Neuroprotective effects of mesenchymal stromal cells in mouse models of Alzheimer's Disease: The Mediating role of gut microbes and their metabolites via the Microbiome-Gut-Brain axis.

The intricacy and multifaceted nature of Alzheimer's disease (AD) necessitate therapies that target multiple aspects of the disease. Mesenchymal stromal cells (MSCs) emerge as potential agents to mitigate AD symptoms; however, whether their therapeutic efficacy involves modulation of gut microbiota and the microbiome-gut-brain axis (MGBA) remains unexplored. In this study, we evaluated the effects of three distinct MSCs types-derived from the umbilical cord (UCMSC), dental pulp (SHED), and adipose tissue (ADSC)-in an APP/PS1 mouse model of AD. In comparison to saline control, MSCs administration resulted in a significant reduction of behavioral disturbances, amyloid plaques, and phosphorylated tau in the hippocampus and frontal cortex, accompanied by an increase in neuronal count and Nissl body density across AD-afflicted brain regions. Through 16S rRNA gene sequencing, we identified partial restoration of gut microbial balance in AD mice post-MSCs treatment, evidenced by the elevation of neuroprotective Akkermansia and reduction of the AD-associated Sphingomonas. To examine whether gut microbiota involved in MSCs efficacy in treating AD, SHED with better anti-inflammatory and gut microbiota recovery effects among three MSCs, and another AD model 5 × FAD mice with earlier and more pathological proteins in brain than APP/PS1, were selected for further studies. Antibiotic-mediated gut microbial inactivation attenuated MSCs efficacy in 5 × FAD mice, implicating the involvement of gut microbiota in the therapeutic mechanism. Functional analysis of altered gut microbiota and targeted bile acid metabolism profiling revealed a significant enhancement in bile acid variety following MSCs therapy. A chief bile acid constituent, taurocholic acid (TCA), was orally administered to AD mice and similarly abated AD symptoms. Nonetheless, the disruption of intestinal neuronal integrity with enterotoxin abrogated the ameliorative impact of both MSCs and TCA treatments. Collectively, our findings substantiate that MSCs confer therapeutic benefits in AD within a paradigm that primarily involves regulation of gut microbiota and their metabolites through the MGBA.

Transplantation of feces from mice with Alzheimer's disease promoted lung cancer growth.

BACKGROUND: Alzheimer's disease (AD) is a progressive neurologic disorder that causes the brain to shrink and brain cells to die. Lung cancer is characterized by high morbidity and mortality, late diagnosis and poor prognosis. And there is no specific mechanism to explain the epidemiological correlation between AD and lung cancer. MATERIALS AND METHODS: Lewis lung cancer cells (LLC) were injected into the left forelimb armpit of APP/PS1 mice to establish a tumor-bearing model. After remodeling the gut microbiota by fecal microbiota transplantation (FMT), the tumor were collected and analyzed for tumor size, Western blotting, and 16S rRNA gene sequencing. RESULTS: Compared with the control group, the AD FMT group showed larger tumors, while C57 FMT group showed smaller tumors. The former group showed the inhibition of AKT/Bax/Bcl-2 pathway, while the latter showed promotion of Caspase-1/IL-1ß and AKT/Bax/Bcl-2 pathway, which induced changes in tumor size. And Prevotella, Prevotella, Mucispirillum and Halomonas in the gut lumen of LLC tumor-bearing mice are increased, and Bacteroides, Coprobacillus, Bifidobacterium, Faecalibacterium and Aggregatiacter are decreased significantly. CONCLUSION: AD and lung cancer showed a positive correlation in APP expression, which proposed a different view from epidemiology on the correlation between AD and lung cancer.

Human gingival mesenchymal stem cells improve movement disorders and tyrosine hydroxylase neuronal damage in Parkinson disease rats.

