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.

The Role of Progesterone in Elf5 Activation and Milk Component Synthesis for Cell-Cultured Milk Production in MAC-T Cells.

Prolactin is essential for mammary gland development and lactation. Progesterone also induces ductal branching and alveolar formation via initial secretory differentiation within the mammary gland. Herein, we aimed to evaluate the role of progesterone as a prolactin substitute for the production of cell-cultured milk components in MAC-T cells. Cells were treated with various hormones such as prolactin (PRL), progesterone (P4), 17ß-estradiol (E2), cortisol (COR), and insulin (INS) for 5 d. MAC-T cells cultured in a P4 differentiation media (2500 ng/mL of P4, 25 ng/mL of E2, 25 ng/mL of COR, and 25 ng/mL of INS) showed similar levels of E74-like factor 5 (Elf5) and milk component synthesis (α-casein, ß-casein, α-lactalbumin, ß-lactoglobulin, and triglycerides) compared to those cultured in a PRL differentiation media (5000 ng/mL of PRL, 500 ng/mL of CORT, and 50 ng/mL of INS). The levels of α-casein and triglycerides in the optimal P4 differentiation media were present at comparable levels to those in the PRL differentiation media. Our results demonstrated that P4 induces the activation of Elf5 and the synthesis of milk components in MAC-T cells, similar to PRL. Therefore, P4 may be used as an effective substitute of PRL for cell-cultured milk production in in vitro frameworks.

In vitro culture of bovine fibroblasts using select serum-free media supplemented with Chlorella vulgaris extract.

Standard cell culture practices require the addition of animal-derived serum to culture media to achieve adequate cell growth. Typically, 5-10% by volume of fetal bovine serum (FBS) is used, which accounts for a vast majority of the media cost while also imposing environmental and ethical concerns associated with the use of animal serum. Here we tested the efficacy of culturing cells by replacing serum in the media with algae extract and select additives. Using LC-MS, we compared molecular signatures of FBS to Chlorella algae extracts and identified NAD(H)/NADP(H) as common and relatively abundant features in their characteristic profiles. Bovine fibroblasts, cultured in serum-free media supplemented with C. vulgaris extract and just two growth factors plus insulin, showed significant growth with enhanced viability compared to control cells cultured without serum, albeit still lower than that of controls cultured with 10% FBS. Moreover, C. vulgaris extract enhanced cell viability beyond that of cells cultured with the two growth factors and insulin alone. These results suggest that key components in serum which are essential for cell growth may also be present in C. vulgaris extract, demonstrating that it may be used at least as a partial alternative to serum for cell culture applications.

Bioprocessing by Decellularized Scaffold Biomaterials in Cultured Meat: A Review.

As novel carrier biomaterials, decellularized scaffolds have promising potential in the development of cellular agriculture and edible cell-cultured meat applications. Decellularized scaffold biomaterials have characteristics of high biocompatibility, bio-degradation, biological safety and various bioactivities, which could potentially compensate for the shortcomings of synthetic bio-scaffold materials. They can provide suitable microstructure and mechanical support for cell adhesion, differentiation and proliferation. To our best knowledge, the preparation and application of plant and animal decellularized scaffolds have not been summarized. Herein, a comprehensive presentation of the principles, preparation methods and application progress of animal-derived and plant-derived decellularized scaffolds has been reported in detail. Additionally, their application in the culture of skeletal muscle, fat and connective tissue, which constitute the main components of edible cultured meat, have also been generally discussed. We also illustrate the potential applications and prospects of decellularized scaffold materials in future foods. This review of cultured meat and decellularized scaffold biomaterials provides new insight and great potential research prospects in food application and cellular agriculture.

Sustainable Future Protein Foods: The Challenges and the Future of Cultivated Meat.

