Human Amniotic Fluid Research Articles

In Utero Amniotic Fluid Stem Cell Therapy Protects Against Myelomeningocele via Spinal Cord Coverage and Hepatocyte Growth Factor Secretion.

Despite the poor prognosis associated with myelomeningocele (MMC), the options for prenatal treatments are still limited. Recently, fetal cellular therapy has become a new option for treating birth defects, although the therapeutic effects and mechanisms associated with such treatments remain unclear. The use of human amniotic fluid stem cells (hAFSCs) is ideal with respect to immunoreactivity and cell propagation. The prenatal diagnosis of MMC during early stages of pregnancy could allow for the ex vivo proliferation and modulation of autologous hAFSCs for use in utero stem cell therapy. Therefore, we investigated the therapeutic effects and mechanisms of hAFSCs‐based treatment for fetal MMC. hAFSCs were isolated as CD117‐positive cells from the amniotic fluid of 15‐ to 17‐week pregnant women who underwent amniocentesis for prenatal diagnosis and consented to this study. Rat dams were exposed to retinoic acid to induce fetal MMC and were subsequently injected with hAFSCs in each amniotic cavity. We measured the exposed area of the spinal cord and hepatocyte growth factor (HGF) levels at the lesion. The exposed spinal area of the hAFSC‐treated group was significantly smaller than that of the control group. Immunohistochemical analysis demonstrated a reduction in neuronal damage such as neurodegeneration and astrogliosis in the hAFSC‐treated group. Additionally, in lesions of the hAFSC‐treated group, HGF expression was upregulated and HGF‐positive hAFSCs were identified, suggesting that these cells migrated to the lesion and secreted HGF to suppress neuronal damage and induce neurogenesis. Therefore, in utero hAFSC therapy could become a novel strategy for fetal MMC. stem cells translational medicine 2019;8:1170–1179

Effect of ascorbic acid on differentiation of human amniotic fluid mesenchymal stem cells into cardiomyocyte-like cells

The aim of this study was to evaluate the efficiency of ascorbic acid (AA) on cell viability, cytotoxicity and the effects on cardiomyogenic differentiation of the human amniotic fluid mesenchymal stem cells (hAF-MSCs). The results of methylthiazole tetrazolium (MTT) assay and cell apoptosis assay indicated that after 24, 48 and 72 h of treatment, AA had no effect on cells viability and cytotoxicity. After treating the hAF-MSCs with 5-azacytidine (5-aza) and a combination of AA and 5-aza, the alamar blue cells proliferation assay showed the normal growth characteristic similar to control group. Especially, the morphological changes were observed between day 0 and day 21, and it was revealed that the hAF-MSCs exhibited myotube-like morphology after 7 days of cell culturing. Moreover, the treatment with a combination of AA and 5-aza was able to up-regulate the cardiomyogenic specific gene levels, which are known to play an important role in cardiomyogenesis. This was specifically notable with the results of immunofluorescence and immunoenzymatic staining in the AA combined with 5-aza treatment group, the highest expression of cardiomyogenic specific proteins was revealed including for GATA4, cTnT, Cx43 and Nkx2.5. It could be concluded that AA may be a good alternative cardiomyogenic inducing factor for hAF-MSCs and may open new insights into future biomedical applications for a clinically treatment.

Effect of Human Amniotic Fluid Stem Cells on Kidney Function in a Model of Chronic Kidney Disease.

