Novel therapeutic activities of dragon blood from palm tree Daemonorops draco for the treatment of chronic diabetic wounds.

BACKGROUND: The clinical efficacy of Jinchuang Ointment, a traditional Chinese medicine (TCM), in treating chronic non-healing diabetic wounds has been demonstrated over the past decades. Both in vitro and in vivo angiogenic activities have been reported for its herbal ingredients, including dragon blood from the palm tree Daemonorops draco and catechu from Uncaria gambir Roxb. Additionally, crude extracts of dragon blood have exhibited hypoglycemic effects not only in animal studies but also in cell-based in vitro assays. RESULTS: Our findings indicate that crude dragon blood extract promotes the differentiation of myoblasts into myotubes. Partially purified fractions of dragon blood crude extract significantly enhance the expression of muscle cell differentiation-related genes such as myoG, myoD, and myoHC. Our results also demonstrate that crude extracts of dragon blood can inhibit platelet-derived growth factor-induced PAI-1 expression in primary rat vascular smooth muscle cells, thereby favoring changes in hemostasis towards fibrinolysis. Consistent with previous reports, reduced expression of plasminogen activator inhibitor 1 (PAI-1) accelerates wound healing. However, further separation resulted in a significant loss of both activities, indicating the involvement of more than one compound in these processes. Stem cells play a crucial role in muscle injury repair. Neither dragon blood nor catechu alone stimulated the proliferation of human telomerase reverse transcriptase (hTERT)-immortalized and umbilical cord mesenchymal stem cells. Interestingly, the proliferation of both types of stem cells was observed when crude extracts of dragon blood and catechu were present together in the stem cell growth medium. CONCLUSIONS: Dragon blood from D. draco offers multifaceted therapeutic benefits for treating chronic nonhealing diabetic wounds from various perspectives. Most drugs in Western medicine consist of small molecules with defined ingredients. However, this is not the case in TCM, as the activities of dragon blood reported in this study. Surprisingly, the activities documented here align with descriptions in ancient Chinese medical texts dating back to A.D. 1625.

The intestinal epithelial-macrophage-crypt stem cell axis plays a crucial role in regulating and maintaining intestinal homeostasis.

The intestinal tract plays a vital role in both digestion and immunity, making its equilibrium crucial for overall health. This equilibrium relies on the dynamic interplay among intestinal epithelial cells, macrophages, and crypt stem cells. Intestinal epithelial cells play a pivotal role in protecting and regulating the gut. They form vital barriers, modulate immune responses, and engage in pathogen defense and cytokine secretion. Moreover, they supervise the regulation of intestinal stem cells. Macrophages, serving as immune cells, actively influence the immune response through the phagocytosis of pathogens and the release of cytokines. They also contribute to regulating intestinal stem cells. Stem cells, known for their self-renewal and differentiation abilities, play a vital role in repairing damaged intestinal epithelium and maintaining homeostasis. Although research has primarily concentrated on the connections between epithelial and stem cells, interactions with macrophages have been less explored. This review aims to fill this gap by exploring the roles of the intestinal epithelial-macrophage-crypt stem cell axis in maintaining intestinal balance. It seeks to unravel the intricate dynamics and regulatory mechanisms among these essential players. A comprehensive understanding of these cell types' functions and interactions promises insights into intestinal homeostasis regulation. Moreover, it holds potential for innovative approaches to manage conditions like radiation-induced intestinal injury, inflammatory bowel disease, and related diseases.

Reversible Host-Guest Crosslinks in Supramolecular Hydrogels for On-Demand Mechanical Stimulation of Human Mesenchymal Stem Cells.