Global pressure from consumers to improve animal welfare, and reduce microbiological risks or the use of antibiotics pose new challenges for the meat industry. Today's livestock production, despite many undertaken measures, is still far from being sustainable. This forced the need to work on alternative protein types that come from plants, insects, fungi, or cell culture processes. Due to some technical and legal barriers, cultivated meat is not present on the European market, however, in 2020 it was approved in Singapore and in 2022 in the USA. While the technology of obtaining cell cultures from animal muscles has been known and successfully practiced for years, the production of a stable piece of meat with appropriate texture, taste, and smell, is still a problem for several scientific groups related to subsequent companies trying to obtain the highest quality product, in line with the expectations of customers. Although the work on optimal cell meat production has been going on for years, it is still in an early stage, mainly due to several limitations that represent milestones for industrial production. The most important are: the culture media (without animal serum), which will provide an environment for optimal muscle development, natural or close to natural (but still safe for the consumer) stable scaffolds for growing cells. Here, we review the actual knowledge about the above-mentioned challenges which make the production of cellular meat not yet developed on an industrial scale.

Chitosan­sodium alginate-collagen/gelatin three-dimensional edible scaffolds for building a structured model for cell cultured meat.

Cell cultured meat (CCM) production is an innovative technology that does not depend on livestock farming practices to produce meat. The construction of structured CCM requires a three-dimensional (3D) scaffold to mimic the extracellular matrix to provide mechanical support for the cells. Furthermore, the 3D scaffolds should be edible and have good biocompatibility and tissue-like texture. Here, we demonstrated a 3D edible chitosan­sodium alginate-collagen/gelatin (CS-SA-Col/Gel) scaffold that can support the adhesion and proliferation of porcine skeletal muscle satellite cells, culminating in the construction of a structured CCM model. The 3D edible scaffolds were prepared by freeze-drying using electrostatic interactions between chitosan and sodium alginate. Initially, the physicochemical properties and structural characteristics of different scaffolds were explored, and the biocompatibility of the scaffolds was evaluated using the C2C12 cell model. The results showed that the 2-CS-SA-Col1-Gel scaffold provided stable mechanical support and abundant adhesion sites for the cells. Subsequently, we inoculated porcine skeletal muscle satellite cells on the 2-CS-SA-Col1-Gel scaffold and induced differentiation for a total of 14 days. Immunofluorescence staining results showed cytoskeleton formation, and Western blotting (WB) and qPCR results showed upregulation of skeletal proteins and myogenic genes. Ultimately, the structured CCM model has similar textural properties (chewiness, springiness and resilience) and appearance to those of fresh pork. In conclusion, the method of constructing 3D edible scaffolds to prepare structured CCM models exhibits the potential to produce cell cultured meat.

Relative Effectiveness of Cell-Cultured versus Egg-Based Seasonal Influenza Vaccines in Preventing Influenza-Related Outcomes in Subjects 18 Years Old or Older: A Systematic Review and Meta-Analysis.

Avian mutations in vaccine strains obtained from embryonated eggs could impair vaccine effectiveness. We performed a systematic review and meta-analysis of the adjusted relative vaccine effectiveness (arVE) of seed cell-cultured influenza vaccines (ccIV) compared to egg-based influenza vaccines (eIV) in preventing laboratory-confirmed influenza related outcomes (IRO) or IRO by clinical codes, in subjects 18 and over. We completed the literature search in January 2021; applied exclusion criteria, evaluated risk of bias of the evidence, and performed heterogeneity, publication bias, qualitative, quantitative and sensitivity analyses. All estimates were computed using a random approach. International Prospective Register of Systematic Reviews, CRD42021228290. We identified 12 publications that reported 26 adjusted arVE results. Five publications reported 13 laboratory confirmed arVE and seven reported 13 code-ascertained arVE. Nine publications with 22 results were at low risk of bias. Heterogeneity was explained by season. We found a significant 11% (8 to 14%) adjusted arVE favoring ccIV in preventing any IRO in the 2017-2018 influenza season. The arVE was 3% (-2% to 7%) in the 2018-2019 influenza season. We found moderate evidence of a significant advantage of the ccIV in preventing IRO, compared to eIV, in a well-matched A(H3N2) predominant season.

Perspective: Unpacking the Wicked Challenges for Alternative Proteins in the United States: Can Highly Processed Plant-Based and Cell-Cultured Food and Beverage Products Support Healthy and Sustainable Diets and Food Systems?