Kidney disease is a major medical problem globally. Chronic kidney disease (CKD) is a progressive loss of kidney function. It causes accumulation of waste and fluid in the body, eventually resulting in kidney failure as well as damaging other organs. Although dialysis and kidney transplantation have been used as primary treatments for renal disease, dialysis does not restore full renal function, and there is a shortage of donor kidneys for transplantation. Recent advances in cell-based therapies have offered a means to augment and restore renal function. Various types of cells have been tested to evaluate their therapeutic effects on injured kidneys. Among various types of cells, amniotic fluid stem cells (AFSCs) share advantages of both embryonic and adult stem cells, such as pluripotent activity, remarkable plasticity, and immunomodulatory effects, which may allow their future therapeutic use as an "off-the-shelf" cell source. AFSC presents advantages of both conventional pluripotent and adult stem cells, such as pluripotent activity, remarkable plasticity, and immunomodulatory effects. This study demonstrates that administration of human-derived AFSC facilitates functional and structural improvement in a rat model of CKD, and suggests that cell therapy with AFSC has potential as a therapeutic strategy to recover renal function in patients with CKD. Impact Statement Patients with chronic kidney disease (CKD) have limited treatment options, and renal transplantation is the only definitive treatment method that restores kidney function. However, challenges associated with transplantation, including donor organ shortage, rejection, and life-long immunosuppression, remain a problem. Recently, stem cell-based therapies have been proposed as an alternative approach to augment and restore renal function. In this study, we used human-derived amniotic fluid stem cells (AFSCs) to treat CKD in a rat model and demonstrated that AFSC treatment facilitated positive effects in terms of improvements of renal function.

Human mid-trimester amniotic fluid (stem) cells lack expression of the pluripotency marker OCT4A

Expression of OCT4A is one of the hallmarks of pluripotency, defined as a stem cell’s ability to differentiate into all the lineages of the three germ layers. Despite being defined as non-tumorigenic cells with high translational potential, human mid-trimester amniotic fluid stem cells (hAFSCs) are often described as sharing features with embryonic stem cells, including the expression of OCT4A, which could hinder their clinical potential. To clarify the OCT4A status of hAFSCs, we first undertook a systematic review of the literature. We then performed extensive gene and protein expression analyses to discover that neither frozen, nor fresh hAFSCs cultivated in multipotent stem cell culture conditions expressed OCT4A, and that the OCT4A positive results from the literature are likely to be attributed to the expression of pseudogenes or other OCT4 variants. To address this issue, we provide a robust protocol for the assessment of OCT4A in other stem cells.

The human amniotic fluid mesenchymal stem cells therapy on, SKOV3, ovarian cancer cell line.

PurposeOne of the most common malignancies peculiar to female health with few symptoms, low response to therapy, difficult diagnosis, frequent relapse, and high mortality, is ovarian cancer. Thus, our experiment, using Human amniotic fluid mesenchymal stem cells (hAFMSCs) as a therapeutic tool, aims to find an efficient treatment approach for patients suffering from SKOV3 ovarian cancer.Material & MethodsIn this study, we obtained 5 ml amniotic fluid from 16–20 week pregnant women who underwent amniocentesis for routine prenatal diagnosis by karyotyping in Al‐Zahra Hospital of Tabriz University of Medical Sciences, Iran. Using trans wells in 24 wells plate, hAFMSCs were isolated from all samples, co‐cultured with SKOV3 ovarian cancer cell line, and characterized via flow cytometry and RT‐PCR. Human skin fibroblast cells (HSFCs) were isolated and used as a negative control. SKOV3 and HSFCs' viability after 5 days was evaluated by MTT assay. Cell cycle and apoptotic genes were analyzed by real‐time PCR.ResultsWe successfully isolated and characterized hAFMSCs through it positivity for CD44 and CD90 specific mesenchymal stem cell markers and negativity for CD31 and CD45. Oct4 and NANOG were evaluated by RT‐PCR as pluripotency markers, and visualized on 2% gel electrophoresis. We established hAFMS cell lines after 5 days of co‐culturing the SKOV3 cells, viability was decreased; however, HSFCs did not show toxicity by MTT assay. The genes indicated upregulation and high expression by a real‐time PCR.ConclusionsOur findings showed that hAFMSCs have natural tumor tropism, and can release soluble factors in a cell culture, which cause an efficient anticancer effect. Thus, we can use hAFMSCs for complete anticancer therapy on SKOV3 cell line at cell culture condition and possibly in vivo in the near future.

Amniotic Fluid Cells, Stem Cells, and p53: Can We Stereotype p53 Functions?

