Revealing role of epigenetic modifiers and DNA oxidation in cell-autonomous regulation of Cancer stem cells.

BACKGROUND: Breast cancer cells (BCCs) can remain undetected for decades in dormancy. These quiescent cells are similar to cancer stem cells (CSCs); hence their ability to initiate tertiary metastasis. Dormancy can be regulated by components of the tissue microenvironment such as bone marrow mesenchymal stem cells (MSCs) that release exosomes to dedifferentiate BCCs into CSCs. The exosomes cargo includes histone 3, lysine 4 (H3K4) methyltransferases - KMT2B and KMT2D. A less studied mechanism of CSC maintenance is the process of cell-autonomous regulation, leading us to examine the roles for KMT2B and KMT2D in sustaining CSCs, and their potential as drug targets. METHODS: Use of pharmacological inhibitor of H3K4 (WDR5-0103), knockdown (KD) of KMT2B or KMT2D in BCCs, real time PCR, western blot, response to chemotherapy, RNA-seq, and flow cytometry for circulating markers of CSCs and DNA hydroxylases in BC patients. In vivo studies using a dormancy model studied the effects of KMT2B/D to chemotherapy. RESULTS: H3K4 methyltransferases sustain cell autonomous regulation of CSCs, impart chemoresistance, maintain cycling quiescence, and reduce migration and proliferation of BCCs. In vivo studies validated KMT2's role in dormancy and identified these genes as potential drug targets. DNA methylase (DNMT), predicted within a network with KMT2 to regulate CSCs, was determined to sustain circulating CSC-like in the blood of patients. CONCLUSION: H3K4 methyltransferases and DNA methylation mediate cell autonomous regulation to sustain CSC. The findings provide crucial insights into epigenetic regulatory mechanisms underlying BC dormancy with KMT2B and KMT2D as potential therapeutic targets, along with standard care. Stem cell and epigenetic markers in circulating BCCs could monitor treatment response and this could be significant for long BC remission to partly address health disparity.

Oncobiology and treatment of breast cancer in young women.

Female breast cancer emerged as the leading cancer type in terms of incidence globally in 2020. Although mortality due to breast cancer has improved during the past three decades in many countries, this trend has reversed in women less than 40 years since the past decade. From the biological standpoint, there is consensus among experts regarding the clinically relevant definition of breast cancer in young women (BCYW), with an age cut-off of 40 years. The idea that breast cancer is an aging disease has apparently broken in the case of BCYW due to the young onset and an overall poor outcome of BCYW patients. In general, younger patients exhibit a worse prognosis than older pre- and postmenopausal patients due to the aggressive nature of cancer subtypes, a high percentage of cases with advanced stages at diagnosis, and a high risk of relapse and death in younger patients. Because of clinically and biologically unique features of BCYW, it is suspected to represent a distinct biologic entity. It is unclear why BCYW is more aggressive and has an inferior prognosis with factors that contribute to increased incidence. However, unique developmental features, adiposity and immune components of the mammary gland, hormonal interplay and crosstalk with growth factors, and a host of intrinsic and extrinsic risk factors and cellular regulatory interactions are considered to be the major contributing factors. In the present article, we discuss the status of BCYW oncobiology, therapeutic interventions and considerations, current limitations in fully understanding the basis and underlying cause(s) of BCYW, understudied areas of BCYW research, and postulated advances in the coming years for the field.

Mesenchymal stem cell secretome alters gene expression and upregulates motility of human endometrial stromal cells.

In brief: Endometrial stromal cell motility is fundamental to regeneration and repair of this tissue and crucial for successful reproduction. This paper shows a role for the mesenchymal stem cell (MSC) secretome in enhancing endometrial stromal cell motility. Abstract: Cyclic regeneration and repair of the endometrium are crucial for successful reproduction. Mesenchymal stem cells (MSCs) derived from bone marrow (BM-MSC) and umbilical cord (UC-MSC) facilitate tissue repair via their secretome, which contains growth factors and cytokines that promote wound healing. Despite the implication of MSCs in endometrial regeneration and repair, mechanisms remain unclear. This study tested the hypothesis that the BM-MSC and UC-MSC secretomes upregulate human endometrial stromal cell (HESC) proliferation, migration, and invasion and activate pathways to increase HESC motility. BM-MSCs were purchased from ATCC and cultured from the BM aspirate of three healthy female donors. UC-MSCs were cultured from umbilical cords of two healthy male term infants. Using indirect co-culture of MSCs and hTERT-immortalized HESCs via a transwell system, we demonstrated that co-culture of HESCs with BM-MSCs or UC-MSCs from all donors significantly increased HESC migration and invasion, whereas effects on HESC proliferation varied among BM-MSC and UC-MSC donors. Analysis of gene expression by mRNA sequencing and RT-qPCR showed that expression of CCL2 and HGF was upregulated in HESCs that had been cocultured with BM-MSCs or UC-MSCs. Validation studies revealed that exposure to recombinant CCL2 for 48 h significantly increased HESC migration and invasion. Increased HESC motility by the BM-MSC and UC-MSC secretome appears to be mediated in part by upregulated HESC CCL2 expression. Our data support the potential for leveraging MSC secretome as a novel cell-free therapy to treat disorders of endometrial regeneration.