Over the past decade, a plethora of alternative protein (AP) products has entered the US food system as plant-based food and beverage products. These AP products, which include plant-based meat and dairy alternatives and cell-cultured meat and seafood products, are being developed for the marketplace to simulate the appearance, texture, taste, and flavor and nutritional profiles of animal products. The new generation of AP plant-based and cell-cultured food and beverage products are part of a market-driven narrative that has embraced technology to address future human health, environmental, ethical, and planetary health challenges. This perspective article synthesizes evidence about the benefits of adopting minimally processed plant-based diets that support sustainable food systems and human and planetary health. Thereafter, it examines 4 wicked challenges related to AP products in the US context that include 1) a confusing marketing landscape for the public; 2) diverse views and varying acceptance among consumers about the health and environmental benefits of these products; 3) inadequate education and labeling provided by federal agencies to enable consumers to understand how these may support healthy sustainable diets; and 4) slow federal policy and regulatory actions to address the range of AP products and provide industry guidance. The article concludes with suggested policies and actions for government agencies and food system actors to address these challenges. Future research and actions are needed to balance the human health, equity, animal welfare, and economic viability goals and to clarify how AP products may support safe, healthy, sustainable diets and food systems.

US and UK Consumer Adoption of Cultivated Meat: A Segmentation Study.

Despite growing evidence of the environmental and public health threats posed by today's intensive animal production, consumers in the west remain largely attached to meat. Cultivated meat offers a way to grow meat directly from cells, circumventing these issues as well as the use of animals altogether. The aim of this study was to assess the overall consumer markets and a range of preferences around cultivated meat in the US and the UK relating to nomenclature, genetic modification, health enhancements, and other features. To this end, we recruited large representative samples to participate in an online survey about cultivated meat, and subsequently analyzed segments (a) in the early majority population (guided by the Diffusion of Innovations Model), (b) by generation, and (c) in the general population. Our findings showed a high level of openness (80%) in both the US and UK populations, with 40% somewhat or moderately likely to try and 40% highly likely to try. Younger generations had the greatest openness: 88% of Gen Z, 85% of Millennials, 77% of Gen X, and 72% of Baby Boomers were at least somewhat open to trying cultivated meat. All segments envisioned cultivated meat to be nearly half of their total meat intake. Findings show that consumers prefer the terms 'cultured' and 'cultivated' over 'cell-based' and 'cell-cultured' for use in a social context and on packages, even though they perceive these terms as less descriptive. The most important on-package label was one indicating government assurances, and participants preferred non-GM products over GM products. We also found that US consumers prefer nutritionally superior meat over nutritionally equivalent meat. We discuss implications for product development, messaging, and understanding the likely adoption path of this food innovation.

Reports of cell-based influenza vaccine administered during pregnancy in the Vaccine Adverse Event Reporting System (VAERS), 2013-2020.

BACKGROUND: In November 2012, the first cell cultured influenza vaccine, a trivalent subunit inactivated influenza vaccine (Flucelvax(®), ccIIV3), was approved in the United States for adults aged ≥18 years. A quadrivalent version (ccIIV4) was later approved in 2016 and replaced ccIIV3. The safety of ccIIV3 or ccIIV4 (ccIIV) was not assessed for pregnant women or their infants during pre-licensure studies. OBJECTIVE: To assess the safety of ccIIV administered during pregnancy in pregnant women and their infants whose reports were submitted to VAERS during 2013-2020. MATERIAL AND METHODS: We searched VAERS for United States reports of adverse events (AEs) in pregnant women who received ccIIV from 1 July 2013 through 31 May 2020. Clinicians reviewed reports and available medical records and assigned a primary clinical category for each report. Reports were coded as serious based on the Code of Federal Regulations definition. RESULTS: VAERS received 391 reports following ccIIV administered to pregnant women. Twenty-four (6.1%) were serious. Two neonatal deaths were reported. No maternal deaths occurred. Among reports with trimester information (n = 340), ccIIV was administered during the second trimester in 170 (50%). The most frequent pregnancy-specific AE was premature delivery in 85 (21.7%) reports, followed by dysmature placenta in 13 (3.3%) and pre-eclampsia/eclampsia in ten (2.3%). The most common non-pregnancy specific conditions were infectious conditions in 32 (8.2%). Among infant conditions, low birth weight was reported in 62 (15.9%) reports. Fifteen birth defects were reported; in 12 with gestational age information, administration of the vaccine occurred late in the second trimester or later. CONCLUSIONS: Review of maternal ccIIV reports in VAERS was not unexpectedly different from other maternal influenza vaccine safety VAERS reviews.

