight years. In this study, eight taxanes were isolated from the suspension cell cultures of this cell line. Based on NMR and MS analyses, and comparison with literature data and standards, their structures were determined to be 2α,5α,10β_triacetoxy_14β_propionyloxy_4(20),11_taxadiene (1), 2α,5α,10β_triacetoxy_14β_(2′_methyl)_butyryloxy_4(20),11_taxadiene (2), 2α,5α,10β_14β_tetra_acetoxy_4 (20),11_taxadiene (3, taxuyunnanine C), 2α,5α,10β_triacetoxy_14β_(2′_methyl_3′_hydroxy)_butyryloxy_4(20),11_taxadiene (4, yunnanxane) and its 3′_epimer (5), baccatin Ⅳ (6), baccatin Ⅲ (7) and taxol (8), respectively. Among those compounds, 3, 5, 6 and 7 were reported to be isolated from the suspension cell cultures of T. yunnanensis for the first time. TLC and HPLC analyses indicated that the chemical constituents of the culture solution were similar to those of cultured cells. Moreover, the highest taxol content of this cell line reached 0.3% and the cell line could be applied for a large_scale culture.

Chemical Constituents of the Suspension Cell Cultures of Maytenus hookeri

Suspension cell cultures of Maytenus hookeri Loos. (Celastraceae) in SH media were established from the calli induced from the leaves and young steins of M. hookeri on MS media with the supplement of 2 mg/L 2,4-D and 0.1 mg/L KIN (kinetin). Ethyl acetate extract of the cultures showed inhibitory activities against Penicillium avellaneum UC-4376 which was sensitive to maytansinoids. Exhaustive isolation of natural products from a large scale of suspension cell cultures did not yield maytansine instead of affording nine compounds including one novel triterpenoid, named 2, 3-diacetoxyl maytenusone (1), and eight known ones including squalene (2), beta-sitosterol (3), 2', 3', 4-triacetyl-sitoindoside I (4), salaspermic acid (5), maytenonic acid (6), 2alpha-hydroxy-maytenonic acid (7), 6, 11,12-trihydroxy-8, 11, 13-abietrien-7-one (8) and 11, 12-dihydroxy-8, 11, 13-abietatrien-7-one (9) elucidated on the basis of 1D and 2D NMR data. The H-1-NMR and C-13-NMR assignments were made for 1, 5, 6 and 7, while the C-13-NMR assignments for 5 and 6 were revised. The chemical results suggested that the suspension cell cultures of M. hookeri did not produce maytansinoids under the reported experiment conditions.

Cell Growth and Ajmalicine Accumulation in a Full Habituated Catharanthus roseus Cell Line C_ (20)hi

A full habituated cell line C_ 20hi was screened from 2,4_D dependent line (C_ 20D) of Catharanthus roseus (L.) G. Don. The investigation involved the cell growth, ajmalicine production and enzyme activity related to indole alkaloid biosynthesis in both cell lines. These results indicated that C_ 20hi cells grew faster than C_ 20D cells, and average ajmalicine content in C_ 20hi cells was 18.4 times more than that in C_ 20D when cultured in the production medium. In the growth medium, average ajmalicine content in C_ 20hi cells was 31.9 times more than that in C_ 20D cells, while the cell growth has no obvious difference. The comparison of enzyme activities in C_ 20hi and C_ 20D cells indicated that tryptophan decarboxylase (TDC), strictosidine synthase (SSS) and geraniol_10_dehydrogenase (G10H) activities have no close relation to ajmalicine accumulation, although the activities of these enzymes were higher when cells were cultured in the production medium than in the growth medium. The C_ 20hi cells are relatively stable in five years of culture.

Single Polar Compound Bioelectret Material and Its Influence on the Cell Growth

To explore the influence of compound bioelectret material′s dielectric property on the cell growth,several kinds of compound bioelectret materials of collagen/chitosan were developed Their TSDC(thermally stimulated depolarization current)spectra were analyzed,and the compound bioelectret collagen/chitosan whose t α and I α were 37℃ and 2×10 -9 A respectively at polarized state was selected The cell culture study showed that the compound bioelectret material could promote normal cell growth when singly negatively polarized,and could inhibit cancer cell growth when singly positively polarized It proves that the rational designation of compound bioelectret has a broad application for clinical medicine

Study on the Browning in Cell Suspension Culture of Taxus cuspidata

[Objective] This study aimed to investigate the browning of T. cuspidata cells in suspension culture and provide the guidance for the cell suspension culture of T. cuspidata. [Method] T. cuspidata callus was used as experimental materials, to explore the effect of different medium, N/P ratio, pH, shaking speed, illumination time and light intensity and other factors on browning of T. cuspidata cells in suspension culture. [Result] Non-browning callus was transferred to 2MB5 medium （pH 7.0） for illumination culture at 22℃ under light intensity of 1 500 lx with shaking speed of 90 r/min for 24 h. Results showed that the cell browning was significantly inhibited. [Conclusion] This study laid the foundation for cell suspension culture of T. cuspidata and had important significance to the large-scale industrial production of paclitaxel.