Neuroimmune/Hematopoietic Axis with Distinct Regulation by the High-Mobility Group Box 1 in Association with Tachykinin Peptides.

Hematopoiesis is tightly regulated by the bone marrow (BM) niche. The niche is robust, allowing for the return of hematopoietic homeostasis after insults such as infection. Hematopoiesis is partly regulated by soluble factors, such as neuropeptides, substance P (SP), and neurokinin A (NK-A), which mediate hematopoietic stimulation and inhibition, respectively. SP and NK-A are derived from the Tac1 gene that is alternately spliced into four variants. The hematopoietic effects of SP and NK-A are mostly mediated via BM stroma. Array analyses with 2400 genes indicated distinct changes in SP-stimulated BM stroma. Computational analyses indicated networks of genes with hematopoietic regulation. Included among these networks is the high-mobility group box 1 gene (HMGB1), a nonhistone chromatin-associated protein. Validation studies indicated that NK-A could reverse SP-mediated HMGB1 decrease. Long-term culture-initiating cell assay, with or without NK-A receptor antagonist (NK2), showed a suppressive effect of HMGB1 on hematopoietic progenitors and increase in long-term culture-initiating cell assay cells (primitive hematopoietic cells). These effects occurred partly through NK-A. NSG mice with human hematopoietic system injected with the HMGB1 antagonist glycyrrhizin verified the in vitro effects of HMGB1. Although the effects on myeloid lineage were suppressed, the results suggested a more complex effect on the lymphoid lineage. Clonogenic assay for CFU- granulocyte-monocyte suggested that HMGB1 may be required to prevent hematopoietic stem cell exhaustion to ensure immune homeostasis. In summary, this study showed how HMGB1 is linked to SP and NK-A to protect the most primitive hematopoietic cell and also to maintain immune/hematopoietic homeostasis.

Mesenchymal stem cell therapies in brain disease.

As treatments for diseases throughout the body progress, treatment for many brain diseases has been at a standstill due to difficulties in drug delivery. While new drugs are being discovered in vitro, these therapies are often hindered by inefficient tissue distribution and, more commonly, an inability to cross the blood brain barrier. Mesenchymal stem cells are thus being investigated as a delivery tool to directly target therapies to the brain to treat wide array of brain diseases. This review discusses the use of mesenchymal stem cells in hypoxic disease (hypoxic ischemic encephalopathy), an inflammatory neurodegenerative disease (multiple sclerosis), and a malignant condition (glioma).

Decoding epigenetic cell signaling in neuronal differentiation.

Neurogenesis is the process by which new neurons are generated in the brain. Neural stem cells (NSCs) are differentiated into neurons, which are integrated into the neural network. Nowadays, pluripotent stem cells, multipotent stem cells, and induced pluripotent stem cells can be artificially differentiated into neurons utilizing several techniques. Specific transcriptional profiles from NSCs during differentiation are frequently used to approach and observe phenotype alteration and functional determination of neurons. In this context, the role of non-coding RNA, transcription factors and epigenetic changes in neuronal development and differentiation has gained importance. Epigenetic elucidation has become a field of intense research due to distinct patterns of normal conditions and different neurodegenerative disorders, which can be explored to develop new diagnostic methods or gene therapies. In this review, we discuss the complexity of transcription factors, non-coding RNAs, and extracellular vesicles that are responsible for guiding and coordinating neural development.

Epigenetic dynamics in cancer stem cell dormancy.