BACKGROUND AIMS: Gingival mesenchymal stem cells (GMSCs) demonstrate high proliferation, trilineage differentiation and immunomodulatory properties. Parkinson disease (PD) is the second most common type of neurodegenerative disease. This study aimed to explore the effect and mechanism of GMSC-based therapy in 6-hydroxydopamine-induced PD rats. METHODS: RNA sequencing and quantitative proteomics technology was used to validate the neuroprotective role of GMSCs therapeutic in 6-Hydroxydopamine -induced PD model in vitro and in vivo. Western blotting, immunofluorescence and real-time quantitative PCR verified the molecular mechanism of GMSCs treatment. RESULTS: Intravenous injection of GMSCs improved rotation and forelimb misalignment behavior, enhanced the anti-apoptotic B-cell lymphoma 2/B-cell lymphoma 2-associated X axis, protected tyrosine hydroxylase neurons, decreased the activation of astrocytes and reduced the astrocyte marker glial fibrillary acidic protein and microglia marker ionized calcium-binding adaptor molecule 1 in the substantia nigra and striatum of PD rats. The authors found that GMSCs upregulated nerve regeneration-related molecules and inhibited metabolic disorders and the activation of signal transducer and activator of transcription 3. GMSCs showed a strong ability to protect neurons and reduce mitochondrial membrane potential damage and reactive oxygen species accumulation. The safety of GMSC transplantation was confirmed by the lack of tumor formation following subcutaneous transplantation into nude mice for up to 8 weeks. CONCLUSIONS: The authors' research helps to explain the mechanism of GMSC-based therapeutic strategies and promote potential clinical application in Parkinson disease.

Metformin pre-treatment of stem cells from human exfoliated deciduous teeth promotes migration and angiogenesis of human umbilical vein endothelial cells for tissue engineering.

BACKGROUND AIMS: Stem cells from human exfoliated deciduous teeth (SHED) play a significant role in tissue engineering and regenerative medicine. Angiogenesis is crucial in tissue regeneration and a primary target of regenerative medicine. As a first-line anti-diabetic drug, metformin demonstrates numerous valuable impacts on stem cells. This study aimed to explore metformin's impact and mechanism of action on SHED-mediated angiogenesis. METHODS: First, cell proliferation; flow cytometry; osteogenic, adipogenic and chondrogenic induction; and proteomics analyses were conducted to explore the role of metformin in SHED. Subsequently, migration and tube formation assays were used to evaluate chemotaxis and angiogenesis enhancement by SHED pre-treated with metformin under co-culture conditions in vitro, and relative messenger RNA expression levels were determined by quantitative reverse transcription polymerase chain reaction. Finally, nude mice were used for in vivo tube formation assay, and sections were analyzed through immunohistochemistry staining with anti-human CD31 antibody. RESULTS: Metformin significantly promoted SHED proliferation as well as osteogenic, adipogenic and chondrogenic differentiation. Proteomics showed that metformin significantly upregulated 124 differentially abundant proteins involved in intracellular processes, including various proteins involved in cell migration and angiogenesis, such as MAPK1. The co-culture system demonstrated that SHED pre-treated with metformin significantly improved the migration and angiogenesis of human umbilical vein endothelial cells. In addition, SHED pre-treated with metformin possessed greater ability to promote angiogenesis in vivo. CONCLUSIONS: In summary, the authors' findings illustrate metformin's mechanism of action on SHED and confirm that SHED pre-treated with metformin exhibits a strong capacity for promoting angiogenesis. This helps in promoting the application of dental pulp-derived stem cells pre-treated with metformin in regeneration engineering.

Chitosan Hydrogel Supplemented with Metformin Promotes Neuron-like Cell Differentiation of Gingival Mesenchymal Stem Cells.

Human gingival mesenchymal stem cells (GMSCs) are derived from migratory neural crest stem cells and have the potential to differentiate into neurons. Metformin can inhibit stem-cell aging and promotes the regeneration and development of neurons. In this study, we investigated the potential of metformin as an enhancer on neuronal differentiation of GMSCs in the growth environment of chitosan hydrogel. The crosslinked chitosan/ß-glycerophosphate hydrogel can form a perforated microporous structure that is suitable for cell growth and channels to transport water and macromolecules. GMSCs have powerful osteogenic, adipogenic and chondrogenic abilities in the induction medium supplemented with metformin. After induction in an induction medium supplemented with metformin, Western blot and immunofluorescence results showed that GMSCs differentiated into neuron-like cells with a significantly enhanced expression of neuro-related markers, including Nestin (NES) and ß-Tubulin (TUJ1). Proteomics was used to construct protein profiles in neural differentiation, and the results showed that chitosan hydrogels containing metformin promoted the upregulation of neural regeneration-related proteins, including ATP5F1, ATP5J, NADH dehydrogenase (ubiquinone) Fe-S protein 3 (NDUFS3), and Glutamate Dehydrogenase 1 (GLUD1). Our results help to promote the clinical application of stem-cell neural regeneration.

Advances in Human Cellular Mechanistic Understanding and Drug Discovery of Brain Organoids for Neurodegenerative Diseases.