Stem cells are regulated not only by biochemical signals but also by biophysical properties of extracellular matrix (ECM). The ECM is constantly monitored and remodeled because the fate of stem cells can be misdirected when the mechanical interaction between cells and ECM is imbalanced. A well-defined ECM model for bone marrow-derived human mesenchymal stem cells (hMSCs) based on supramolecular hydrogels containing reversible host-guest crosslinks is fabricated. The stiffness (Young's modulus E) of the hydrogels can be switched reversibly by altering the concentration of non-cytotoxic, free guest molecules dissolved in the culture medium. Fine-adjustment of substrate stiffness enables the authors to determine the critical stiffness level E* at which hMSCs turn the mechano-sensory machinery on or off. Next, the substrate stiffness across E* is switched and the dynamic adaptation characteristics such as morphology, traction force, and YAP/TAZ signaling of hMSCs are monitored. These data demonstrate the instantaneous switching of traction force, which is followed by YAP/TAZ signaling and morphological adaptation. Periodical switching of the substrate stiffness across E* proves that frequent applications of mechanical stimuli drastically suppress hMSC proliferation. Mechanical stimulation across E* level using dynamic hydrogels is a promising strategy for the on-demand control of hMSC transcription and proliferation.

Spectroscopic techniques for monitoring stem cell and organoid proliferation in 3D environments for therapeutic development.

Spectroscopic techniques for monitoring stem cell and organoid proliferation have gained significant attention in therapeutic development. Spectroscopic techniques such as fluorescence, Raman spectroscopy, and infrared spectroscopy offer noninvasive and real-time monitoring of biochemical and biophysical changes that occur during stem cell and organoid proliferation. These techniques provide valuable insight into the underlying mechanisms of action of potential therapeutic agents, allowing for improved drug discovery and screening. This review highlights the importance of spectroscopic monitoring of stem cell and organoid proliferation and its potential impact on therapeutic development. Furthermore, this review discusses recent advances in spectroscopic techniques and their applications in stem cell and organoid research. Overall, this review emphasizes the importance of spectroscopic techniques as valuable tools for studying stem cell and organoid proliferation and their potential to revolutionize therapeutic development in the future.

Molecular hydrogen promotes wound healing by inducing early epidermal stem cell proliferation and extracellular matrix deposition.

BACKGROUND: Despite progress in developing wound care strategies, there is currently no treatment that promotes the self-tissue repair capabilities. H2 has been shown to effectively protect cells and tissues from oxidative and inflammatory damage. While comprehensive effects and how H2 functions in wound healing remains unknown, especially for the link between H2 and extracellular matrix (ECM) deposition and epidermal stem cells (EpSCs) activation. METHODS: Here, we established a cutaneous aseptic wound model and applied a high concentration of H2 (66% H2) in a treatment chamber. Molecular mechanisms and the effects of healing were evaluated by gene functional enrichment analysis, digital spatial profiler analysis, blood perfusion/oxygen detection assay, in vitro tube formation assay, enzyme-linked immunosorbent assay, immunofluorescent staining, non-targeted metabonomic analysis, flow cytometry, transmission electron microscope, and live-cell imaging. RESULTS: We revealed that a high concentration of H2 (66% H2) greatly increased the healing rate (3 times higher than the control group) on day 11 post-wounding. The effect was not dependent on O2 or anti-reactive oxygen species functions. Histological and cellular experiments proved the fast re-epithelialization in the H2 group. ECM components early (3 days post-wounding) deposition were found in the H2 group of the proximal wound, especially for the dermal col-I, epidermal col-III, and dermis-epidermis-junction col-XVII. H2 accelerated early autologous EpSCs proliferation (1-2 days in advance) and then differentiation into myoepithelial cells. These epidermal myoepithelial cells could further contribute to ECM deposition. Other beneficial outcomes include sustained moist healing, greater vascularization, less T-helper-1 and T-helper-17 cell-related systemic inflammation, and better tissue remodelling. CONCLUSION: We have discovered a novel pattern of wound healing induced by molecular hydrogen treatment. This is the first time to reveal the direct link between H2 and ECM deposition and EpSCs activation. These H2-induced multiple advantages in healing may be related to the enhancement of cell viability in various cells and the maintenance of mitochondrial functions at a basic level in the biological processes of life.