Morphological Characteristics of Smooth Muscle Cells Isolated from the Rat Ductus Deferens

The aim of this study was to establish a method of isolating and culturing smooth muscle cells from the ductus deferens of rats. Smooth muscle cells were prepared from ductus deferens by explanting technique after dissection of adventitia and intimae, and cultured in vitro. The identification of the smooth muscle cells were verified by using anti u-smooth muscle actin （a-SMA） immunohistochemistry studies. The result suggested that the cells are multi-morphous, showing long fusiform or star shapes. The apophysis of cells contacted and coalesced to each other, in some regions the cells overlapped in multilayer, while in the other regions they formed monolayer that fluctuated and showed a ＂peak-valley＂ shape. They presented a positive reaction through immunohistochemistry studies. The purity of the cells was more than 99% through this method. The culturing of smooth muscle cells by explanting technique is simple and stable.

MORPHOLOGICAL MODEL FOR RHEOLOGICAL PROPERTIES OF PLANT CELL SUSPENSION CULTURE SYSTEM

This paper puts forward a physical and mathematical model for the rheological properties of a plant cell suspension culture system.The model can explain why the system is pseudoplastic satisfactorily,thus the rheological properties of the system as the effect of the flow behavior index on plant cell concentration are interpreted correctly and the mechanism of the rheological properties of the system is further understood.Therefore the model can be applied in the technological design and optimum conditions of the system and the reformation,evaluation and scale up of reactors.

Detection of Mycoplasma Contamination in Animal Cell Cultures

[ Objective ] To develop a rapid efficient method for detecting mycoplasma contamination in cell cultures. [ Method] A pair of primers was designed according to two highly conserved nucleotide sequences of the 16S RNA from six kinds of mycoplasma that commonly contaminated cells. Then the mycoplasma contamination of 25 cell samples was defected by PCR and DNA fluorescence staining. EResultl When these cell samples were detected by DNA fluorescence staining, the positive rate and probable positive rate were respectively 24% and 16%. And when they were detected by PCR, the positive rate was 36%. [ Condusion] The PCR method is more sensitive and specific than the DNA fluorescence staining, and combining these two methods is the optimal way to detect mycoplasma contamination in cell cultures.

Selective 3-OH Isomerization of Resibufogenin by Cell Suspension Cultures of Ginkgo biloba

Aim To modify the structure of resibufogenin by using Ginkgo bilobasuspension. Methods Young leaves of Ginkgo biloba were differentiated into callus in MS medium withonly 2,4-D as plant growth regulator. The callus was then transferred aseptically to liquid MSmedium exoge-nously supplemented with appropriate concentration of 6-BA, NAA and 2,4-D to establishsuspension cell culture system. Resibufogenin was administered into the well-grown cell cultures andincubated for 4 d. The products dissolved in the liquid phase of the cultures were extracted andpurified by silica gel column chromatography gradiently eluted with petroleum ether and acetonesystem. Results One transformed product was obtained in 40% yield after 4 d incubation, which wasidentified as 3-epi-resibufogenin on the basis of FAB MS, ~1H NMR and ^(13)C NMR spectroscopicanalysis and corresponding data reported in literature. Conclusion G. biloba suspension cultures canbe used as an enzyme system to biotransform resibufogenin, an animal-originated bufadienolide, into3-epi-resibufogenin.

Influence of Bioelectret Materials on cell Culture

The aim of this work is to study the effects of the electret property of natural biopolymers (such as collagen) on tissue repair. Type I collagen was prepared from pigskin,and polarized by using the TSDC (thermally stimulated depolarization current) technique. The polarized materials are used for cell culture. and then the cell growth curve is drawn. It was found that the polarized biomaterials accelerated cell differentiation. which indicates that they may be applied in the biomedical field.

Effect of norcantharidin on proliferation and invasion of human gallbladder carcinoma GBC-SD cells

AIM: To investigate the effect of norcantharidin on proliferation and invasion of human gallbladder carcinoma GBC-SD cells in vitro and its anticancer mechanism. METHODS: Human gallbladder carcinoma GBC-SD cells were cultured by cell culture technique. The growth and the invasiveness of GBC-SD cells in vitro were evaluated by the tetrazolium-based colorimetric assay and by the Matrigel experiment and the crossing-river test. Expression of PCNA, Ki-67, MMP2 and TIMP2 proteins of GBC-SD cells was determined by streptavidin-biotin complex method. RESULTS: In vitro norcantharidin inhibited the growth and proliferation of GBC-SD cells in a dose- and time-dependent manner, with the IC50 value of 56.18 μ/mL at 48 h. Norcantharidin began to inhibit the invasion of GBC-SD cells at the concentration of 5 μg/mL, and the invasive action of GBC-SD cells was inhibited completely and their crossing-river time was prolonged significantly at 40 μg/mL. After treatment with norcantharidin, the expression of PCNA, Ki-67, and MMP2 was significantly decreased. With the increase in TIMP2 expression, the MMP2 to TIMP2 ratio was decreased significantly (P<0.05). CONCLUSION: Norcantharidin inhibits the proliferation and growth of human gallbladder carcinoma cells in vitro at relatively low concentrations by inhibiting PCNA and Ki-67 expression. Its anti-invasive activity may be the result of decrease in MMP2 to TIMP2 ratio and reduced cell motility.

