Profiling Dynamic Patterns of Single-Cell Motility.

Cell motility plays an essential role in many biological processes as cells move and interact within their local microenvironments. Current methods for quantifying cell motility typically involve tracking individual cells over time, but the results are often presented as averaged values across cell populations. While informative, these ensemble approaches have limitations in assessing cellular heterogeneity and identifying generalizable patterns of single-cell behaviors, at baseline and in response to perturbations. In this study, CaMI is introduced, a computational framework designed to leverage the single-cell nature of motility data. CaMI identifies and classifies distinct spatio-temporal behaviors of individual cells, enabling robust classification of single-cell motility patterns in a large dataset (n = 74 253 cells). This framework allows quantification of spatial and temporal heterogeneities, determination of single-cell motility behaviors across various biological conditions and provides a visualization scheme for direct interpretation of dynamic cell behaviors. Importantly, CaMI reveals insights that conventional cell motility analyses may overlook, showcasing its utility in uncovering robust biological insights. Together, a multivariate framework is presented to classify emergent patterns of single-cell motility, emphasizing the critical role of cellular heterogeneity in shaping cell behaviors across populations.

Highly Ordered Nanotube-Like Microstructure on Titanium Dental Implant Surface Fabricated via Anodization Enhanced Cell Adhesion and Migration of Human Gingival Fibroblasts.

Background: Titanium (Ti) surface with nanotubes array via anodization has been used in dental implants to enhance bone regeneration but little research was carried out to evaluate whether the presence of highly ordered or disorderly distributed nanotubes array on titanium surface would have an effect on cell behaviors of gingival fibroblasts. Methods: The present study fabricated nanotubes arrays with varied topography under different constant voltage of electrochemical anodization in fluorine-containing electrolyte. Human gingival fibroblasts (HGFs) from extracted third molar were harvested and co-cultured with titanium disks with different nanotubes topography. Then cell behaviors of gingival fibroblasts including cell proliferation, adhesive morphology and cell migration were estimated to investigate the influence of titanium nanotubes on cell biology. Besides, gene and protein expression of adhesion molecule (integrin ß1/ß4/α6, fibronectin, intracellular adhesion molecule-1 and collagen type I) were detected to evaluate the influence of different surfaces on cell adhesion. Results: Highly ordered arrays of nanotubes with pore diameter of 60 nm and 100 nm were fabricated under 30 and 40 V of anodization (TNT-30 and TNT-40) while disorderedly distributed nanotube arrays formed on the titanium surface under 50 V of anodization (TNT-50). Our results demonstrated that compared with raw titanium surface and disorderly nanotubes, surface with orderly nanotubes array increased cell area and aspect ratio, as well as cell migration ability in the early phase of cell adhesion (p<0.05). Besides, compared with raw titanium surface, gene and protein expression of adhesion molecules were upregulated in nanotubes groups to different extents, no matter whether in an orderly or disorderly array. Conclusion: Within the limitations of our study, we conclude that compared with raw titanium surface, the presence of nanotubes array on titanium surface could enhance cells adhesion and cell migration in the early phase. And compared with disorderly distributed nanotubes, highly ordered nanotubes array might provide a much more favorable surface for gingival fibroblasts to achieve a tight adhesion on the materials.

Circ_0027446 promotes malignant development of glioblastoma by interacting with miR-346 to up-regulate PGK1.

Circular RNAs (circRNAs) can play essential roles in tumor development, including glioblastoma (GBM). The current study was performed to explore the function and mechanism of circ_0027446 in GBM progression. Circ_0027446, microRNA-346 (miR-346) and Phosphoglycerate kinase 1 (PGK1) levels were detected using reverse transcription-quantitative polymerase chain reaction assay. Cell behaviors were examined using Cell Counting Kit-8 assay, colony formation assay, EdU assay, flow cytometry, and transwell assay. Glycolytic metabolism was analyzed by commercial kits. The protein level was determined via western blot. The target interaction was analyzed by dual-luciferase reporter assay. Circ_0027446 function in vivo was explored by tumor xenograft assay. Circ_0027446 expression was significantly up-regulated in GBM samples and cells. Circ_0027446 down-regulation suppressed proliferation, invasion, glycolytic metabolism and enhanced apoptosis of GBM cells. MiR-346 was a target of circ_0027446, and circ_0027446 promoted GBM progression by sponging miR-346. PGK1 acted as a target gene of miR-346, and circ_0027446 interacted with miR-346 to regulate PGK1 expression. Overexpression of miR-346 inhibited malignant behaviors of GBM cells through down-regulating PGK1. Circ_0027446 contributed to tumor growth in vivo via miR-346/PGK1 axis. The current evidences demonstrated that circ_0027446 facilitated malignant progression of GBM through binding to miR-346 to up-regulate PGK1.

