Callicarpa is an important medicinal plant in China, which has hemostatic, antibacterial, and antioxidant pharmacological effects, and the efficacy of astringing and arresting bleeding, clearing heat and detoxification, activating blood, and resolving stasis is outstanding. At the same time, Callicarpa can be used as an ornamental plant because of its gorgeous flowers and fruits. Callicarpa has good market development prospects, but the long seed reproduction cycle directly limits the large demand for seedlings in its industrial development. Asexual reproduction technology is the basis for the industrialization development of Callicarpa, which is helpful in producing high-quality seedlings and medicinal materials. Although Chinese and foreign scholars have achieved remarkable results in the study of asexual reproduction of Callicarpa, there is no report on the large-scale production of seedlings of Callicarpa. Integrating and improving its asexual reproduction technology can promote the development and utilization of Callicarpa, improve its medicinal value, and create significant economic benefits. Therefore, the authors reviewed the effects of cutting, season, plant growth regulators, substrates, environment, and management measures on the cutting of Callicarpa and the research progress of tissue culture propagation affected by explants, basic media, exogenous additives, subculture cycles, culture conditions, and transplanting substrates. The mechanism of adventitious root formation was reviewed at the cellular, physiological, and biochemical levels, so as to put forward the problems and corresponding solutions in the study of asexual propagation technology and regulatory mechanism of Callicarpa and point out the future research directions. The study aims to provide a reference for in-depth research on the asexual propagation technology of Callicarpa and the commercial production of its high-quality seedlings.
Reproduction, Asexual ; Plants, Medicinal/physiology* ; Seedlings/growth & development* ; Tissue Culture Techniques

As a high-quality moxibustion material, Artemisia stolonifera has high economic value and research prospects. However, due to difficulties in seed germination, its wild germplasm resources are sparsely distributed in China. This study used young stem segments grown in the current year to investigate the effects of explant sterilization, different combinations and concentrations of plant growth regulators on the proliferation and rooting of adventitious shoots, with the aim of constructing an in vitro rapid propagation technology system for A. stolonifera. The results showed that the lowest contamination rate of 25.83% was achieved when sterilizing the stem segments by rinsing with running water for 30 min, soaking in 75% ethanol for 30 s, followed by a 5 min treatment with 0.1% HgCl_2, 10 min with 8% NaClO, and 10 min with 0.6% phytosaniline. There was no browning of the stem segments, and surface sterilization of the A. stolonifera stem segments was successfully achieved. In the induction culture phase, when the concentration of kinetin(KT) was 0.05 mg·L~(-1) and 6-benzylaminopurine(6-BA) was 0.05 mg·L~(-1), the adventitious shoot proliferation coefficient was 2.02, effectively promoting the proliferation and growth of A. stolonifera. In the rooting culture phase, 0.1 mg·L~(-1) 1-naphthaleneacetic acid(NAA) effectively induced A. stolonifera test-tube seedlings to root within a short period, achieving a rooting rate of 100%. The addition of a small amount of activated charcoal also promoted rooting and strengthened seedling growth. The survival rate of A. stolonifera seedlings transplanted into a substrate consisting of 90% nutrient soil and 10% perlite was 100%. This study established an efficient in vitro rapid propagation system for A. stolonifera, overcoming difficulties with seed germination, shortening the breeding cycle, and reducing production and planting costs. It provides technical support for the introduction, domestication, seedling propagation, germplasm conservation, and industrial development of A. stolonifera.
Artemisia/drug effects* ; Tissue Culture Techniques/methods* ; Plant Growth Regulators/pharmacology* ; Plant Stems/drug effects* ; Plant Shoots/drug effects*

Organoids are three-dimensional (3D) cellular structures formed through the differentiation and self-organization of pluripotent stem cells or tissue-derived cells, showing considerable potential in the research on disease mechanism, personalized medicine, and developmental biology. However, the development of organoids is limited by the complex composition, batch-to-batch variations, and immunogenicity of basement-membrane matrix in the current culture system, which hinders the clinical translation and in vivo applications of organoids. Hydrogels are highly hydrated 3D polymer network materials, with modifiable mechanical and biochemical properties by engineering, representing an ideal alternative to basement-membrane matrix. This article reviews the research progress in engineered hydrogels with defined composition currently used in organoid culture. We introduce the structural characteristics and engineering design considerations of hydrogels, emphasize the latest research progress and specific application cases, and discuss the future development of these engineered hydrogels, provide valuable insights for the further advancement and optimization of engineered hydrogels for organoid.
Hydrogels/chemistry* ; Organoids/cytology* ; Tissue Engineering/methods* ; Humans ; Animals ; Pluripotent Stem Cells/cytology* ; Cell Culture Techniques, Three Dimensional/methods* ; Tissue Scaffolds