Cancer remains one of the most challenging diseases despite significant advances of early diagnosis and therapeutic treatments. Cancerous tumors are composed of various cell types including cancer stem cells capable of self-renewal, proliferation, differentiation, and invasion of distal tumor sites. Most notably, these cells can enter a dormant cellular state that is resistant to conventional therapies. Thereby, cancer stem cells have the intrinsic potential for tumor initiation, tumor growth, metastasis, and tumor relapse after therapy. Both genetic and epigenetic alterations are attributed to the formation of multiple tumor types. This review is focused on how epigenetic dynamics involving DNA methylation and DNA oxidations are implicated in breast cancer and glioblastoma multiforme. The emergence and progression of these cancer types rely on cancer stem cells with the capacity to enter quiescence also known as a dormant cellular state, which dictates the distinct tumorigenic aggressiveness between breast cancer and glioblastomas.

Exosomes in the Healthy and Malignant Bone Marrow Microenvironment.

The bone marrow (BM) is a complex organ that sustains hematopoiesis via mechanisms involving the microenvironment. The microenvironment includes several cell types, neurotransmitters from innervated fibers, growth factors, extracellular matrix proteins, and extracellular vesicles. The main function of the BM is to regulate hematopoietic function to sustain the production of blood and immune cells. However, the BM microenvironment can also accommodate the survival of malignant cells. A major mechanism by which the cancer cells communicate with cells of the BM microenvironment is through the exchange of exosomes, a subset of extracellular vesicles that deliver molecular signals bidirectionally between malignant and healthy cells. The field of exosomes is an active area of investigation since an understanding of how the exosomal packaging, cargo, and production can be leveraged therapeutically to deter cancer progression and sensitize malignant cells to other therapies. Altogether, this chapter discusses the crucial role of exosomes in the development and progression of BM-associated cancers, such as hematologic malignancies and marrow-metastatic breast cancer. Exosome-based therapeutic strategies and their limitations are also considered.

Therapeutic Potential of Mesenchymal Stem Cells in Immune-Mediated Diseases.

Mesenchymal stem cells (MSCs) are multipotent cells that can self-renew and differentiate into cells of all germ layers. MSCs can be easily attracted to the site of tissue insult with high levels of inflammatory mediators. The general ability of MSCs to migrate at the sites of tissue injury suggested an innate ability for these cells to be involved in baseline tissue repair. The bone marrow is one of the primary sources of MSCs, though they can be ubiquitous. An attractive property of MSCs for clinical application is their ability to cross allogeneic barrier. However, alone, MSCs are not immune suppressive cells. Rather, they can be licensed by the tissue microenvironment to become immune suppressor cells. Immune suppressor functions of MSCs include those that blunt cytotoxicity of natural killer cells, suppression of T-cell proliferation, and "veto" function. MSCs, as third-party cells, suppress the immune response that generally recapitulates graft-versus-host disease (GvHD) responses. Based on the plastic functions of MSCs, these cells have dominated the field of cell-based therapies, such as anti-inflammatory and drug delivery. Here, we focus on the potential use of MSC for immunological disorders such as Crohn's disease and GvHD.

Immune modulation by a cellular network of mesenchymal stem cells and breast cancer cell subsets: Implication for cancer therapy.

The immune modulatory properties of mesenchymal stem cells (MSCs) are mostly controlled by the particular microenvironment. Cancer stem cells (CSCs), which can initiate a clinical tumor, have been the subject of intense research. This review article discusses investigative studies of the roles of MSCs on cancer biology including on CSCs, and the potential as drug delivery to tumors. An understanding of how MSCs behave in the tumor microenvironment to facilitate the survival of tumor cells would be crucial to identify drug targets. More importantly, since CSCs survive for decades in dormancy for later resurgence, studies are presented to show how MSCs could be involved in maintaining dormancy. Although the mechanism by which CSCs survive is complex, this article focus on the cellular involvement of MSCs with regard to immune responses. We discuss the immunomodulatory mechanisms of MSC-CSC interaction in the context of therapeutic outcomes in oncology. We also discuss immunotherapy as a potential to circumventing this immune modulation.