The prevalence of neurodegenerative diseases (NDs) is increasing rapidly as the aging population accelerates, and there are still no treatments to halt or reverse the progression of these diseases. While traditional 2D cultures and animal models fail to translate into effective therapies benefit patients, 3D cultured human brain organoids (hBOs) facilitate the use of non-invasive methods to capture patient data. The purpose of this study was to review the research and application of hBO in disease models and drug screening in NDs. The pluripotent stem cells are induced in multiple stages to form cerebral organoids, brain region-specific organoids and their derived brain cells, which exhibit complex brain-like structures and perform electrophysiological activities. The brain region-specific organoids and their derived neurons or glial cells contribute to the understanding of the pathogenesis of NDs and the efficient development of drugs, including Alzheimer's disease, Parkinson's disease, Huntington's disease and Amyotrophic lateral sclerosis. Glial-rich brain organoids facilitate the study of glial function and neuroinflammation, including astrocytes, microglia, and oligodendrocytes. Further research on the maturation enhancement, vascularization and multi-organoid assembly of hBO will help to enhance the research and application of NDs cellular models.

Implementation of blood-brain barrier on microfluidic chip: Recent advance and future prospects.

The complex structure of the blood-brain barrier (BBB) hinders its modeling and the treatment of brain diseases. The microfluidic technology promotes the development of BBB-on-a-chip platforms, which can be used to reproduce the complex brain microenvironment and physiological reactions. Compared with traditional transwell technology, microfluidic BBB-on-a-chip shows great technical advantages in terms of flexible control of fluid shear stress in the chip and fabrication efficiency of the chip system, which can be enhanced by the development of lithography and three-dimensional (3D) printing. It is convenient to accurately monitor the dynamic changes of biochemical parameters of individual cells in the model by integrating an automatic super-resolution imaging sensing platform. In addition, biomaterials, especially hydrogels and conductive polymers, solve the limitations of microfluidic BBB-on-a-chip by compounding onto microfluidic chip to provide a 3D space and special performance on the microfluidic chip. The microfluidic BBB-on-a-chip promotes the development of basic research, including cell migration, mechanism exploration of neurodegenerative diseases, drug barrier permeability, SARS-CoV-2 pathology. This study summarizes the recent advances, challenges and future prospects of microfluidic BBB-on-a-chip, which can help to promote the development of personalized medicine and drug discovery.

Potential of astrocytes in targeting therapy for Alzheimer's disease.

As the most common neurodegenerative disease, Alzheimer's disease (AD) exhibits an incomprehensible pathogenesis, which has led to the continuous failure of drug development in recent years. Although neuronal damage is considered a pathological feature of AD, treatment strategies targeting ß-amyloid (Aß) have not achieved beneficial effects. In-depth research on glial cells has revealed the strong importance and application prospects of astrocytes in the recovery of cognitive functions. This review summarizes the role of astrocytes in AD and the possibility of therapeutic strategies targeting astrocytes. Astrocytes are involved in brain lipid metabolism and can regulate the synthesis and degradation of Aß to affect the pathology of AD. The tau protein is phosphorylated by astrocytes, and this phosphorylation leads to the formation of neurofibrillary tangles (NFTs). Astrocytes can express a variety of receptors and inflammatory factors and participate in the neuroinflammatory process and the release of proinflammatory mediators. When the glutamate produced by the neurons is not cleared by astrocytes, neurons undergo apoptosis due to blocked cell metabolism. Therapies for astrocytes are highly efficient, and these include stem cell therapy, gene editing technology, astrocyte transformation and chemical drugs. Here, we discuss the advantages and disadvantages of animal and cell models applied to the study of targeted astrocyte therapies. This study helps elucidate the mechanism of astrocytes in AD and promotes the clinical application of potential therapeutic strategies targeting astrocytes.

Targeting small heat shock proteins to degrade aggregates as a potential strategy in neurodegenerative diseases.

Neurodegenerative diseases (NDs) are aging-related diseases that involve the death of neurons in the brain. Dysregulation of protein homeostasis leads to the production of toxic proteins or the formation of aggregates, which is the pathological basis of NDs. Small heat shock proteins (HSPB) is involved in the establishment of a protein quality control (PQC) system to maintain cellular homeostasis. HSPB can be secreted into the extracellular space and delivered by various routes, especially extracellular vehicles (EVs). HSPB plays an important role in influencing the aggregation phase of toxic proteins involved in heat shock transcription factor (HSF) regulation, oxidative stress, autophagy and apoptosis pathways. HSPB conferred neuroprotective effects by resisting toxic protein aggregation, reducing autophagy and reducing neuronal apoptosis. The HSPB treatment strategies, including targeted PQC system therapy and delivery of EVs-HSPB, can improve disease manifestations for NDs. This review aims to provide a comprehensive insight into the impact of HSPB in NDs and the feasibility of new technology to enhance HSPB expression and EVs-HSPB delivery for neurodegenerative disease.