An empirical assessment of common or usual names to label cell-based seafood products.

An important consideration in the commercialization of cell-based meat, poultry, and seafood is what common or usual name to use on package labels to meet U.S. Food and Drug Administration (FDA) regulations. However, naming these products has been the subject of considerable debate. This study used a 3 × 10 between-subjects online experiment involving a quota sample of 3,186 U.S. adult panel participants to test common or usual names using images of realistic packages of three types of seafood that a consumer might encounter in a supermarket. The terms tested were, "cell-based seafood," "cell-cultured seafood," "cultivated seafood," and "cultured seafood" and the phrases, "produced using cellular aquaculture," "cultivated from the cells of ____," and "grown directly from the cells of ____," where the blanks are filled by the name of the seafood product. Five criteria were used for evaluation, including each term's ability to: enable consumers to distinguish cell-based seafood from wild and farmed fish, to signal potential allergenicity, be seen by consumers as an appropriate term to identify the product, not disparage either cell-based or conventional products, and not evoke thoughts, images, or emotions that are inconsistent with the idea that the products are safe, healthy, and nutritious. The results showed that "cell-based seafood" outperforms the other names tested. It enables consumers to recognize that the products are neither wild caught nor farm raised, signals potential allergenicity, is seen as an appropriate name for describing the technology/process, and it performs well with respect to measures of consumer acceptance, particularly in comparison to conventional products. PRACTICAL APPLICATION: Creating consensus around a single common or usual name for cell-based meat, poultry, and seafood products is clearly important both for regulatory reasons and for shaping public perceptions and understanding of the products that are labeled with it. Our findings suggest that "cell-based" is the best common or usual name for seafood products that both meets FDA regulatory requirements and performs well with respect to potential consumer acceptance. Consistent use of this term by industry, advocates, and regulators would help orient consumers to what is likely to be a transformational food technology.

Prospects and challenges for cell-cultured fat as a novel food ingredient.

BACKGROUND: In vitro meat production has been proposed as a solution to environmental and animal welfare issues associated with animal agriculture. While most academic work on cell-cultured meat has focused on innovations for scalable muscle tissue culture, fat production is an important and often neglected component of this technology. Developing suitable biomanufacturing strategies for adipose tissue from agriculturally relevant animal species may be particularly beneficial due to the potential use of cell-cultured fat as a novel food ingredient. SCOPE AND APPROACH: Here we review the relevant studies from areas of meat science, cell biology, tissue engineering, and bioprocess engineering to provide a foundation for the development of in vitro fat production systems. We provide an overview of adipose tissue biology and functionality with respect to meat products, then explore cell lines, bioreactors, and tissue engineering strategies of potential utility for in vitro adipose tissue production for food. Regulation and consumer acceptance are also discussed. KEY FINDINGS AND CONCLUSIONS: Existing strategies and paradigms are insufficient to meet the full set of unique needs for a cell-cultured fat manufacturing platform, as tradeoffs are often present between simplicity, scalability, stability, and projected cost. Identification and validation of appropriate cell lines, bioprocess strategies, and tissue engineering techniques must therefore be an iterative process as a deeper understanding of the needs and opportunities for cell-cultured fat develops.

Relative Effectiveness of Influenza Vaccines Among the United States Elderly, 2018-2019.