Mineralocorticoid receptor agonist aldosterone rescues hippocampal neural stem cell proliferation defects and improves postoperative cognitive function in aged mice.

OBJECTIVES: Hippocampal neurogenesis is closely related to learning and memory, and hippocampal neurogenesis disorders are involved in the development of many neurodegenerative diseases. Mineralocorticoid receptor (MR) plays a vital role in regulating stress response, neuroendocrine and cognitive functions, and is involved in regulating the integrity and stability of neural networks. However, the potential role of MR in the pathogenesis of postoperative cognitive dysfunction (POCD) is unclear. Therefore, this study evaluated the effect and mechanism of MR activation on postoperative hippocampal neurogenesis and cognitive function in aged mice. METHODS: 18-month-old male Kunming mice were randomly divided into Control group (C group), Surgery group (S group), Surgery+ Aldosterone group (S+Aldo group), Surgery + Wortmannin group (S+Wort group), Surgery + Aldosterone + Wortmannin group (S+Aldo+Wort group). Laparotomy was used to establish an animal model of postoperative cognitive dysfunction. After surgery, mice were intraperitoneally injected with aldosterone (100 ug/kg,150 ug/kg,200 ug/kg) and / or wortmannin (1 mg/kg); One day before the sacrifice, mice were injected intraperitoneally with BrdU (100 mg / kg / time, 3 times in total). Mice were subjected to Morris water maze and field tests at 1, 3, 7, and 14 days after surgery. Immunofluorescence was used to detect the number of BrdU +, Nestin +, BrdU/Nestin + positive cells in the hippocampal dentate gyrus of mice at 1, 3, 7 and 14 days after surgery. Western-blot was used to detect PI3K/Akt/GSK-3ß signaling pathway related proteins Akt, p-Akt, GSK-3ß, P-GSK-3ß expression. RESULTS: Stress impairs the performance of aged mice in water maze and open field tests, reduces the number of BrdU/Nestin+ cells in the hippocampal dentate gyrus, and inhibits the phosphorylation of Akt and GSK-3ß proteins in the hippocampus. Aldosterone treatment promotes P-Akt, P-GSK-3ß protein expression and hippocampal neural stem cell proliferation, and improves postoperative cognitive dysfunction. However, wortmannin treatment significantly reversed these effects of aldosterone. CONCLUSIONS: The mineralocorticoid receptor agonist aldosterone promotes the proliferation of hippocampal neural stem cells and improves cognitive dysfunction in aged mice after surgery, and the mechanism may be related to activation of PI3K/Akt/GSK-3ß signaling.

Bacterial Cellulose Composites with Polysaccharides Filled with Nanosized Cerium Oxide: Characterization and Cytocompatibility Assessment.

A new biocompatible nanocomposite film material for cell engineering and other biomedical applications has been prepared. It is based on the composition of natural polysaccharides filled with cerium oxide nanoparticles (CeONPs). The preparative procedure consists of successive impregnations of pressed bacterial cellulose (BC) with a sodium alginate (ALG) solution containing nanoparticles of citrate-stabilized cerium oxide and a chitosan (CS) solution. The presence of CeONPs in the polysaccharide composite matrix and the interaction of the nanoparticles with the polymer, confirmed by IR spectroscopy, change the network architecture of the composite. This leads to noticeable changes in a number of properties of the material in comparison with those of the matrix's polysaccharide composition, viz., an increase in mechanical stiffness, a decrease in the degree of planar orientation of BC macrochains, an increase in hydrophilicity, and the shift of the processes of thermo-oxidative destruction of the material to a low-temperature region. The latter effect is considered to be caused by the redox activity of cerium oxide (reversible transitions between the states Ce4+ and Ce3+) in thermally stimulated processes in the nanocomposite films. In the equilibrium swollen state, the material retains a mechanical strength at the level of ~2 MPa. The results of in vitro tests (cultivation of multipotent mesenchymal stem cells) have demonstrated the good biocompatibility of the BC-ALG(CeONP)-CS film as cell proliferation scaffolds.