Methods for studying mammalian aquaporin biology.

Aquaporins (AQPs), transmembrane water-conducting channels, have earned a great deal of scrutiny for their critical physiological roles in healthy and disease cell states, especially in the biomedical field. Numerous methods have been implemented to elucidate the involvement of AQP-mediated water transport and downstream signaling activation in eliciting whole cell, tissue, and organ functional responses. To modulate these responses, other methods have been employed to investigate AQP druggability. This review discusses standard in vitro, in vivo, and in silico methods for studying AQPs, especially for biomedical and mammalian cell biology applications. We also propose some new techniques and approaches for future AQP research to address current gaps in methodology.

On-chip modeling of tumor evolution: Advances, challenges and opportunities.

Tumor evolution is the accumulation of various tumor cell behaviors from tumorigenesis to tumor metastasis and is regulated by the tumor microenvironment (TME). However, the mechanism of solid tumor progression has not been completely elucidated, and thus, the development of tumor therapy is still limited. Recently, Tumor chips constructed by culturing tumor cells and stromal cells on microfluidic chips have demonstrated great potential in modeling solid tumors and visualizing tumor cell behaviors to exploit tumor progression. Herein, we review the methods of developing engineered solid tumors on microfluidic chips in terms of tumor types, cell resources and patterns, the extracellular matrix and the components of the TME, and summarize the recent advances of microfluidic chips in demonstrating tumor cell behaviors, including proliferation, epithelial-to-mesenchymal transition, migration, intravasation, extravasation and immune escape of tumor cells. We also outline the combination of tumor organoids and microfluidic chips to elaborate tumor organoid-on-a-chip platforms, as well as the practical limitations that must be overcome.

Who's really in charge: Diverse follower cell behaviors in collective cell migration.

Collective cell migrations drive morphogenesis, wound healing, and cancer dissemination. Cells located at the front are considered leaders while those behind them are defined topologically as followers. Leader cell behaviors, including chemotaxis and their coupling to followers, have been well-studied and reviewed. However, the contributions of follower cells to collective cell migration represent an emerging area of interest. In this perspective, we highlight recent research into the broadening array of follower cell behaviors found in moving collectives. We describe examples of follower cells that possess cryptic leadership potential and followers that lack that potential but contribute in diverse and sometimes surprising ways to collective movement, even steering from behind. We highlight collectives in which all cells both lead and follow, and a few passive passengers. The molecular mechanisms controlling follower cell function and behavior are just emerging and represent an exciting frontier in collective cell migration research.

Crisscross multilayering of cell sheets.

Hydrostatic skeletons such as the Hydra's consist of two stacked layers of muscle cells perpendicularly oriented. In vivo, these bilayers first assemble, and then the muscle fibers of both layers develop and organize with this crisscross orientation. In the present work, we identify an alternative mechanism of crisscross bilayering of myoblasts in vitro, which results from the prior local organization of these active cells in the initial monolayer. The myoblast sheet can be described as a contractile active nematic in which, as expected, most of the +1/2 topological defects associated with this nematic order self-propel. However, as a result of the production of extracellular matrix (ECM) by the cells, a subpopulation of these comet-like defects does not show any self-propulsion. Perpendicular bilayering occurs at these stationary defects. Cells located at the head of these defects converge toward their core where they accumulate until they start migrating on top of the tail of the first layer, while the tail cells migrate in the opposite direction under the head. Since the cells keep their initial orientations, the two stacked layers end up perpendicularly oriented. This concerted process leading to a crisscross bilayering is mediated by the secretion of ECM.

Application of functionalized hyaluronic acid hydrogel with the activity of regulating the behaviors of stem cells in repairing rabbit knee articular cartilage.