OBJECTIVE: To review the application of cell derived decellularized extracellular matrix (CDM) in tissue engineering. METHODS: The literature related to the application of CDM in tissue engineering was extensively reviewed and analyzed. RESULTS: CDM is a mixture of cells and their secretory products obtained by culturing cells in vitro for a period of time, and then the mixture is treated by decellularization. Compared with tissue derived decellularized extracellular matrix (TDM), CDM can screen and utilize pathogen-free autologous cells, effectively avoiding the possible shortcomings of TDM, such as immune response and limited sources. In addition, by selecting the cell source, controlling the culture conditions, and selecting the template scaffold, the composition, structure, and mechanical properties of the scaffold can be controlled to obtain the desired scaffold. CDM retains the components and microstructure of extracellular matrix and has excellent biological functions, so it has become the focus of tissue engineering scaffolds. CONCLUSION CDM is superior in the field of tissue engineering because of its outstanding adjustability, safety, and high bioactivity. With the continuous progress of technology, CDM stents suitable for clinical use are expected to continue to emerge.
Tissue Engineering/methods* ; Tissue Scaffolds/chemistry* ; Humans ; Decellularized Extracellular Matrix/chemistry* ; Cells, Cultured ; Extracellular Matrix ; Animals ; Biocompatible Materials/chemistry* ; Cell Culture Techniques

In order to establish a stable in vitro culture platform for chicken small intestine three-dimensional (3D) organoids, in this study, crypt cells were collected from the small intestine of 18-day-old embryos of AA broilers. On the basis of the L-WRN conditioned medium, we optimized the culture conditions of chicken small intestinal organoids by adjusting the proportions of nicotinamide, N-acetylcysteine, LY2157299, CHIR99021, Jagged-1, FGF, and other cytokines to select the medium suitable for the long-term stable growth of the organoids. The optimization results showed that the addition of 1.5 µmol/L CHIR99021 significantly improved the organoid formation efficiency and organoid diameter. When 0.5 µmol/L Jagged-1 was added, a small amount of bud-like tissue appeared in organoids. After the addition of 50 ng/mL FGF-2, the rate of organoid germination was significantly increased. The 1.5 µmol/L CHIR99021, 0.5 µmol/L Jagged-1, and 50 ng/mL FGF-2 added in the medium can cooperate with each other to improve the formation and speed up the proliferation and differentiation of organoids, while improving the stemness maintenance of cells. The morphology, cell types, and culture characteristics of chicken small intestinal organoids were studied by HE staining, transmission electron microscopy, reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), indirect immunofluorescence, and immunohistochemistry. The results showed that the 3D organoids of the chicken small intestine cultured in vitro were morphologically consistent with the chicken intestinal tissue and contained differentiated epithelial cells. In summary, we successfully established an in vitro culture system for chicken small intestinal organoids, providing a new method for the subsequent research on chicken intestinal physiology, pathology, and host-pathogen interaction mechanism and the development of relevant drugs.
Animals ; Organoids/metabolism* ; Intestine, Small/drug effects* ; Chickens ; Cell Culture Techniques/methods* ; Culture Media ; Chick Embryo ; Tissue Culture Techniques/methods*

Since researchers have found that the conditioned medium and exosomes of mesenchymal stem cells (MSCs) had the biological effects equivalent to those of MSCs, MSC exosomes (MSC-Exos), the representative product of MSCs' paracrine effect, have become the research focus of the "cell-free" therapy of MSCs. However, most researchers currently use conventional culture condition to culture MSCs and then isolate exosomes for the treatment of wound or other diseases. Theoretically, the paracrine effect of MSCs is directly associated with the pathological condition of the wound (disease) microenvironment or in vitro culture condition, and their paracrine components and biological effects may be altered with the changes of the wound (disease) microenvironment or in vitro culture condition. Thus, the feasibility of using traditional culture condition to culture MSCs for exosome extraction for the treatment of different diseases without considering the actual situation of the disease to be treated needs further discussion. Therefore, the author suggests that the research of MSC-Exos should consider the microenvironment of the wound (disease) to be treated. as much as possible, otherwise the extracted MSC-Exos may not be "accurate" or may not really achieve the treatment effect of MSCs. In this article, we summarized some thoughts of the author and problems related to the researches about MSC-Exos and wound microenvironment, and hoped to discuss with researchers.
Exosomes ; Cell- and Tissue-Based Therapy ; Culture Media, Conditioned ; Mesenchymal Stem Cells

Salivary glands are important organs in the oral and maxillofacial region. Environment and genetic factors may cause salivary gland tumors or non-neoplastic diseases, but the mechanisms of those diseases are still unclear. One of the important reasons is the short of researching media and model. As a new technique and research model, organoids have been widely used in the research of various diseases. Organoid culture plays a bridging role between two-dimensional cell culture and living animal models, and it is also the most promising translational research model that could connect the clinical research to basic research. This review will discuss the recent development of organoid techniques in the culture of normal salivary glands and salivary gland tumors, also their applications and challenges in tissue engineering, etiological research, and tumor therapy.
Animals ; Cell Culture Techniques ; Organoids ; Salivary Gland Neoplasms ; Salivary Glands ; Tissue Engineering