BACKGROUND: Studies among individuals ages ≥65 years have found a moderately higher relative vaccine effectiveness (RVE) for the high-dose (HD) influenza vaccine compared with standard-dose (SD) products for most seasons. Studies during the A(H3N2)-dominated 2017-2018 season showed slightly higher RVE for the cell-cultured vaccine compared with SD egg-based vaccines. We investigated the RVE of influenza vaccines among Medicare beneficiaries ages ≥65 years during the 2018-2019 season. METHODS: This is a retrospective cohort study using inverse probability of treatment weighting and Poisson regression to evaluate RVE in preventing influenza hospital encounters. RESULTS: Among 12 777 214 beneficiaries, the egg-based adjuvanted (RVE, 7.7%; 95% confidence interval [CI], 3.9%-11.4%) and HD (RVE, 4.9%; 95% CI, 1.7%-8.1%) vaccines were marginally more effective than the egg-based quadrivalent vaccines. The cell-cultured quadrivalent vaccine was not significantly more effective than the egg-based quadrivalent vaccine (RVE, 2.5%; 95% CI, -2.4% to 7.3%). CONCLUSIONS: We did not find major effectiveness differences between licensed vaccines used among the elderly during the 2018-2019 season. Consistent with prior research, we found that the egg-based adjuvanted and HD vaccines were slightly more effective than the egg-based quadrivalent vaccines.

Relative Effectiveness of Cell-Cultured and Egg-Based Influenza Vaccines Among Elderly Persons in the United States, 2017-2018.

BACKGROUND: The low influenza vaccine effectiveness (VE) observed during the A(H3N2)-dominated 2017-2018 season may be due to vaccine virus adaptation to growth in eggs. We compared the effectiveness of cell-cultured and egg-based vaccines among Medicare beneficiaries. METHODS: Retrospective cohort study on Medicare beneficiaries aged ≥65 years who received an influenza vaccine (cell-cultured, egg-based quadrivalent; egg-based high-dose, adjuvanted, or standard-dose trivalent) during the 2017-2018 season. We used Poisson regression to evaluate relative VE (RVE) in preventing influenza-related hospital encounters. RESULTS: Of >13 million beneficiaries, RVE for cell-cultured vaccines relative to egg-based quadrivalent vaccines was 10% (95% confidence interval [CI], 7%-13%). In a midseason interim analysis, this estimate was 16.5% (95% CI, 10.3%-22.2%). In a 5-way comparison, cell-cultured (RVE, 11%; 95% CI, 8%-14%) and egg-based high-dose (RVE, 9%; 95% CI, 7%-11%) vaccines were more effective than egg-based quadrivalent vaccines. CONCLUSIONS: The modest VE difference between cell-cultured and egg-based vaccines only partially explains the low overall VE reported by the Centers for Disease Control and Prevention, suggesting that egg adaptation was not the main contributor to the low VE found among individuals aged ≥65 years. The midseason interim analysis we performed demonstrates that our methods can be used to evaluate VE actively during the influenza season.

Quantitative proteomic analysis of human retinal microvascular endothelial cells stimulated with 4-Hydroxynonenal / 中华眼底病杂志

Objective To detect the protein expression change in the proliferation of human retinal microvascular endothelial cells (hRMECs) stimulated with 4-Hydroxynonenal (4-HNE).Methods hRMECs were in a logarithmic growth phase, and then were separated into 4-HNE-stimulated group and negative control group. The concentration of 4-HNE included 5, 10, 20 and 50 μmol/L in 4-HNE-stimulated group, while the negative control group was added in the same volume of ethanol (the solvent of 4-HNE). Then the cells were stimulated with 4-HNE for 24 hours following by CCK-8 kits incubating for 4 hours to detect absorbance. It was found that 10 μmol/L 4-HNE had the most obvious effect on the proliferation of hRMECs. Therefore, the cellular proteins from 10 μmol/L 4-HNE-stimulated group and negative control group were acquired and prepared by FASP sample preparation method. Data independent acquisition was used for data acquisition, and the GO analysis and pathway enrichment were performed for analysis of differentially expressed proteins. Results CCK-8 kits detection results showed that theA value of the 10 and 20 μmol/L 4-HNE-stimulated groups were significantly higher than negative control group and 5 μmol/L 4-HNE-stimulated group (F=25.42, P<0.01), while there were no differences between 10 and 20 μmol/L 4-HNE-stimulated groups, and theA value of 50 μmol/L 4-HNE-stimulated groups was lower than negative control. A total of 2710 quantifiable proteins were identified by peoteomics,and 118 proteins were differentially expressed (fold change>1.5,P<0.05). Seventy-two proteins were up-regulated after 4-HNE stimulation, whereas 46 proteins were down-regulated. Particularly, the expressions of Heme oxygenase-1, Sulfoxdoxin-1, Heat shock protein A1B, Thioredoxin reductase-1, Glutathione reductase, ATPase and prothrombin were increased when cells were added in 4-HNE, whereas the expressions of apolipoprotein A1 and programmed cell death protein 4 were decreased. These differentially expressed proteins were mainly involved in the biological processes such as oxidative stress, cell detoxification, and ATPase-coupled membrane transport.Conclusion After stimulated with 4-HNE, the oxidative stress, cell detoxification, and ATPase-coupled membrane transport protein expression may change in hRMECs in order to regulate oxidative stress and growth situation.