In recent years, great interest has been devoted to finding alternative sources for human stem cells which can be easily isolated, ideally without raising ethical objections. These stem cells should furthermore have a high proliferation rate and the ability to differentiate into all three germ layers. Amniotic fluid, ordinarily discarded as medical waste, is potentially such a novel source of stem cells, and these amniotic fluid derived stem cells are currently gaining a lot of attention. However, further information will be required about the properties of these cells before they can be used for therapeutic purposes. For example, the risk of tumor formation after cell transplantation needs to be explored. The tumor suppressor protein p53, well known for its activity in controlling Cell Prolif.eration and cell death in differentiated cells, has more recently been found to be also active in amniotic fluid stem cells. In this review, we summarize the major findings about human amniotic fluid stem cells since their discovery, followed by a brief overview of the important role played by p53 in embryonic and adult stem cells. In addition, we explore what is known about p53 in amniotic fluid stem cells to date, and emphasize the need to investigate its role, particularly in the context of cell tumorigenicity.

Human platelet lysate as an alternative to fetal bovine serum for culture and endothelial differentiation of human amniotic fluid mesenchymal stem cells.

Human amniotic fluid (hAF) mesenchymal stem cells (MSCs) are commonly cultured in medium containing FBS. However, there are concerns about using animal serum in therapeutic applications due to the potential for immunogenic reactions and the risk of transmission of pathogens. For safety reasons, human platelet lysate (hPL) has been suggested as a replacement for FBS because it appears to be a natural source of growth factors. In this present study, it was investigated whether FBS could be substituted with hPL in hAF-MSCs culture without affecting their properties. Pooled hPL was generated by the freeze-thaw method. The concentration of hPL was selected after evaluation by MTT assay. The hAF-MSCs were cultured in FBS- or hPL-supplemented conditions and shared a fibroblast-like morphology. Cell proliferation assays showed that the growth characteristic of hAF-MSCs cultured in 10% hPL-supplemented media was similar to those cultured in 10% FBS-supplemented media. The expression of MSC markers did not differ between the cells cultured in the different conditions. The endothelial differentiation potential was also investigated. Reverse transcription-quantitative (RT-q)PCR revealed that induced cells supplemented with hPL showed an increase level of endothelial specific gene expression compared to the FBS-supplemented cells. Immunofluorescence analysis showed specific protein localization in both induced cell groups. Additionally, induced cells supplemented with hPL had the potential to form networks on Matrigel. This present study indicated that hPL could be used to culture and enhance the endothelial differentiation potential of hAF-MSCs.

Reactivating endogenous mechanisms of cardiac regeneration via paracrine boosting using the human amniotic fluid stem cell secretome

BackgroundThe adult mammalian heart retains residual regenerative capability via endogenous cardiac progenitor cell (CPC) activation and cardiomyocyte proliferation. We previously reported the paracrine cardioprotective capacity of human amniotic fluid-derived stem cells (hAFS) following ischemia or cardiotoxicity. Here we analyse the potential of hAFS secretome fractions for cardiac regeneration and future clinical translation. MethodshAFS were isolated from amniotic fluid leftover samples from prenatal screening. hAFS conditioned medium (hAFS-CM) was obtained following hypoxic preconditioning. Anti-apoptotic, angiogenic and proliferative effects were evaluated on rodent neonatal cardiomyocytes (r/mNVCM), human endothelial colony forming cells (hECFC) and human CPC. Mice undergoing myocardial infarction (MI) were treated with hAFS-CM, hAFS-extracellular vesicles (hAFS-EV), or EV-depleted hAFS-CM (hAFS-DM) by single intra-myocardial administration and evaluated in the short and long term. ResultshAFS-CM improved mNVCM survival under oxidative and hypoxic damage, induced Ca2+-dependent angiogenesis in hECFC and triggered hCPC and rNVCM proliferation. hAFS-CM treatment after MI counteracted scarring, supported cardiac function, angiogenesis and cardiomyocyte cell cycle progression in the long term. hAFS-DM had no effect. hAFS-CM and hAFS-EV equally induced epicardium WT1+ CPC reactivation. Although no CPC cardiovascular differentiation was observed, our data suggests contribution to local angiogenesis by paracrine modulation. hAFS-EV alone were able to recapitulate all the beneficial effects exerted by hAFS-CM, except for stimulation of vessel formation. ConclusionshAFS-CM and hAFS-EV can improve cardiac repair and trigger cardiac regeneration via paracrine modulation of endogenous mechanisms. While both formulations are effective in sustaining myocardial renewal, hAFS-CM retains higher pro-angiogenic potential, while hAFS-EV particularly enhances cardiac function.