A Novel Vaccine Targeting Glypican-3 as a Treatment for Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) has a high morbidity and mortality rate worldwide, with limited treatment options. Glypican-3 (GPC3) is a glycosylphosphatidylinositol-anchored glycoprotein that is overexpressed in most HCC tissues but not in normal tissues. GPC3-targeting antibody therapy shows limited response in a clinical trial due to the lack of a tumor-specific cytotoxic T lymphocyte (CTL) response. Here, in C57/B6 mice, we demonstrated that intravenous infusion of GPC3-coupled lymphocytes (LC/GPC3+) elicited robust GPC3-specific antibody and CTL responses, which effectively restricted proliferation and lysed cultured-HCC cells. Treatment with LC/GPC3+ induced durable tumor regression in HCC-bearing C57/B6 mice. Administration of LC/GPC3+ induced elevated levels of the cytotoxic T cell bioactive factors tumor necrosis factor alpha (TNF-α), interferon-γ (IFN-γ), granzyme B, and perforin, and substantially increased the number of infiltrating CD8+ T cells in tumor tissues. Moreover, immune responses elicited by LC/GPC3+ selectively suppressed GPC3+ tumors, but didn't affect the GPC3- tumors in BALB/c mice. Our findings provide the first preclinical evidence that intravenous infusion of the LC/GPC3+ complex can induce a strong anti-HCC effect through regulating systemic and local immune responses. These results indicate that the LC/GPC3+ complex could be developed as precision therapeutics for HCC patients in the future.

Human Aging and Cancer: Role of miRNA in Tumor Microenvironment.

Human aging is an inevitable and complex phenomenon characterized by a progressive, gradual degradation of physiological and cellular processes that leads from vulnerability to death. Mammalian somatic cells display limited proliferative properties in vitro that results in a process of permanent cell cycle arrest commonly known as senescence. Events leading to cellular senescence are complex but may be due to the increase in tumor suppressor genes, caused by lifetime somatic mutations. Cumulative mutation leaves an imprint on the genome of the cell, an important risk factor for the occurrence of cancer. Adults over the age of 65+ are vulnerable to age related diseases such as cancers but such changes may begin at middle age. MicroRNAs (miRNAs), which are small non-coding RNA, can regulate cancer progression, recurrence and metastasis. This chapter discusses the role of miRNA in tumor microenvironment, consequent to aging.

Mesenchymal stromal/stem cells in drug therapy: New perspective.

Mesenchymal stromal/stem cells (MSC) have emerged as a class of cells suitable for cellular delivery of nanoparticles, drugs and micro-RNA cargo for targeted treatments such as tumor and other protective mechanisms. The special properties of MSC underscore the current use for various clinical applications. Examples of applications include but are not limited to regenerative medicine, immune disorders and anti-cancer therapies. In recent years, there has been intense research in modifying MSC to achieve targeted and efficient clinical outcomes. This review discusses effects of MSC in an inflammatory microenvironment and then explains how these properties could be important to the overall application of MSC in cell therapy. The article also advises caution in the application of these cells because of their role in tumorigenesis. The review stresses the use of MSC as vehicles for drug delivery and discusses the accompanying challenges, based on the influence of the microenvironment on MSC.

The RNA-binding protein Musashi 1 stabilizes the oncotachykinin 1 mRNA in breast cancer cells to promote cell growth.

Substance P and its truncated receptor exert oncogenic effects. The high production of substance P in breast cancer cells (BCCs) is caused by the enhancement of tachykinin (TAC)1 translation by cytosolic factor. In vitro translational studies and mRNA stabilization analyses indicate that BCCs contain the factor needed to increase TAC1 translation and to stabilize the mRNA. Prediction of protein folding, RNA-shift analysis, and proteomic analysis identified a 40 kDa molecule that interacts with the noncoding exon 7. Western blot analysis and RNA supershift identified Musashi 1 (Msi1) as the binding protein. Ectopic expression of TAC1 in nontumorigenic breast cells (BCs) indicates that TAC1 regulates its stability by increasing Msi1. Using a reporter gene system, we showed that Msi1 competes with microRNA (miR)130a and -206 for the 3' UTR of exon 7/TAC1. In the absence of Msi1 and miR130a and -206, reporter gene activity decreased, indicating that Msi1 expression limits TAC1 expression. Tumor growth was significantly decreased when nude BALB/c mice were injected with Msi1-knockdown BCCs. In summary, the RNA-binding protein Msi1 competes with miR130a and -206 for interaction with TAC1 mRNA, to stabilize and increase its translation. Consequently, these interactions increase tumor growth.

Mesenchymal Stem Cell-Macrophage Crosstalk Provides Specific Exosomal Cargo to Direct Immune Response Licensing of Macrophages during Inflammatory Responses.