Using injectable hydrogels loaded with mesenchymal stem cells (MSCs) to repair chondral defects is a new trend of cartilage tissue engineering in recent years. In this study, hyaluronic acid (HA) hydrogels containing the system of sustained-release Kartogenin (KGN) and modified by RGD and HAV peptides were used to facilitate repair of cartilage defect in the knee joint of rabbits. Different groups of implants were injected into osteochondral defects, and samples were taken 4 weeks after operation. Through the qualitative and quantitative analysis of Micro-CT, it can be seen that both FH (unloaded cell group) and R + FH groups (allogeneic cell group) can repair osteochondral defects well, and the amount of bone formation is high, which is close to the intact cartilage groups. Macroscopic observation and histological staining analysis showed that except for the intact cartilage group, FH group obtained the highest score. The morphology of the cartilage tissue in the FH groups was more regular and continuous than that in R + FH and H + FH (xenogeneic cell group) groups, approaching that of native cartilage. Immunohistochemical staining of Collagen II (Col II) showed that the expression and morphology of Col II in FH groups were similar to those in intact cartilage tissue. Interestingly, through in vivo experiments, this functionalized hyaluronic acid hydrogel can effectively promote the rapid repair of rabbit knee cartilage defects within one month.

Vascular cells responses to controlled surface structure and properties of bacterial nanocellulose artificial blood vessel after mercerization.

Therapeutic benefits of small caliber artificial blood vessels to cure cardio and cerebrovascular diseases are mainly limited by their low patency during long-term transplantation. Bacterial nanocellulose (BNC), as a natural polysaccharide mainly synthesized by a bacterium Komagataeibatacter xylinus, has shown great potential in small-caliber vascular graft applications due to its shape controllability, and furthermore its physical surface structure can be adjusted with different treatments. However, influences of physical surface structure and properties of BNC conduits on behaviors of vascular cells have not been investigated. In this work, mercerized BNC conduits (MBNC) with different surface roughness and stiffness were constructed by controlled alkali (NaOH) treatment. The changes of surface structures and properties significantly affected the behaviors of vascular cells and gene expression; meanwhile, the cell seeding density also affected the cell responses. After mercerization with NaOH concentration > 10 %, it was observed that the increased stiffness of MBNC decreased several functional gene expressions of human vascular endothelial cells, and the pathological transformation of smooth muscle cells was inhibited. This study demonstrates physical surface structure of MBNC conduits will critically regulate functions and behaviors of vascular cells and it also provides important designing parameters to improve the long-term patency of BNC-based conduits.

Cell Behavioral Dynamics as a Cue in Optimizing Culture Stabilization in the Bioprocessing of Pluripotent Stem Cells.

Pluripotent stem cells (PSCs) are important for future regenerative medicine therapies. However, in the production of PSCs and derivatives, the control of culture-induced fluctuations in the outcome of cell quality remains challenging. A detailed mechanistic understanding of how PSC behaviors are altered in response to biomechanical microenvironments within a culture is necessary for rational bioprocessing optimization. In this review, we discuss recent insights into the role of cell behavioral and mechanical homeostasis in modulating the states and functions of PSCs during culture processes. We delineate promising ways to manipulate the culture variability through regulating cell behaviors using currently developed tools. Furthermore, we anticipate their potential implementation for designing a culture strategy based on the concept of Waddington's epigenetic landscape that may provide a feasible solution for tuning the culture quality and stability in the bioprocessing space.

Recent Advances in Macroporous Hydrogels for Cell Behavior and Tissue Engineering.

Hydrogels have been extensively used as scaffolds in tissue engineering for cell adhesion, proliferation, migration, and differentiation because of their high-water content and biocompatibility similarity to the extracellular matrix. However, submicron or nanosized pore networks within hydrogels severely limit cell survival and tissue regeneration. In recent years, the application of macroporous hydrogels in tissue engineering has received considerable attention. The macroporous structure not only facilitates nutrient transportation and metabolite discharge but also provides more space for cell behavior and tissue formation. Several strategies for creating and functionalizing macroporous hydrogels have been reported. This review began with an overview of the advantages and challenges of macroporous hydrogels in the regulation of cellular behavior. In addition, advanced methods for the preparation of macroporous hydrogels to modulate cellular behavior were discussed. Finally, future research in related fields was discussed.

Engineering Extracellular Microenvironment for Tissue Regeneration.