BACKGROUND AND OBJECTIVES: Oxidative stress (OS) is known to be an important factor of male infertility. Adipose-derived mesenchymal stem cells (AD-MSCs) are known to have immune-modulatory and anti-oxidant effects through their secretions, hence raising the idea of their potential benefit to improve sperm parameters. This study aims at investigating the effect of AD-MSCs conditioned medium (CM) on human sperm parameters in the presence and absence of H2O2-induced OS.METHODS AND RESULTS: Sperm samples were collected from 30 healthy men and divided into two groups: non-stressed and H2O2-stressed. Isolated AD-MSCs from healthy donors undergoing liposuction were cultured and CM was collected at 24, 48 and 72 h. Both sperm groups were cultured with CM and a time course was performed followed by an evaluation of sperm parameters. The incubation of non-stressed and stressed sperm samples with AD-MSCs-CM for 24 h was found to have the optimum impact on sperm vacuolization, DNA fragmentation and OS levels in comparison to other incubation timings, while preserving motility, viability and morphology of cells. Incubation with CM improved all sperm parameters except morphology in comparison to the non-treated group, with the best effect noted with CM collected at 24 h rather than 48 or 72 h for sperm vacuolization and DNA fragmentation. When compared to fresh semen parameters (T0), samples cultured with CM 24 h showed a significant decrease in sperm vacuolization and DNA fragmentation while keeping other parameters stable.CONCLUSIONS: AD-MSCSs-CM improves sperm quality, and hence can be used in treating infertility and subsequently enhancing IVF outcomes.
Antioxidants ; Culture Media, Conditioned ; DNA Fragmentation ; DNA ; Humans ; In Vitro Techniques ; Infertility ; Infertility, Male ; Lipectomy ; Male ; Mesenchymal Stromal Cells ; Oxidative Stress ; Semen ; Spermatozoa ; Tissue Donors

BACKGROUND: Tissue engineering is a multidisciplinary field which attracted much attention in recent years. One of the most important issue in tissue engineering is how to obtain high cell numbers and tissue regeneration while maintaining appropriate cellular characteristics in vitro for restoring damaged or dysfunctional body tissues and organs. These demands can be achieved by the use of three dimensional (3D) dynamic cultures of cells combined with cell-adhesive micro-carriers. METHODS: In this study, human mesenchymal stem cells (hMSCs) were cultured in a wave-bioreactor system for up to 100 days, after seeding on Cultisphere-S porous gelatin micro-carriers. Cell counting was performed at the time points of 7, 12, 17, 31 days and compared to those of hMSCs cultured under static condition. Higher growth and proliferation rates was achieved in wave-type dynamic culture, when cell culture continued to day 31. A scanning electron microscope (SEM) photographs, both live and dead and MTT assays were taken to confirm the survival and distribution of cells on porous gelatin micro-carrier surfaces. The results of histological stains such as hematoxylin and eosin, Masson’s trichrome, Alcian blue and Alizarin red S also showed improved proliferation and tissue regeneration of hMSCs on porous gelatin micro-carriers. CONCLUSION: The experimental results demonstrated the effect and importance of both micro-carriers and bioreactor in hMSC expansion on cell proliferation and migration as well as extracellular matrix formation on the superficial and pore surfaces of the porous gelatin micro-carriers, and then their inter-connections, leading to tissue regeneration.
Alcian Blue ; Bioreactors ; Cell Count ; Cell Culture Techniques ; Cell Proliferation ; Coloring Agents ; Eosine Yellowish-(YS) ; Extracellular Matrix ; Gelatin ; Hematoxylin ; Humans ; In Vitro Techniques ; Mesenchymal Stromal Cells ; Regeneration ; Tissue Engineering

As one of the top 10 breakthrough and emerging technologies in the world in 2018, cultured meat has attracted extensive attention due to its advantages of traceable origin, food safety and green sustainable development. Europe and the United States have invested a lot of resources to focus on research about cultured meat, which will affect our domestic meat and food market in the future. At present, the challenge of cultured meat production is how to efficiently simulate the growth environment of animal muscle tissue and realize large-scale production in bioreactor. Although cell tissue engineering has been deeply studied and achieved varying successful application, it is still difficult to obtain large-scale cultured meat production due to the high cost and technical requirements. Therefore, the development of efficient and safe cell culture technology is an urgent problem for large-scale cultured meat production, which can effectively reduce costs and achieve industrial application. In this review, we summarize the research progress of animal cell tissue culture technology used for cultured meat, and highlighted the current challenges and possible strategies in further applications.
Animals ; Bioreactors ; Cell Culture Techniques ; Meat ; Tissue Engineering ; United States