Neurodegenerative diseases (NDDs) continue to be a significant healthcare problem. The economic and social implications of NDDs increase with longevity. NDDs are linked to neuroinflammation and activated microglia and astrocytes play a central role. There is a growing interest for stem cell-based therapy to deliver genes, and for tissue regeneration. The promise of mesenchymal stem cells (MSC) is based on their availability as off-the-shelf source, and ease of expanding from discarded tissues. We tested the hypothesis that MSC have a major role of resetting activated microglial cells. We modeled microglial cell lines by using U937 cell-derived M1 and M2 macrophages. We studied macrophage types, alone, or in a non-contact culture with MSCs. MSCs induced significant release of exosomes from both types of macrophages, but significantly more of the M1 type. RNA sequencing showed enhanced gene expression within the exosomes with the major changes linked to the inflammatory response, including cytokines and the purinergic receptors. Computational analyses of the transcripts supported the expected effect of MSCs in suppressing the inflammatory response of M1 macrophages. The inflammatory cargo of M1 macrophage-derived exosomes revealed involvement of cytokines and purinergic receptors. At the same time, the exosomes from MSC-M2 macrophages were able to reset the classical M2 macrophages to more balanced inflammation. Interestingly, we excluded transfer of purinergic receptor transcripts from the co-cultured MSCs by analyzing these cells for the identified purinergic receptors. Since exosomes are intercellular communicators, these findings provide insights into how MSCs may modulate tissue regeneration and neuroinflammation.

Serum exosomal microRNA pathway activation in placenta accreta spectrum: pathophysiology and detection.

BACKGROUND: Placenta accreta spectrum disorders are a complex range of placental pathologies that are associated with significant maternal morbidity and mortality. A diagnosis of placenta accreta spectrum relies on ultrasonographic findings with modest positive predictive value. Exosomal microRNAs are small RNA molecules that reflect the cellular processes of the origin tissues. OBJECTIVE: We aimed to explore exosomal microRNA expression to understand placenta accreta spectrum pathology and clinical use for placenta accreta spectrum detection. STUDY DESIGN: This study was a biomarker analysis of prospectively collected samples at 2 academic institutions from 2011 to 2022. Plasma specimens were collected from patients with suspected placenta accreta spectrum, placenta previa, or repeat cesarean deliveries. Exosomes were quantified and characterized by nanoparticle tracking analysis and western blotting. MicroRNA were assessed by polymerase chain reaction array and targeted single quantification. MicroRNA pathway analysis was performed using the Ingenuity Pathway Analyses software. Placental biopsies were taken from all groups and analyzed by polymerase chain reaction and whole cell enzyme-linked immunosorbent assay. Receiver operating characteristic curve univariate analysis was performed for the use of microRNA in the prediction of placenta accreta spectrum. Clinically relevant outcomes were collected from abstracted medical records. RESULTS: Plasma specimens were analyzed from a total of 120 subjects (60 placenta accreta spectrum, 30 placenta previa, and 30 control). Isolated plasma exosomes had a mean size of 71.5 nm and were 10 times greater in placenta accreta spectrum specimens (20 vs 2 particles/frame). Protein expression of exosomes was positive for intracellular adhesion molecule 1, flotilin, annexin, and CD9. MicroRNA analysis showed increased detection of 3 microRNAs (mir-92, -103, and -192) in patients with placenta accreta spectrum. Pathway interaction assessment revealed differential regulation of p53 signaling in placenta accreta spectrum and of erythroblastic oncogene B2 or human epidermal growth factor 2 in control specimens. These findings were subsequently confirmed in placental protein analysis. Placental microRNA paralleled plasma exosomal microRNA expression. Biomarker assessment of placenta accreta spectrum signature microRNA had an area under the receiver operating characteristic curve of 0.81 (P<.001; 95% confidence interval, 0.73-0.89) with a sensitivity and specificity of 89.2% and 80%, respectively. CONCLUSION: In this large cohort, plasma exosomal microRNA assessment revealed differentially expressed pathways in placenta accreta spectrum, and these microRNAs are potential biomarkers for the detection of placenta accreta spectrum.

Experimental evidence for bone marrow as a source of nonhematopoietic endometrial stromal and epithelial compartment cells in a murine model.