Condrogenesis of mesenchymal stem cells derived from human amniotic fluid in chitosan-xanthan scaffolds under TGF- beta 3 stimuli

Condrogenesis of mesenchymal stem cells derived from human amniotic fluid in chitosan-xanthan scaffolds under TGF- beta 3 stimuli

083 Critical Analysis of Human Amniotic Fluid as a Novel Biological Therapy

083 Critical Analysis of Human Amniotic Fluid as a Novel Biological Therapy

Evaluation of Percutaneous Intradiscal Amniotic Suspension Allograft in a Rabbit Model of Intervertebral Disc Degeneration.

A laboratory study using a rabbit annular puncture model of intervertebral disc degeneration (IDD). The aims of this study were to assess whether an amniotic suspension allograft (ASA) containing particulated human amnion and amniotic fluid derived cells regains intervertebral disc height and morphology and improves histologic scoring in a rabbit model of IDD. In contrast to current surgical interventions for IDD, in which the primary goal is to relieve symptomatic pain, one novel strategy involves the direct injection of anabolic cytokines. Current therapies for IDD are limited by both the short half-life of therapeutic proteins and general decline in anabolic cell populations. Intervertebral discs in New Zealand white rabbits were punctured using 18-gauge needle under fluoroscopic guidance. Four weeks post-puncture, two groups of rabbits were injected with either ASA or a vehicle/sham control, while a third group was untreated. Weekly radiographs were obtained for 12 weeks to assess disc height index (DHI). Magnetic resonance imaging (MRI) T2 relaxation time was evaluated at weeks 4 and 12 to assess morphological changes. Histologic sections were evaluated on a semi-quantitative grading scale. Before treatment at week 4, DHIs and normalized T2 relaxation times between the three groups were not significantly different. At week 12, ASA-treated rabbits exhibited significantly greater DHIs and MRI T2 relaxation times than vehicle and untreated control groups. The ASA group had higher mean histologic score than the vehicle group, which demonstrated extensive fiber disorganization and delamination with reduced proteoglycan staining on histology. Minimally invasive intervention with intradiscal injection of ASA was successful in reducing IDD in a reproducible rabbit model, with significant improvement in disc height and morphology when compared with vehicle and untreated control groups on radiographic and MRI analyses. N/A.

Stem Cells Transplantation Improves Kidney Function in Cardio-Renal Syndrome Type II: Insight into Pathophysiological Mechanisms

Purpose we investigated the effects of human amniotic fluid stem cells (hAFS) and rat adipose tissue stromal vascular fraction GFP-positive cells (rSVC-GFP) in a model of cardio-renal syndrome type II (CRSII). Methods RHF was induced by monocrotaline (MCT) in 28 Sprague-Dawley rats. Three weeks later, four million hAFS or rSVC-GFP cells were injected via tail vein. BNP, sCreatinine, kidney and heart NGAL and MMP9, sCytokines, kidney and heart apoptosis and cells (Cs) engraftment were evaluated. Results Cell-treated rats showed a significant reduction of serum NGAL and Creatinine compared to CRSII. In both hAFS and rSVC-GFP group, kidney protein expression of NGAL was significantly lower than in CRSII (hAFS p=0.036 and rSVC-GFP p Conclusion Cell therapy produced improvement in kidney function in rats with CRSII. This was the result of interstitial, vessel and tubular cell engraftment leading to tubular and vessel regeneration, decreased tubular cells apoptosis and mitigated pro-inflammatory milieu. Reduction of NGLA-MMP9 complexes mainly due to decrease MMPs activity prevented further negative heart remodelling.