The extracellular microenvironment is a highly dynamic network of biophysical and biochemical elements, which surrounds cells and transmits molecular signals. Extracellular microenvironment controls are of crucial importance for the ability to direct cell behavior and tissue regeneration. In this review, we focus on the different components of the extracellular microenvironment, such as extracellular matrix (ECM), extracellular vesicles (EVs) and growth factors (GFs), and introduce engineering approaches for these components, which can be used to achieve a higher degree of control over cellular activities and behaviors for tissue regeneration. Furthermore, we review the technologies established to engineer native-mimicking artificial components of the extracellular microenvironment for improved regenerative applications. This review presents a thorough analysis of the current research in extracellular microenvironment engineering and monitoring, which will facilitate the development of innovative tissue engineering strategies by utilizing different components of the extracellular microenvironment for regenerative medicine in the future.

MiR-193a-5p serves as an inhibitor in ovarian cancer cells through RAB11A.

Ovarian cancer is often not diagnosed until it is in advanced stages and its cure rate is relatively low. Thus, this investigation is concerned about the pathogenesis of this cancer. Differential expression analysis was undertaken on messenger RNA (mRNA) and microRNA (miRNA) data of ovarian cancer in Gene Expression Profiling Interactive Analysis and Gene Expression Omnibus databases. RAB11A mRNA and miR-193a-5p expression levels were tested by quantitative polymerase chain reaction. The targeting relationship between RAB11A and miR-193a-5p was verified by dual-luciferase assay. Cell behaviors of ovarian cancer were tested by Cell-Counting-Kit-8, colony formation and transwell assays. Expression of RAB11A protein and the proteins associated with Wnt/ß-catenin was tested by western blot. RAB11A high expression and miR-193a-5p low expression were found in ovarian cancer cells. RAB11A was targeted by miR-193a-5p. Cellular function experiments proved that RAB11A facilitated Wnt/ß-catenin signaling activation and deteriorated ovarian cancer progression. Rescue experiments exhibited two results: miR-193a-5p hindered proliferation, migration and invasion of ovarian cancer cells, and this suppression was counteracted by overexpression of RAB11A and miR-193a-5p. Furthermore, miR-193a-5p repressed RAB11A-mediated Wnt/ß-catenin activation. Altogether, miR-193a-5p served as a modulator in ovarian cancer cells via targeting RAB11A.

Regenerative Potential of Solid Bone Marrow Aspirate Concentrate Compared with Platelet-Rich Fibrin.

Platelet-rich fibrin (PRF) prepared from venous blood is used in the clinic to improve soft tissue wound healing. Nevertheless, arterial blood or bone marrow aspirate might also be a candidate for the source of PRF-like concentrates. The purpose of the present study was to investigate blood/bone marrow aspirate concentrates obtained from arterial blood, venous blood, and bone marrow aspirate to determine its respective regenerative potential in vitro. Arterial blood-derived PRF (Ar-PRF), venous blood-derived PRF (Ve-PRF), and solid-type bone marrow aspirate concentrate (sBMAC) were prepared from New Zealand white rabbits. Each clot was evaluated for its cytocompatibility and regenerative potential on primary rabbit gingival fibroblasts and osteoblasts. Both gingival fibroblasts and osteoblasts treated with each concentrate showed excellent viability. Interestingly, the sBMAC-treated cells demonstrated a significantly greater migratory potential than the other treatment groups. Furthermore, higher mRNA levels of transforming growth factor-beta, vascular endothelial growth factor, and collagen 1 (COL1) in gingival fibroblasts were observed in the sBMAC group compared with the Ar-PRF and Ve-PRF groups. Greater osteoblast differentiation potential, including higher osteocalcin (OCN) expression and mineralization potential, was found in osteoblasts treated with sBMAC. However, minor differences between the behaviors of cells treated with Ar-PRF and Ve-PRF were observed. In conclusion, sBMAC might be a new candidate for promoting wound healing and bone regeneration. Further preclinical and clinical experiments are necessary to prove the regenerative potential of sBMAC in the body. Impact Statement Blood concentrate material such as platelet-rich plasma or platelet-rich fibrin (PRF) is used in clinical practice to promote tissue regeneration in the field of dentistry, orthopedic surgery, and plastic surgery. The present study introduces a new type of solid bone marrow aspirate concentrate material and, for the first time, shows its excellent regenerative potential in both gingival fibroblasts and osteoblasts in vitro compared with that of conventional PRF.

Continuous live imaging reveals a subtle pathological alteration with cell behaviors in congenital heart malformation.