Human endometrium has the remarkable ability to regenerate all cellular compartments with every menstrual cycle; the cellular source remains unknown. The objective of the present study was to determine whether the bone marrow (BM) is a source of multiple endometrial cell types using a murine BM transplant model. BM cells were harvested from transgenic donor mice that ubiquitously express green fluorescent protein (GFP) and were injected into lethally irradiated, syngeneic female recipient mice. Recipients with successful hematopoietic reconstitution were sacrificed at 3, 5, 9, and 12 mo posttransplant, after which hysterectomy was performed. Numbers of GFP-positive, CD45-positive, and CD45-negative cells in the endometrial stromal and epithelial compartments were determined. In the stromal compartment, BM-derived cells (BMDCs) were detectable as early as 3 mo posttransplant, and the BM remained a long-term contributor of nonhematopoietic endometrial cells. Nonhematopoietic endometrial cells comprised 47.3%-72.2% of total BMDCs in the stromal compartment at 12 mo posttransplant. In contrast, BMDCs were not detected in the glandular or luminal epithelial compartments until 12 mo posttransplant. These data demonstrate that the BM is a significant source of nonhematopoietic endometrial stromal compartment cells and contributes to a much lesser extent to the epithelial compartments. That BM is a source of nonhematopoietic cells in the endometrial stromal and epithelial compartments provides a potential mechanism by which monthly regeneration of the endometrium may occur.

An indirect role for oncomir-519b in the expression of truncated neurokinin-1 in breast cancer cells.

Neurokinin 1 (NK1) encodes full-length (NK1-FL) and truncated (NK1-Tr) receptors, with distinct 3' UTR. NK1-Tr exerts oncogenic functions and is increased in breast cancer (BC). Enhanced transcription of NK1 resulted in higher level of NK1-Tr. The 3' UTR of these two transcripts are distinct with NK1-Tr terminating at a premature stop codon. NK1-Tr mRNA gained an advantage over NK1-FL with regards to translation. This is due to the ability of miR519B to interact with sequences within the 3' UTR of NK1-FL, but not NK1-Tr since the corresponding region is omitted. MiR519b suppressed the translation of NK1-FL in T47D and MDA-MB-231 resulting in increased NK1-Tr protein. Cytokines can induce the transcription of NK1. However, our studies indicated that translation appeared to be independent of cytokine production by the BC cells (BCCs). This suggested that transcription and translation of NK1 might be independent. The findings were validated in vivo. MiR-519b suppressed the growth of MDA-MB-231 in 7/10 nude BALB/c. In total, increased NK1-Tr in BCCs is due to enhanced transcription and suppressed translation of NK1-FL by miR-519b to reduced tumor growth. In summary, we report on miRNA as a method to further regulate the expression of a spiced variant to promote oncogenesis. In addition, the findings have implications for therapy with NK1 antagonists. The oncogenic effect of NK1-Tr must be considered to improve the efficacy of current drugs to NK1.

Feline bone marrow-derived mesenchymal stromal cells (MSCs) show similar phenotype and functions with regards to neuronal differentiation as human MSCs.

Mesenchymal stromal cells (MSCs) show promise for treatment of a variety of neurological and other disorders. Cat has a high degree of linkage with the human genome and has been used as a model for analysis of neurological disorders such as stroke, Alzheimer's disease and motor disorders. The present study was designed to characterize bone marrow-derived MSCs from cats and to investigate the capacity to generate functional peptidergic neurons. MSCs were expanded with cells from the femurs of cats and then characterized by phenotype and function. Phenotypically, feline and human MSCs shared surface markers, and lacked hematopoietic markers, with similar morphology. As compared to a subset of human MSCs, feline MSCs showed no evidence of the major histocompatibility class II. Since the literature suggested Stro-1 as an indicator of pluripotency, we compared early and late passages feline MSCs and found its expression in >90% of the cells. However, the early passage cells showed two distinct populations of Stro-1-expressing cells. At passage 5, the MSCs were more homogeneous with regards to Stro-1 expression. The passage 5 MSCs differentiated to osteogenic and adipogenic cells, and generated neurons with electrophysiological properties. This correlated with the expression of mature neuronal markers with concomitant decrease in stem cell-associated genes. At day 12 induction, the cells were positive for MAP2, Neuronal Nuclei, tubulin ßIII, Tau and synaptophysin. This correlated with electrophysiological maturity as presented by excitatory postsynaptic potentials (EPSPs). The findings indicate that the cat may constitute a promising biomedical model for evaluation of novel therapies such as stem cell therapy in such neurological disorders as Alzheimer's disease and stroke.