Mechanical Stress Improves Fat Graft Survival by Promoting Adipose-Derived Stem Cells Proliferation.

Cell-assisted lipotransfer (CAL), defined as co-transplantation of aspirated fat with enrichment of adipose-derived stem cells (ASCs), is a novel technique for cosmetic and reconstructive surgery to overcome the low survival rate of traditional fat grafting. However, clinically approved techniques for increasing the potency of ASCs in CAL have not been developed yet. As a more clinically applicable method, we used mechanical stress to reinforce the potency of ASCs. Mechanical stress was applied to the inguinal fat pad by needling . Morphological and cellular changes in adipose tissues were examined by flow cytometric analysis 1, 3, 5, and 7 days after the procedure. The proliferation and adipogenesis potencies of ASCs were evaluated. CAL with ASCs treated with mechanical stress or sham control were performed, and engraftment was determined at 4 weeks post-operation. Flow cytometry analysis revealed that mechanical stress significantly increased the number as well as the frequency of ASC proliferation in fat. Proliferation assays and adipocyte-specific marker gene analysis revealed that mechanical stress promoted proliferation potential but did not affect the differentiation capacity of ASCs. Moreover, CAL with cells derived from mechanical stress-treated fat increased the engraftment. Our results indicate that mechanical stress may be a simple method for improving the efficacy of CAL by enhancing the proliferation potency of ASCs.

Low-energy Shockwave Therapy in the Management of Wound Healing Following Fournier's Gangrene.

We report on postoperative management of wound healing in four cases of Fournier's gangrene successfully treated with low-intensity shockwave therapy (LI-ESWT). In three cases, LI-ESWT (3 sessions per week with 2000 shockwaves at 3 Hz applied at 0.25 mJ/mm2) was able to close wound dehiscence secondary to plastic surgery with skin flaps. In one patient, LI-ESWT resulted in complete closure of an extensive wound with restoration of the local scrotal and penile skin. This is the first report of successful application of LI-ESWT for this indication. Restoration of local skin rather than wound closure by fibrous tissue could be related to promotion of stem cells, which has been discussed previously for other indications, such as treatment of chronic ulcers and restoration of the pelvic floor.

ClC-c regulates the proliferation of intestinal stem cells via the EGFR signalling pathway in Drosophila.

OBJECTIVES: Adult stem cells uphold a delicate balance between quiescent and active states, which is crucial for tissue homeostasis. Whereas many signalling pathways that regulate epithelial stem cells have been reported, many regulators remain unidentified. MATERIALS AND METHODS: Flies were used to generate tissue-specific gene knockdown and gene knockout. qRT-PCR was used to assess the relative mRNA levels. Immunofluorescence was used to determine protein localization and expression patterns. Clonal analyses were used to observe the phenotype. RNA-seq was used to screen downstream mechanisms. RESULTS: Here, we report a member of the chloride channel family, ClC-c, which is specifically expressed in Drosophila intestinal stem/progenitor cells and regulates intestinal stem cell (ISC) proliferation under physiological conditions and upon tissue damage. Mechanistically, we found that the ISC loss induced by the depletion of ClC-c in intestinal stem/progenitor cells is due to inhibition of the EGFR signalling pathway. CONCLUSION: Our findings reveal an ISC-specific function of ClC-c in regulating stem cell maintenance and proliferation, thereby providing new insights into the functional links among the chloride channel family, ISC proliferation and tissue homeostasis.

Visualization of individual cell division history in complex tissues using iCOUNT.