To form fully functional four-chambered structure, mammalian heart development undergoes a transient finger-shaped trabeculae, crucial for efficient contraction and exchange for gas and nutrient. Although its developmental origin and direct relevance to congenital heart disease has been studied extensively, the time-resolved cellular mechanism underlying hypotrabeculation remains elusive. Here, we employed in toto live imaging and reconstructed the holistic cell lineages and cellular behavior landscape of control and hypotrabeculed hearts of mouse embryos from E9.5 for up to 24 h. Compared to control, hypotrabeculation in ErbB2 mutants arose mainly through dual mechanisms: both reduced proliferation of trabecular cardiomyocytes from early cell fate segregation and markedly impaired oriented cell division and migration. Further examination of mosaic mutant hearts confirmed alterations in cellular behaviors in a cell autonomous manner. Thus, our work offers a framework for continuous live imaging and digital cell lineage analysis to better understand subtle pathological alterations in congenital heart disease.

Conjunction of gallium doping and calcium silicate mediates osteoblastic and osteoclastic performances of tricalcium phosphate bioceramics.

Gallium-containing biomaterials are considered promising for reconstructing osteoporotic bone defects, owing to the potent effect of gallium on restraining osteoclast activities. Nevertheless, the gallium-containing biomaterials were demonstrated to disturb the osteoblast activities. In this study, tricalcium phosphate (TCP) bioceramics were modified by gallium doping in conjunction with incorporation of calcium silicate (CS). The results indicated that the incorporation of CS promoted transition ofß-TCP toα-TCP, and accelerated densification process, but did not improve the mechanical strength of bioceramics. The silicon released from the composite bioceramics diminished the inhibition effect of released gallium on osteoblast activities, and maintained its effect on restraining osteoclast activities. The TCP-based bioceramics doped with 2.5 mol% gallium and incorporated with 10 mol% CS are considered suitable for treating the bone defects in the osteoporotic environment.

PSPH induces cell autophagy and promotes cell proliferation and invasion in the hepatocellular carcinoma cell line Huh7 via the AMPK/mTOR/ULK1 signaling pathway.

Phosphoserine phosphatase (PSPH), a key enzyme of the l-serine synthesis pathway, has been involved in cancer progression and survival. However, limited evidence revealed the PSPH influence on hepatocellular carcinoma (HCC). Herein, we observed that PSPH expression was upregulated in both HCC tissues and cell lines, which was determined by western blotting. TCGA database showed that the PSPH protein levels were significantly upregulated and affected patient survival rates in HCC. Then gain- and loss-of-function manipulations were performed by transfection with a pcDNA-PSPH expression vector or a specific short interfering RNA against PSPH in Huh7 cells. Huh7 cell proliferation, stemness, invasion, and apoptosis were assessed by using CCK-8 test, colony formation assay, Transwell assay, and Flow cytometry analysis, respectively, and levels of autophagy-related proteins were detected by using western blotting. The results showed that PSPH could induce Huh7 cell autophagy, promote cell proliferation and invasion, and inhibit apoptosis. The knockdown of PSPH could inhibit Huh7 cell proliferation, invasion, and autophagy. Furthermore, PSPH activated Liver kinase B1 (LKB1) and TGF beta-activated kinase 1 (TAK1), affected the adenosine 5'-monophosphate-activated protein kinase (AMPK)/mTOR/ULK1 signaling pathway, but could not activate calcium/calmodulin-dependent protein kinase kinase (CaMKK) in Huh7 cells. Inhibition of either LKB1, TAK1, or AMPK could eliminate the effect of PSPH overexpression on Huh7 cell behaviors. However, inhibition of CaMKK could not influence the effect of PSPH overexpression on Huh7 cell behaviors. In conclusion, PSPH could induce autophagy, promote proliferation and invasion, and inhibit apoptosis in HCC cells via the AMPK/mTOR/ULK1 signaling pathway.

Down-Regulation of AHNAK2 Inhibits Cell Proliferation, Migration and Invasion Through Inactivating the MAPK Pathway in Lung Adenocarcinoma.