Composition of intestinal flora affects the risk relationship between Alzheimer's disease/Parkinson's disease and cancer.

An increasing number of epidemiological studies have shown that there is a significant inverse relationship between the onset of Alzheimer's disease/Parkinson's disease (AD/PD) and cancer, but the mechanism is still unclear. Considering that intestinal flora can connect them, we tried to explain this phenomenon from the intestinal flora. This review briefly introduced the relationship among AD/PD, cancer, and intestinal flora, studied metabolites or components of the intestinal flora and the role of intestinal barriers and intestinal hormones in AD/PD and cancer. After screening, a part of the flora capable of participating in the occurrence processes of the three diseases at the same time was obtained, the abundance changes of the special flora in AD/PD and various types of cancers were summarized, and they were classified according to the flora function and abundance, which in turn innovatively and reasonably explained the fact that AD/PD and cancer showed certain antagonism in epidemiological statistics from the perspective of intestinal flora. This review also proposed that viewing the risk relationship between diseases from the perspective of intestinal flora may provide new research ideas for the treatment of fecal microbiota transplantation (FMT) and related diseases.

Stem cells from human exfoliated deciduous teeth relieves Alzheimer's disease symptoms in SAMP8 mice by up-regulating the PPARγ pathway.

The pathology of Alzheimer's disease (AD) is complex and heterogeneous, and there are currently no drugs that can stop its progression. The failure of traditional chemical small-molecule drug development showed the weakness of single target and made researchers look to cell therapy with multiple regulatory effects. Stem cells from human exfoliated deciduous teeth (SHED) are a kind of neural crest-derived mesenchymal stem cells which have broad prospects in the treatment of neurodegenerative diseases. In this study, we demonstrated the therapeutic effects of SHED in AD mice, including behavioral improvement, neuronal protection, and alleviation of neuroinflammation. Tracking experiments on SHED showed that some of the transplanted cells could enter the brain. To elucidate the role played by the majority of cells transplanted into veins, blood proteomic assays were performed. Data are available via ProteomeXchange with identifier PXD030313. Among the altered proteins, the PPAR pathway related to energy metabolism was considered to be an important signaling pathway involved in regulation through gene ontology analysis and pathway analysis. Western blot showed that the transplantation of SHED improved the glucose metabolism in AD mice by increasing the PPARγ signaling pathway. These results suggested that SHED have a potential in relieving AD pathological symptoms and improving behavioral cognition. The therapeutic mechanism of SHED is related to up-regulating PPARγ signaling pathway and reducing neuronal damage.

Transfer of Tumor-Bearing Mice Intestinal Flora Can Ameliorate Cognition in Alzheimer's Disease Mice.

BACKGROUND: Fecal microbiota transplant (FMT) is a potential treatment approach for many diseases. Alzheimer's disease (AD) and cancer have been proven to have a specific antagonistic relationship to FMT. OBJECTIVE: This article aims to explore whether intestinal flora transplantation from cancer individuals can ameliorate cognitive impairment. METHODS: Morris water maze and object recognition tests were performed to assess cognitive function after the fecal flora from tumor-bearing and WT mice were transplanted into AD mice by gavage. The effect of flora transplantation on AD was analyzed by thioflavin T staining, western blot, and 16S RNA sequencing. RESULTS: AD mice with FMT significantly improved short-term memory level and cognitive ability compared with Tg + NaCl group. Inflammatory factors in the plasma were regulated, and Aß plaques burden in the hippocampus and cortex were decreased. FMT in the tumor-bearing group showed a higher significant amelioration in symptoms compared to the healthy group. 16S RNA sequencing revealed that FMT treatments could reverse the increased Firmicutes and Prevotella and the decreased Bacteroidetes, Bacteroides, and Sutterella in AD mice. AD mice transplanted with tumor-bearing mice feces additionally increased the density of Oscillospira, Odoribacter, and AF12. Furthermore, the predicted functional analyses showed that the metabolism of inorganic and organic salts in the intestinal flora of AD mice was also reversed by FMT. CONCLUSION: Intestinal flora transplantation from tumor-bearing mice can ameliorate the cognitive impairment of AD mice.