The division potential of individual stem cells and the molecular consequences of successive rounds of proliferation remain largely unknown. Here, we developed an inducible cell division counter (iCOUNT) that reports cell division events in human and mouse tissues in vitro and in vivo. Analyzing cell division histories of neural stem/progenitor cells (NSPCs) in the developing and adult brain, we show that iCOUNT can provide novel insights into stem cell behavior. Further, we use single-cell RNA sequencing (scRNA-seq) of iCOUNT-labeled NSPCs and their progenies from the developing mouse cortex and forebrain-regionalized human organoids to identify functionally relevant molecular pathways that are commonly regulated between mouse and human cells, depending on individual cell division histories. Thus, we developed a tool to characterize the molecular consequences of repeated cell divisions of stem cells that allows an analysis of the cellular principles underlying tissue formation, homeostasis, and repair.

Neurobiology of zinc and its role in neurogenesis.

BACKGROUND: Zinc (Zn) has a diverse role in many biological processes, such as growth, immunity, anti-oxidation system, homeostatic, and repairing. It acts as a regulatory and structural catalyst ion for activities of various proteins, enzymes, and signal transcription factors, as well as cell proliferation, differentiation, and survival. The Zn ion is essential for neuronal signaling and is mainly distributed within presynaptic vesicles. Zn modulates neuronal plasticity and synaptic activity in both neonatal and adult stages. Alterations in brain Zn status results in a dozen neurological diseases including impaired brain development. Numerous researchers are working on neurogenesis, however, there is a paucity of knowledge about neurogenesis, especially in neurogenesis in adults. Neurogenesis is a multifactorial process and is regulated by many metal ions (e.g. Fe, Cu, Zn, etc.). Among them, Zn has an essential role in neurogenesis. At the molecular level, Zn controls cell cycle, apoptosis, and binding of DNA and several proteins including transcriptional and translational factors. Zn is needed for protein folding and function and Zn acts as an anti-apoptotic agent; organelle stabilizer; and an anti-inflammatory agent. Zn deficiency results in aging, neurodegenerative disease, immune deficiency, abnormal growth, cancer, and other symptoms. Prenatal deficiency of Zn results in developmental disorders in humans and animals. CONCLUSION: Both in vitro and in vivo studies have shown an association between Zn deficiency and increased risk of neurological disorders. This article reviews the existing knowledge on the role of Zn and its importance in neurogenesis.

Chromatin Dynamics in Intestinal Epithelial Homeostasis: A Paradigm of Cell Fate Determination versus Cell Plasticity.

The rapid renewal of intestinal epithelium is mediated by a pool of stem cells, located at the bottom of crypts, giving rise to highly proliferative progenitor cells, which in turn differentiate during their migration along the villus. The equilibrium between renewal and differentiation is critical for establishment and maintenance of tissue homeostasis, and is regulated by signaling pathways (Wnt, Notch, Bmp…) and specific transcription factors (TCF4, CDX2…). Such regulation controls intestinal cell identities by modulating the cellular transcriptome. Recently, chromatin modification and dynamics have been identified as major actors linking signaling pathways and transcriptional regulation in the control of intestinal homeostasis. In this review, we synthesize the many facets of chromatin dynamics involved in controlling intestinal cell fate, such as stemness maintenance, progenitor identity, lineage choice and commitment, and terminal differentiation. In addition, we present recent data underlying the fundamental role of chromatin dynamics in intestinal cell plasticity. Indeed, this plasticity, which includes dedifferentiation processes or the response to environmental cues (like microbiota's presence or food ingestion), is central for the organ's physiology. Finally, we discuss the role of chromatin dynamics in the appearance and treatment of diseases caused by deficiencies in the aforementioned mechanisms, such as gastrointestinal cancer, inflammatory bowel disease or irritable bowel syndrome. Graphical abstract.

Tsc1 Regulates the Proliferation Capacity of Bone-Marrow Derived Mesenchymal Stem Cells.