The Unique Features of Proteins Depicting the Chicken Amniotic Fluid

In many amniotes, the amniotic fluid is depicted as a dynamic milieu that participates in the protection of the embryo (cushioning, hydration, and immunity). However, in birds, the protein profile of the amniotic fluid remains unexplored, even though its proteomic signature is predicted to differ compared with that of humans. In fact, unlike humans, chicken amniotic fluid does not collect excretory products and its protein composition strikingly changes at mid-development because of the massive inflow of egg white proteins, which are thereafter swallowed by the embryo to support its growth. Using GeLC-MS/MS and shotgun strategies, we identified 91 nonredundant proteins delineating the chicken amniotic fluid proteome at day 11 of development, before egg white transfer. These proteins were essentially associated with the metabolism of nutrients, immune response and developmental processes. Forty-eight proteins were common to both chicken and human amniotic fluids, including serum albumin, apolipoprotein A1 and alpha-fetoprotein. We further investigated the effective role of chicken amniotic fluid in innate defense and revealed that it exhibits significant antibacterial activity at day 11 of development. This antibacterial potential is drastically enhanced after egg white transfer, presumably due to lysozyme, avian beta-defensin 11, vitelline membrane outer layer protein 1, and beta-microseminoprotein-like as the most likely antibacterial candidates. Interestingly, several proteins recovered in the chicken amniotic fluid prior and after egg white transfer are uniquely found in birds (ovalbumin and related proteins X and Y, avian beta-defensin 11) or oviparous species (vitellogenins 1 and 2, riboflavin-binding protein). This study provides an integrative overview of the chicken amniotic fluid proteome and opens stimulating perspectives in deciphering the role of avian egg-specific proteins in embryonic development, including innate immunity. These proteins may constitute valuable biomarkers for poultry production to detect hazardous situations (stress, infection, etc.), that may negatively affect the development of the chicken embryo.

Processed human amniotic fluid retains its antibacterial activity

BackgroundHuman amniotic fluid (AF) contains numerous nutrients, trophic factors and defense proteins that provide a nurturing and protective environment for fetal development. Based on reports that AF has antibacterial, anti-inflammatory and regenerative properties, we designed a novel method to process AF for use in clinical care.MethodsSix randomly selected lots of processed AF (pAF) were examined to determine whether they retained their antibacterial activity against a panel of wound-associated pathogens E. faecium, S. aureus, K. pneumoniae, A. baumannii, P. aeruginosa, and E. aerogenes (ESKAPE). To identify proteins in pAF that might be responsible for its antibacterial activity, three different lots of pAF were analyzed with quantitative cytokine arrays that consisted of 400 unique human proteins. One protein identified by microarrays, lactoferrin, and a second prominent antibacterial protein that was not identified by microarrays, lysozyme, were examined by depletion experiments to determine their contribution to the antibacterial activity of pAF.ResultsAll six lots of pAF exhibited antibacterial activity against ESKAPE microorganisms, especially against the pathogens predominately found in chronic wounds (i.e. S. aureus and P. aeruginosa). Thirty-one of the peptides on the microarray were annotated as having antibacterial activity and 26 of these were detected in pAF. Cystatin C and lactoferrin were among the most highly expressed antibacterial proteins in pAF. Cystatin C and lactoferrin were confirmed by ELISA to be present in pAF along with lysozyme. Immunoprecipitation of lactoferrin and lysozyme reduced, but did not abolish the antibacterial activities of pAF.ConclusionOur data demonstrate that pAF maintains antibacterial activity via the preservation of antibacterial proteins against a broad spectrum of wound-associated pathogens.

Osteogenic Differentiation Potential of Human Bone Marrow and Amniotic Fluid-Derived Mesenchymal Stem Cells in Vitro & in Vivo.