Effect of atorvastatin on the expression of chemokine receptor-4 and migration ability of rat bone marrow mesenchymal stem cells cultivated in vitro / 中华眼底病杂志

Objective To investigate the effects of migration and expression from chemokine receptor 4 (chemokine receptor-4,CXCR4) of rat bone marrow mesenchymal stem cells (BMSCs) which were pretreated by atorvastatin (ATV) in vitro.Methods Isolated,cultivated,identified the BMSCs,pretreated P4-P6 of BMSCs with different concentrations of ATV for 12 hours.The experimental group was divided into control group,0.1 nM/L (group 0.1 nM),1 nM/L (1 nM group),10 nM/L (10 nM group),100 nM/L (100 nM group),1 000 nM/L (1 000 nM group).The mRNA and protein of CXCR4 were determined by real time-polymerase chain reaction and Western blot.Immunofluoreseence assay were used to detect the expression levels of CXCR4.The migration ability of BMSCs were measured by transwell chamber.Results Immunofluoreseence assay showed the protein level of CXCR4 of group 1 nM and 10 nM were significantly higher than the other group.RT-PCR and Western blot showed the protein and mRNA level of CXCR4 in 10 nM was higher than that in group 1 rM.The migration ability of group 10 nM was higher than 1 nM and control group.Conclusions ATV can be dose-dependent promote expression levels of CXCR4 of BMSCs cultivated in vitro.

Enhanced production of enveloped viruses in BST-2-deficient cell lines.

Despite all the advantages that cell-cultured influenza vaccines have over egg-based influenza vaccines, the inferior productivity of cell-culture systems is a major drawback that must be addressed. BST-2 (tetherin) is a host restriction factor which inhibits budding-out of various enveloped viruses from infected host cells. We developed BST-2-deficient MDCK and Vero cell lines to increase influenza virus release in cell culture. BST-2 gene knock-out resulted in increased release of viral particles into the culture medium, by at least 2-fold and up to 50-fold compared to release from wild-type counterpart cells depending on cell line and virus type. The effect was not influenza virus/MDCK/Vero-specific, but was also present in a broad range of host cells and virus families; we observed similar results in murine, human, canine, and monkey cell lines with viruses including MHV-68 (Herpesviridae), influenza A virus (Orthomyxoviridae), porcine epidemic diarrhea virus (Coronaviridae), and vaccinia virus (Poxviridae). Our results suggest that the elimination of BST-2 expression in virus-producing cell lines can enhance the production of viral vaccines. Biotechnol. Bioeng.2017;114: 2289-2297. © 2017 Wiley Periodicals, Inc.

Semi-degradable porous poly (vinyl alcohol) hydrogel scaffold for cartilage repair: Evaluation of the initial and cell-cultured tribological properties.