TSC1 is a tumor suppressor that inhibits cell growth via negative regulation of the mammalian target of rapamycin complex (mTORC1). TSC1 mutations are associated with Tuberous Sclerosis Complex (TSC), characterized by multiple benign tumors of mesenchymal and epithelial origin. TSC1 modulates self-renewal and differentiation in hematopoietic stem cells; however, its effects on mesenchymal stem cells (MSCs) are unknown. We investigated the impact of Tsc1 inactivation in murine bone marrow (BM)-MSCs, using tissue-specific, transgelin (Tagln)-mediated cre-recombination, targeting both BM-MSCs and smooth muscle cells. Tsc1 mutants were viable, but homozygous inactivation led to a dwarfed appearance with TSC-like pathologies in multiple organs and reduced survival. In young (28 day old) mice, Tsc1 deficiency-induced significant cell expansion of non-hematopoietic BM in vivo, and MSC colony-forming potential in vitro, that was normalized upon treatment with the mTOR inhibitor, everolimus. The hyperproliferative BM-MSC phenotype was lost in aged (1.5 yr) mice, and Tsc1 inactivation was also accompanied by elevated ROS and increased senescence. ShRNA-mediated knockdown of Tsc1 in BM-MSCs replicated the hyperproliferative BM-MSC phenotype and led to impaired adipogenic and myogenic differentiation. Our data show that Tsc1 is a negative regulator of BM-MSC proliferation and support a pivotal role for the Tsc1-mTOR axis in the maintenance of the mesenchymal progenitor pool.

Linc-MYH configures INO80 to regulate muscle stem cell numbers and skeletal muscle hypertrophy.

Chromatin remodeling complexes have functions in transcriptional regulation and chromosome maintenance, but it is mostly unknown how the function of these normally ubiquitous complexes is specified in the cellular context. Here, we describe that the evolutionary conserved long non-coding RNA linc-MYH regulates the composition of the INO80 chromatin remodeler complex in muscle stem cells and prevents interaction with WDR5 and the transcription factor YY1. Linc-MYH acts as a selective molecular switch in trans that governs the pro-proliferative function of the ubiquitous INO80 complex but does not affect its role in maintaining genomic stability. The molecular switch is essential for restricting generation of quiescent MuSCs and proliferation of myoblasts in homeostasis and regeneration. Since linc-MYH is expressed in proliferating myoblasts but not in quiescent MuSCs, we reason that the extent of myoblast proliferation has decisive effects on the size of the quiescent MuSC pool.

Analysis of GPI-anchored proteins involved in germline stem cell proliferation in the Caenorhabditis elegans germline stem cell niche.

Stem cells divide and undergo self-renewal depending on the signals received from the stem cell niche. This phenomenon is indispensable to maintain tissues and organs in individuals. However, not all the molecular factors and mechanisms of self-renewal are known. In our previous study, we reported that glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) synthesized in the distal tip cells (DTCs; the stem cell niche) are essential for germline stem cell proliferation in Caenorhabditis elegans. Here, we characterized the GPI-APs required for proliferation. We selected and verified the candidate GPI-APs synthesized in DTCs by RNA interference screening and found that F57F4.3 (GFI-1), F57F4.4 and F54E2.1 are necessary for germline proliferation. These proteins are likely involved in the same pathway for proliferation and activated by the transcription factor PQM-1. We further provided evidence suggesting that these GPI-APs act through fatty acid remodelling of the GPI anchor, which is essential for association with lipid rafts. These findings demonstrated that GPI-APs, particularly F57F4.3/4 and F54E2.1, synthesized in the germline stem cell niche are located in lipid rafts and involved in promoting germline stem cell proliferation in C. elegans. The findings may thus shed light on the mechanisms by which GPI-APs regulate stem cell self-renewal.

Environmental Stimulation Counteracts the Suppressive Effects of Maternal High-Fructose Diet on Cell Proliferation and Neuronal Differentiation in the Dentate Gyrus of Adult Female Offspring via Histone Deacetylase 4.