BACKGROUND:Cell therapies offer a promising potential in promoting bone regeneration. Stem cell therapy presents attractive care modality in treating degenerative conditions or tissue injuries. The rationale behind this is both the expansion potential of stem cells into a large cell population size and its differentiation abilities into a wide variety of tissue types, when given the proper stimuli. A progenitor stem cell is a promising source of cell therapy in regenerative medicine and bone tissue engineering.AIM:This study aimed to compare the osteogenic differentiation and regenerative potentials of human mesenchymal stem cells derived from human bone marrow (hBM-MSCs) or amniotic fluid (hAF-MSCs), both in vitro and in vivo studies.SUBJECTS AND METHODS:Human MSCs, used in this study, were successfully isolated from two human sources; the bone marrow (BM) and amniotic fluid (AF) collected at the gestational ages of second or third trimesters.RESULTS:The stem cells derived from amniotic fluid seemed to be the most promising type of progenitor cells for clinical applications. In a pre-clinical experiment, attempting to explore the therapeutic application of MSCs in bone regeneration, Rat lumbar spines defects were surgically created and treated with undifferentiated and osteogenically differentiated MSCs, derived from BM and second trimester AF. Cells were loaded on gel-foam scaffolds, inserted and fixed in the area of the surgical defect. X-Ray radiography follows up, and histopathological analysis was done three-four months post- operation. The transplantation of AF-MSCs or BM-MSCs into induced bony defects showed promising results. The AF-MSCs are offering a better healing effect increasing the likelihood of achieving successful spinal fusion. Some bone changes were observed in rats transplanted with osteoblasts differentiated cells but not in rats transplanted with undifferentiated MSCs. Longer observational periods are required to evaluate a true bone formation. The findings of this study suggested that the different sources; hBM-MSCs or hAF-MSCs exhibited remarkably different signature regarding the cell morphology, proliferation capacity and osteogenic differentiation potentialCONCLUSIONS:AF-MSCs have a better performance in vivo bone healing than that of BM-MSCs. Hence, AF derived MSCs is highly recommended as an alternative source to BM-MSCs in bone regeneration and spine fusion surgeries. Moreover, the usage of gel-foam as a scaffold proved as an efficient cell carrier that showed bio-compatibility with cells, bio-degradability and osteoinductivity in vivo.

DRUGPATH \u2013 a novel bioinformatic approach identifies DNA-damage pathway as a regulator of size maintenance in human ESCs and iPSCs

Genetic and biochemical screening approaches often fail to identify functionally relevant pathway networks because many signaling proteins contribute to multiple gene ontology pathways. We developed a DRUGPATH-approach to predict pathway-interactomes from high-content drug screen data. DRUGPATH is based upon combining z-scores of effective inhibitors with their corresponding and validated targets. We test DRUGPATH by comparing homeostatic pathways in human embryonic stem cells (hESCs), human induced pluripotent stem cells (hiPSCs) and human amniotic fluid stem cells (hAFSCs). We show that hAFSCs utilize distinct interactomes compared to hESCs/hiPSCs and that pathways orchestrating cell cycle and apoptosis are strongly interconnected, while pathways regulating survival and size are not. Interestingly, hESCs/hiPSCs regulate their size by growing exact additional sizes during each cell cycle. Chemical and genetic perturbation studies show that this “adder-model” is dependent on the DNA-damage pathway. In the future, the DRUGPATH-approach may help to predict novel pathway interactomes from high-content drug screens.

Human amniotic fluid stem cells (hAFSCs) expressing p21 and cyclin D1 genes retain excellent viability after freezing with (dimethyl sulfoxide) DMSO.

Human amniotic fluid stem cells (hAFSCs) have features intermediate between embryonic and adult SCs, can differentiate into lineages of all three germ layers, and do not develop into tumors in vivo. Moreover, hAFSCs can be easily obtained in routine procedures and there is no ethical or legal limitations regarding their use for clinical and experimental applications. The aim of this study was to assess the effect of slow freezing/thawing and two different concentrations of DMSO (10% DMSO + 90% fetal bovine serum [FBS] and 5% DMSO + 95% FBS) on the survival of hAFSCs. hAFSCs were obtained from 5 pregnant women during amniocentesis at 16-22 weeks of gestation. The expression of pluripotency markers (Octamer-binding transcription factor 4 [Oct4] and NANOG) by reverse transcription polymerase chain reaction and cell surface markers (cluster of differentiation [CD31], CD44, CD45, and CD90) by flow cytometry was analyzed before and after the slow-freezing. Cell viability was assessed by trypan blue exclusion or MTT assay. Quantitative mRNA expression of Oct4, NANOG, cyclin D1 and p21 was determined by real-time PCR before and after the slow-freezing. Pluripotency of hAFSCs was confirmed by NANOG and POU5F1 (Oct4) gene expression before and after slow-freezing. All hAFSC cultures were positive for CD44 and CD90. A higher viability of hAFSCs was observed after freezing with 90% FBS + 10% DMSO. There was increased expression of NANOG and decreased expression of POU5F1 gene after freezing, compared to control cells (before freezing). DMSO and the process of freezing did not significantly change the expression of p21 and cyclin D1 genes in hAFSCs. Overall, our results indicate the applicability of slow-freezing and DMSO in cryopreservation of SCs.