Tissue engineering for articular cartilage repair has shown success in ensuring the integration of neocartilage with surrounding tissue, but the rapid restoration of biomechanical and biotribological functions remains a significant challenge. Poly (vinyl alcohol) (PVA) hydrogel is regarded as a potential articular cartilage replacement for its fair mechanical strength and low surface friction, while its lack of bioactivity limits its utility. Combining the advantages of tissue engineering materials and PVA hydrogel, we developed a semi-degradable porous PVA hydrogel through addition of ploy (lactic-co-glycolic acid) (PLGA) microspheres and salt-leaching technique. Friction coefficient, continuous friction tested by a ball-plate tribometer and worn surface observed by Environmental Scanning Electron Microscopy (ESEM) were characterized for scaffolds prepared with variable porogen and PLGA content. Scaffolds cultured with rabbit chondrocytes for 4 weeks were also studied. The results showed that friction coefficient increased with a rise in porogen content, while it firstly increased and then decreased with increasing PLGA content. Similar results were obtained from cell-cultured scaffolds. In continuous friction test and worn morphology characterization, samples were more prone to be damaged with an increase in porogen and PLGA content. However, wear resistance was obviously improved for all scaffolds after 4 weeks of culture, though friction coefficient went up to a certain extent.

Lowing-intraocular pressure and improving aqueous humor dynamic effects of qingguangan drug granules in spontaneous glaucoma mice / 中华实验眼科杂志

Background Researches showed that the increase of intraocular pressure (IOP) in glaucomatous eye is associated with the increasing resistance to aqueous humor outflow effects of transforming growth factor-β (TGF-β) and CD44.Qingguangan is a traditional Chinese medicine and used to treat glaucoma.However,its mechanism of lowing-IOP effect is not elucidated.Objective This study was to investigate the lowing-IOP effect and mechanism of qingguangan granule in DBA/2J mouse,a spontaneous glaucoma model mice.Methods Ten 3 month-old female DBA/2J mice with normal IOP were chosen as control group,and 20 spontaneous ocular hypertension mice aged 9 months were randomized into high IOP group and qingguangan-treated group,with 10 mice for each group.The qingguangan (2.5 g/kg) was administered by gavaging twice per day for consecutive 15 days in the qingguangan-treated group,and normal saline solution was used in the same way in the control group and high IOP group.IOP was measured by anterior chamber injection/suction system at a perfusion rate of 2.5 and 5.0 μl/min,respectively,and the coefficient of aqueous outflow facility (C value) and outflow resistance (R value) were calculated.Another 60 3-month-old DBA/2J mice were randomized into blank control group gavaged with normal saline solution and high-,middle-and low-dose qingguangan groups gavaged with 25.00,12.50 and 6.25 g/kg drugs,respectively,and the mouse serum containing drugs was extracted 7 days after treatment.The scleral tissue with trabecular meshwork were obtained for the culture of trabecular meshwork cells and the cells were identified by immunohistochemistry of fibronectin (FN),laminin (LN) and neuronspecific enolase (NSE).TGF-β was added into the medium for 24 hours with the final concentration of 0,5,10,20,50 and 100 ng/ml,and MMT chromatometry was employed to detect the cell vitality.The cells pre-treated with 20 ng/ml TGF-β were treated with different concentration of drug serum for 24,48 and 72 hours,and the level of TGF-β2 receptor in cell supernatant and the expression of CD44 protein in the cells were detected by ELISA and Western blot assay,respectively.Results The IOPs with perfusion both 2.5 μl/min and 5.0μl/min in the qingguangan-treated group and the control group were significantly lower than those in the high IOP group (all at P＜0.01).Compared with the high IOP group,the C value was significantly reduced (2.35±1.34 vs.1.08±0.36) and the R value was evidently elevated (0.64±0.55 vs.1.05± 0.47) in the qingguangan-treated group (all at P＜0.01).Cultured cells were spindle-shaped with the positive response to FN,LN and NSE antibody.The cell vitality was lower in the 5,10 and 20 ng/ml TGF-β group than that in the 0 ng/ml TGF-β group (all at P＜0.05).Compared with the blank control group,the TGF-β2 receptor content in the supernatant and the related expression level of CD44 protein in the cells were elevated in the TGF-β-treated group (all at P＜0.01),and TGF-β2 receptor contents and CD44 expression levels in the TGF-β+high dose drug serum group was significantly lower than those in the TGF-β group and TGF-β +low dose drug serum group 24,48 and 72 hours after culture (all at P＜0.01).Conclusions Qingguangan can lower IOP of spontaneous glaucoma mice by affecting aqueous humor dynamics.Serum containing qingguangan down-regulates the expressions of TGF-β2 receptors and CD44 in trabecular meshwork cells in vitro.