Maternal high-fructose diets (HFD) impair the learning and memory capacity of adult female offspring via histone deacetylase 4 (HDAC4). Hippocampal adult neurogenesis is important for supporting the function of existing neural circuits. In this study, we investigated the effects of maternal HFD on hippocampal neural stem cell (NSC) proliferation and neuronal differentiation in adult offspring. Increased nuclear HDAC4 enzyme activity was detected in the hippocampus of HFD female offspring. The Western blot analyses indicated that the expressions of sex-determining region Y box2 (SOX2) and the transcription factor Paired Box 6 (PAX6), which are critical for the progression of NSC proliferation and differentiation, were downregulated. Concurrently, the expression of Ki67 (a cellular marker for proliferation) and doublecortin (DCX), which are related to NSC division and neuronal differentiation, was suppressed. Intracerebroventricular infusion with class II HDAC inhibitor (Mc1568, 4 weeks) led to the upregulation of these proteins. Environmental stimulation reversed the expression of Ki67 and DCX and the counts of Ki67- and DCX-positive cells in the hippocampi of HFD offspring as a result of providing the enriched housing for 4 weeks. Together, these results demonstrate that the suppressive effects of maternal HFD on hippocampal NSC proliferation and neuronal differentiation are reversibly mediated through HDAC4 and can be effectively reversed by environmental stimulation. The advantageous effects of environmental enrichment were possibly mediated by HDAC4 suppression.

[Penetrative needling of scalp-points improves neurological function by promoting stem cell proli-feration and activating Shh/Gil1 signaling in intracerebral hemorrhage rats].

OBJECTIVE: To observe the effect of penetrative needling from "Baihui" (GV20) to "Qubin" (GB7) on neural stem cell proliferation and sonic hedgehog (Shh) signaling in subventricular zone (SVZ) in intracerebral hemorrhage (ICH) rats so as to explore its mechanisms underlying improvement of ischemic injury of brain. METHODS: Male SD rats were randomly divided into blank control, model, acupuncture and agonist (Purmorphamine, an activator of Shh signaling pathway) groups (n＝18 in each group, 6 for H．E. stain, 6 for examining neuronal cell proliferation, and 6 for immunohistochemistry). The ICH model was established by injecting autogenous blood (50 µL) into the right caudate nucleus. The neurological defect was scored with refe-rence to Bederson's method. Penetrative needling from GV20 to GB7 was performed by manipulating the needle for 6 min (repeated 3 times in 30 min), once daily for 7 days. Intraperitoneal injection of Purmorphamine (1 mg/mL, 1 mg/kg) was performed, once daily for 7 days. Histopathological changes of the hemorrhagic penumbra region were observed under microscope after H．E. stain, the newborn neural stem cell proliferation (BrdU＋/Nestin＋ double labeled cells) in the SVZ was observed by immunofluorescence after intraperitoneal injection of BrdU (50 mg/kg), and the expression of Shh and glioma-associated hemolog-1 (Gli1) detected by immunohistochemistry. RESULTS: After modeling, the neurological score and expression levels of Shh and Gil1 proteins were significantly increased in the model group relevant to the blank control group (P<0.001). Following the interventions, the neurological score was evidently decreased (P<0.05), while the number of BrdU＋/Nestin＋ double labeled cells and the expression levels of Shh and Gil1 proteins were significantly increased in both acupuncture and agonist groups in comparison with the model group (P<0.001). No significant differences were found between the acupuncture and agonist groups in down-regulating the neurological score and in up-regulating the number of BrdU＋/Nestin＋ double labeled cells and the expression of Shh and Gil1 proteins (P>0.05). Outcomes of H．E. stain showed obvious edema, disordered arrangement of cells, infiltration of inflammatory cells and red blood cells with glial cell hyperplasia around the hematoma area in the model group, which was relatively milder in both acupuncture and agonist groups such as in basic disappearance of edema and inflammatory reaction. CONCLUSION: Penetrative needling from GV20 to GB7 can obviously improve neurological function in ICH rats, which is related to its effects in activating Shh/Gil1 signaling and in further promoting neural stem cell proliferation in the SVZ region.