The Effects of human amniotic fluid on periodontal ligament fibroblast cell viability, proliferation, and cytokine/growth factor expression

Background: The importance of the amniotic fluid (AF) to the fetus is clear. However, very few studies have been published to examine the potential uses of this fluid in various areas such as tissue regeneration. AF contains epidermal growth factor, transforming growth factor-alpha, transforming growth factor beta-1, insulin-like growth factor-I, erythropoietin and granulocyte colony-stimulating factor, as well as hyaluronic acid and hyaluronic acid-stimulating factor. Previous studies suggest that AF can increase fibroblast proliferation and chemotaxis, and decrease apoptosis as well as promote wound healing. Furthermore, evidence showed that human AF inhibits hyaluronidase, elastase, and cathepsin. The current study examined the effects of human AF on periodontal ligament fibroblasts (PDLF) in terms of cell toxicity, cell proliferation, and cytokine/growth factor expression. Materials and Methods: Cytotoxicity of AF on PDLF was determined using lactate dehydrogenase assays. PDLF proliferation was determined using water-soluble tetrazolium-1 assays. Cytokine/growth factor expression was determined on AF-treated PDLF, AF alone, and PDLF alone utilizing protein arrays. Results: Human AF at 10% and below did not affect cell growth and was not toxic. AF-treated PDLF cells showed a decrease in cytokine/growth factor levels compared to the sum of cytokine/growth factor levels in AF only and cells only for 39 of the 80 proteins examined (48.8%). Of the 39 examined cytokines, 20 inflammatory cytokines, 11 cell cycle cytokines, 1 anti-inflammatory cytokine, and 7 other cytokines were decreased. Conclusion: Human AF at the examined concentrations was not toxic to PDLF cells and did not influence their proliferation. In addition, AF (10%) caused a decrease in the total protein levels of cytokines/growth factors expressed in 39 of the 80 proteins examined (48.8%). Of the 39 examined cytokines, 20 inflammatory cytokines, 11 cell cycle cytokines, 1 anti-inflammatory cytokine, and 7 other cytokines were decreased.

Isolation and characterization of human amniotic fluid and SH-SY5Y/BE(2)-M17 cell derived exosomes

The application of stem cells as a therapy for degenerative disease holds great promise. Substantial evidence suggests that stem cell derived exosomes are a novel cell-free therapy for the corresponding cells. Exosomes are less complex as compared to their parental cells, due to the fewer number of membrane proteins. In addition, the smaller size and lower risk of immunogenicity makes exosomes potentially safe therapeutic nano-carriers. A large number of ongoing research studies are focused on characterizing exosomes that were derived from different sources, for their potential use in various therapeutic applications. In the present study, we focused on characterizing human amniotic fluid stem cell derived exosomes for future therapeutic applications, such as paracrine therapy/nano carrier. In addition, we characterized exosomes derived from SH-SY5Y and BE(2)-M17 cells, which are a known neuronal model, for further characteristic analyses of neuronal differentiation and neurobiology. Finally, we compared various exosome isolation techniques and procedures and evaluated exosome yield.

Activity of p53 in human amniotic fluid stem cells increases their potentiality as a candidate for stem cell therapy.

The potential use of stem cells as a therapeutic treatment for many neurological disorders, such as stroke, has spiked an interest in their properties. Due to limitations of the present-day treatments, regenerative and protective therapies could prove very beneficial if a safe and effective treatment is identified. Using human amniotic fluid stem (hAFS) cells could theoretically provide both neuroprotective and regenerative properties to patients, and knowledge of p53's activity and function could be a key component in understanding the behavior and characteristics of these stem cells to harness their full potential. Many recent studies on p53 have provided new and valuable information that could give rise to new ideas for treatment options. More specifically, p53's activity inside hAFS cells lead them closer to becoming a potential therapeutic stem cell. Other neuroprotective treatments, such as hyperoxia and hypoxia sessions, are showing positive results. In combination, these data are helping to get closer to an effective treatment for neurological disorders.