Effects of ursolic acid and oleanolic acid on human colon carcinoma cell line HCT15

AIM: Ursolic acid (UA) and oleanolic acid (OA) are triperpene acids having a similar chemical structure and are distributed wildly in plants all over the world. In recent years, it was found that they had marked anti-tumor effects. There is little literature currently available regarding their effects on colon carcinoma cells. The present study was designed to investigate their inhibitory effects on human colon carcinoma cell line HCT15. METHODS: HCT15 cells were cultured with different drugs. The treated cells were stained with hematoxylin-eosin and their morphologic changes observed under a light microscope. The cytotoxicity of these drugs was evaluated by tetrazolium dye assay. Cell cycle analysis was performed by flow cytometry (FCM). Data were expressed as means +/-SEM and Analysis of variance and Student' t-test for individual comparisons. RESULTS: Twenty-four to 72 h after UA or OA 60 micromol/L treatment, the numbers of dead cells and cell fragments were increased and most cells were dead at the 72nd hour. The cytotoxicity of UA was stronger than that of OA. Seventy-eight hours after 30 micromol/L of UA or OA treatment, a number of cells were degenerated, but cell fragments were rarely seen. The IC(50) values for UA and OA were 30 and 60 micromol/L, respectively. Proliferation assay showed that proliferation of UA and OA-treated cells was slightly increased at 24h and significantly decreased at 48 h and 60 h, whereas untreated control cells maintained an exponential growth curve. Cell cycle analysis by FCM showed HCT15 cells treated with UA 30 and OA 60 for 36 h and 72 h gradually accumulated in G(0)/G(1) phase (both drugs P【0.05 for 72 h), with a concomitant decrease of cell populations in S phase (both drugs P【0.01 for 72 h) and no detectable apoptotic fraction. CONCLUSION: UA and OA have significant anti-tumor activity. The effect of UA is stronger than that of OA. The possible mechanism of action is that both drugs have an inhibitory effect on tumor cell proliferation through cell-cycle arrest.

Culture and Identification of Human Amniotic Mesenchymal Stem Cells

Objective To establish the method of isolation, purification, and identification of human amniotic mesenchymal stem cells (hAMSCs). Methods hAMSCs were isolated from human amniotic membrane by trypsin-collagenase digestion, and cultured in Dulbecco's modified Eagle's medinm/F12 medium supplemented with 10% fetal bovine serum. Phenotypic characteristics of these cells were analyzed by means of immunocytochemistry and flow cytometry. Results The cells successfully isolated from human amniotic membrane expressed representative mesenchymal cell surface markers CD44, CD90, and vimentin, but not CD45. Conclusions This study establishes a potential method for isolation of hAMSCs from human amnion, in vitro culture, and identification. The isolated cells show phenotypic characteristics of mesenchymal stem cells.

Protective Effects of Quercetin on Cadmium-induced Cytotoxicity in Primary Cultures of Rat Proximal Tubular Cells

Objective To investigate the protective effects of quercetin on cadmium-induced cytotoxicity in primary cultures of rat proximal tubular （rPT） cells. Methods Primary cultures of rPT cells undergoing exponential growth were incubated with 1.0 ug/mL quercetin and/or cadmium （2.5, 5.0 umol/L）, in a serum-free medium at 37℃ at different time intervals. Commercial kits were used and flow cytometric analyses were performed on rPT cell cultures to assay apoptosis and oxidative stress. Results Exposure of rPT cells to cadmium acetate （2.5, 5.0 umol/L） induced a decrease in cell viability, caused an increase in apoptotic rate and apoptotic morphological changes. Simultaneously, elevation of intracellular reactive oxygen species, malondialdehyde and calcium levels, depletion of mitochondrial membrane potential and intracellular glutathione, and inhibition of Na＋, K＋ -ATPase, Ca2＋ -ATPase, glutathione peroxidase （GSH-Px）, catalase （CAT）, and superoxide dismutase （SOD） activities were revealed during the cadmium exposure of rPT cells. However, simultaneous supplementation with 1 ug/mL quercetin protected rPT cells against cadmium-induced cytotoxicity through inhibiting apoptosis, attenuating lipid peroxidation, renewing mitochondrial function and elevating the intracellular antioxidants （non-enzymatic and enzymic） levels. Conclusion The present study has suggested that quercetin, as a widely distributed dietary antioxidant, contributes potentially to prevent cadmium-induced cytotoxicity in rPT cells.