Synthesis and formation mechanism of bone mineral, whitlockite nanocrystals in tri-solvent system.

Whitlockite (WH, Ca18Mg2(HPO4)2(PO4)12) is the second most abundant bone mineral and has attracted attention as one of the novel bone regenerative materials. It has proven to enhance growth and promote osteogenesis of stem cells. However, investigating the mechanism of formation of pure phase WH nanocrystals remains a challenge. In this study, we introduced an interesting synthesis approach of WH nanocrystals using a tri-solvent system for the solid-liquid-solution (SLS) process. The ratio of precursor and reaction solvent composition was optimized to generate WH nanocrystals with tunable size, morphology (nanoplates, nanospheres), and surface properties (hydrophobic, hydrophilic), which is impossible to achieve using the traditional precipitation method. Molecular dynamics (MD) simulations revealed that the growth direction of nanoplates is highly related to the surfactant and its binding affinity. Finite element method (FEM) simulations elucidated that the ratio of ethanol/water plays an important role in defining the crystallinity and morphology of WH. In this study, we demonstrated that the cell proliferation of human bone marrow-derived mesenchymal stem cells (hBMSCs) is enhanced upon treatment with WH. The results of quantitative real-time polymerase chain reaction (qPCR) revealed that WH can positively accelerate the osteogenic differentiation in hBMSCs. The as-synthesized WH has a great potential in the future to be used in osteogenic tissue engineering. This study opens a new horizon for the synthesis and application of WH.

Fibroblast growth factor-2 stimulates proliferation of human adipose-derived stem cells via Src activation.

BACKGROUND: Human adipose-derived stem cells (hASCs) are a subset of mesenchymal stem cells (MSCs); it has been regarded as one of the most promising stem cells. We previously found that fibroblast growth factor-2 (FGF-2) enhanced the proliferation and differentiation of hASC. However, the mechanisms involved in the growth of hASCs by FGF-2 have not been investigated. METHODS: Human adipose-derived stem cells (hASCs) were cultured with FGF-2, and cell growth was assessed. Effects of FGF Receptor (FGFR) inhibitor (NVP-BGJ398), ERK1/2 inhibitor (PD98059), PI3K/Akt inhibitor (LY294002), JNK inhibitor (SP600125), and p38 MAPK inhibitor (SB203580) and Src inhibitor (PP1) on the proliferation were investigated. At the same time, we assessed the effect of FGFR inhibitor on several signaling enzymes such as ERK1/2, JNK, p38, and Akt, in protein level. The involvement of Src activation by FGF-2 was also examined. RESULTS: FGF-2 markedly promoted proliferation of hASCs at concentrations lower than 10 ng/ml and stimulated cell progression to the S and G2/M phases. Proliferation was blocked by the FGFR inhibitor (NVP-BGJ398) and various signaling pathway inhibitors, such as Erk1/2 inhibitor (PD98059), PI3K/Akt inhibitor (LY294002), JNK inhibitor (SP600125), and p38MAPK inhibitor (SB203580). The FGFR inhibitor reduced the activation of protein kinases, such as AKT, Erk1/2, JNK, and p38, in several signaling pathways. The downstream kinase of FGFR, Src, was activated by FGF-2, and its activation was canceled by the FGFR inhibitor. MEK1/2, a downstream kinase of Src, was parallelly regulated by FGF-2. The Src inhibitor (PP1) markedly blocked the proliferation of hASCs via inhibition of Src and MEK1/2. CONCLUSION: Src activation is indispensable for FGF-2-mediated proliferation of ASCs, as well as the subsequent activation of multi-signaling pathways.