Combined Treatment of Tanshinone I and Epirubicin Revealed Enhanced Inhibition of Hepatocellular Carcinoma by Targeting PI3K/AKT/HIF-1α.

Background: Epirubicin (EADM) is a common chemotherapeutic agent in hepatocellular carcinoma (HCC). The accumulation of hypoxia-inducible factor-1α (HIF-1α) is an important cause of drug resistance to EADM in HCC. Tanshinone I (Tan I) is an agent with promising anti-cancer effects alone or with other drugs. Some tanshinones mediate HIF-1α regulation via PI3K/AKT. However, the role of Tan I combined with EADM to reduce the resistance of HCC to EADM has not been investigated. Therefore, this study aimed to investigate the combined use of Tan I and EADM in HCC and the underlying mechanism of PI3K/AKT/HIF-1α. Methods: HCC cells were treated with Tan I, EADM, or the combined treatment for 48 hrs. Cell transfection was used to construct HIF-1α overexpression HCC stable cells. Cell viability, colony formation, and flow cytometric assays were used to detect the viability, proliferation, and apoptosis in HCC cells. Synergism between Tan I and EADM were tested by calculating the Bliss synergy score, positive excess over bliss additivism (EOBA), and the combination index (CI). Western blotting analyses were used to detect the levels of ß-actin, HIF-1α, PI3K p110α, p-Akt Thr308, Cleaved Caspase-3, and Cleaved Caspase-9. Toxicity parameters were used to evaluate the safety of the combination in mice. The xenograft model of mice was built by HCC stable cell lines, which was administrated with Tan I, EADM, or a combination of them for 8 weeks. Immunohistochemistry staining (IHC) was used to assess tumor apoptosis in mouse models. Results: Hypoxia could upregulate HIF-1α to induce drug resistance in HCC cancer cells. The combination of Tan I and EADM was synergistic. Although Tan I or EADM alone could inhibit HCC cancer cells, the combination of them could further enhance the cytotoxicity and growth inhibition by targeting the PI3K/AKT/HIF-1α signaling pathway. Furthermore, Tan I and EADM synergistically reversed HIF-1α-mediated drug resistance to inhibit HCC. The results of toxicity parameters showed that the combination was safe in mice. Meanwhile, animal models showed that Tan I not only improved the anti-tumor effect of EADM, but also reduced the drug reactions of EADM-induced weight loss. Conclusion: Our results suggested that Tan I could effectively improve the anti-tumor effect of EADM, and synergize EADM to reverse HIF-1α mediated resistance via targeting PI3K/AKT/HIF-1α signaling pathway.

Metformin enhances the osteogenesis and angiogenesis of human umbilical cord mesenchymal stem cells for tissue regeneration engineering.

Human umbilical cord mesenchymal stem cells (hUC-MSCs) are a potential clinical material in regenerative medicine applications. Metformin has shown safety and effectiveness as a clinical drug. However, the effect of metformin as a treatment on hUC-MSCs is unclear. Our research aimed to explore the effects of metformin on the osteogenesis, adipogenesis and angiogenesis of hUC-MSCs, and attempted to explain the molecular fluctuations of metformin through the mapping of protein profiles. Proliferation assay, osteogenic and adipogenic differentiation induction, cell cycle, flow cytometry, quantitative proteomics techniques and bioinformatics analysis were used to detect the influences of metformin treatment on hUC-MSCs. Our results demonstrated that low concentrations of metformin promoted the proliferation of hUC-MSCs, but high concentrations of metformin inhibited it. Metformin exhibited promotion of osteogenesis but inhibition of adipogenesis. Metformin treated hUC-MSCs up-regulated the expression of osteogenic marker ALP, OCN and RUNX2, but down-regulated the expression of adipogenic markers PPARγ and LPL. Proteomics analysis found that up-regulation of differentially expressed proteins in metformin treatment group involved the biological process of cell migration in Gene Ontology analysis. Metformin enhanced cell migration of HUVEC in a co-culture system, and hUC-MSCs treated with metformin exhibited stronger angiogenesis in vitro and in vivo compared to the hUC-MSCs group. The results of RT-qPCR revealed that the SCF and VEGFR2 were raised in metformin treatment. This study can promote the application of hUC-MSCs treated with metformin to tissue engineering for vascular reconstruction and angiogenesis.