AHNAK nucleoprotein 2 (AHNAK2) has been emerged as a crucial protein for neuroblast differentiation and cell migration, thereby involving in the development of various cancers. However, the specific molecular mechanism of AHNAK2 in lung adenocarcinoma is inconclusive. By accessing to the Oncomine dataset and GEPIA website, a higher expression level of AHNAK2 was observed in lung adenocarcinoma tissue samples. Overall survival (OS) curve plotted by Kaplan-Meier method showed that up-regulation of AHNAK2 was related with poor prognosis of lung adenocarcinoma patients. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis and western blot were conducted to examine the expression level of genes in lung adenocarcinoma cells. Through functional in vitro experiments, cell proliferation, migration and invasion were all suppressed after AHNAK2 knockdown using Cell counting kit-8 (CCK-8) assay, wound-healing and transwell analysis. Reduction of AHNAK2 decreased the apoptosis rate using flow cytometry analysis. Moreover, the key markers of MAPK pathway, p-MEK, p-ERK and p-P90RSK were decreased due to the transfection of si-AHNAK2 in A549 cells. U0126, a MEK inhibitor, showed the similar effects on MAPK-related protein levels with si-AHNAK2. To sum up, AHNAK2 is significantly increased in lung adenocarcinoma and plays a carcinogenic role by activating the MAPK signaling pathway, providing a novel insight and raising possibility for lung adenocarcinoma treatment.

Identifying disease-associated signaling pathways through a novel effector gene analysis.

BACKGROUND: Signaling pathway analysis methods are commonly used to explain biological behaviors of disease cells. Effector genes typically decide functional attributes (associated with biological behaviors of disease cells) by abnormal signals they received. The signals that the effector genes receive can be quite different in normal vs. disease conditions. However, most of current signaling pathway analysis methods do not take these signal variations into consideration. METHODS: In this study, we developed a novel signaling pathway analysis method called signaling pathway functional attributes analysis (SPFA) method. This method analyzes the signal variations that effector genes received between two conditions (normal and disease) in different signaling pathways. RESULTS: We compared the SPFA method to seven other methods across 33 Gene Expression Omnibus datasets using three measurements: the median rank of target pathways, the median p-value of target pathways, and the percentages of significant pathways. The results confirmed that SPFA was the top-ranking method in terms of median rank of target pathways and the fourth best method in terms of median p-value of target pathways. SPFA's percentage of significant pathways was modest, indicating a good false positive rate and false negative rate. Overall, SPFA was comparable to the other methods. Our results also suggested that the signal variations calculated by SPFA could help identify abnormal functional attributes and parts of pathways. The SPFA R code and functions can be accessed at https://github.com/ZhenshenBao/SPFA.

Traditional Chinese Medicine Xiaoai Jiedu Recipe Suppresses the Development of Hepatocellular Carcinoma via Regulating the microRNA-29a/Signal Transducer and Activator of Transcription 3 Axis.

OBJECTIVE: Hepatocellular carcinoma (HCC) is one of the most frequent and lethal tumors affecting human health worldwide. The aim of this study was to investigate the anti-cancer effects of Xiaoai Jiedu Recipe (XJR) on HCC development and its underlying mechanisms. METHODS: The expression of microRNA-29a (miR-29a) and signal transducer and activator of transcription 3 (STAT3) in HCC tissues and cells was determined by quantitative real-time polymerase chain reaction. The proliferation, migration, and invasion of HCC cells were measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide, wound-healing, and transwell assays, respectively. The regulatory relationship between miR-29a and STAT3 in HCC was predicted by TargetScan and analyzed by luciferase reporter and RNA pull-down assays. The protein expression of matrix metalloproteinase (MMP)-2/9 and STAT3 was detected by Western blotting. A xenograft tumor mouse model was established, and tumor weight and volume were measured. RESULTS: The expression of miR-29a was significantly decreased in HCC tissues and cells compared with that in normal tissues and cells. The up-regulation of miR-29a was related with lymph node metastasis and tumor node metastasis stage. XJR treatment significantly increased the expression of miR-29a, decreased cell viability, migration, and invasion, and reduced the protein expression of MMP-2/9 in HCC cells in a concentration-dependent manner. The anti-tumor effect of XJR on HCC cells was reversed by treatment with miR-29a inhibitor. STAT3 was predicted as a target of miR-29a, and its expression was negatively regulated by miR-29a. Moreover, STAT3 knockdown suppressed the malignant behavior of HCC cells, and its anti-tumor function was reversed by treatment with miR-29a inhibitor. Furthermore, XJR treatment inhibited tumor growth in mice through elevating miR-29a expression and inhibiting STAT3 expression. CONCLUSION: XJR suppressed the development of HCC via regulating miR-29a and STAT3.