Science.gov

Sample records for acellular human amniotic

  1. A new candidate substrate for cell-matrix adhesion study: the acellular human amniotic matrix.

    PubMed

    Guo, Qianchen; Lu, Xuya; Xue, Yuan; Zheng, Hong; Zhao, Xiaotao; Zhao, Huajian

    2012-01-01

    In vivo adhesions between cells and the extracellular matrix play a crucial role in cell differentiation, proliferation, and migration as well as tissue remodeling. Natural three-dimensional (3D) matrices, such as self-assembling matrices and Matrigel, have limitations in terms of their biomechanical properties. Here, we present a simple method to produce an acellular human amniotic matrix (AHAM) with preserved biomechanical properties and a favorable adhesion potential. On the stromal side of the AHAM, human foreskin fibroblasts (HFFs) attached and extended with bipolar spindle-shaped morphology proliferated to multilayer networks, invaded into the AHAM, and migrated in a straight line. Moreover, αV integrin, paxillin, and fibronectin were observed to colocalize after 24 h of HFF culture on the stromal side of the AHAM. Our results indicate that the AHAM may be an ideal candidate as a cell-matrix adhesion substrate to study cell adhesion and invasion as well as other functions in vitro under a tensile force that mimics the in vivo environment.

  2. Production of an acellular matrix from amniotic membrane for the synthesis of a human skin equivalent.

    PubMed

    Sanluis-Verdes, Anahí; Yebra-Pimentel Vilar, Maria Teresa; García-Barreiro, Juan Javier; García-Camba, Marta; Ibáñez, Jacinto Sánchez; Doménech, Nieves; Rendal-Vázquez, Maria Esther

    2015-09-01

    Human amniotic membrane (HAM) has useful properties as a dermal matrix substitute. The objective of our work was to obtain, using different enzymatic or chemical treatments to eliminate cells, a scaffold of acellular HAM for later use as a support for the development of a skin equivalent. The HAM was separated from the chorion, incubated and cryopreserved. The membrane underwent different enzymatic and chemical treatments to eliminate the cells. Fibroblasts and keratinocytes were separately obtained from skin biopsies of patients following a sequential double digestion with first collagenase and then trypsin-EDTA (T/E). A skin equivalent was then constructed by seeding keratinocytes on the epithelial side and fibroblasts on the chorionic side of the decellularizated HAM. Histological, immunohistochemical, inmunofluorescent and molecular biology studies were performed. Treatment with 1% T/E at 37 °C for 30 min totally removed epithelial and mesenchymal cells. The HAM thus treated proved to be a good matrix to support adherence of cells and allowed the achievement of an integral and intact scaffold for development of a skin equivalent, which could be useful as a skin substitute for clinical use.

  3. The effects of acellular amniotic membrane matrix on osteogenic differentiation and ERK1/2 signaling in human dental apical papilla cells.

    PubMed

    Chen, Yi-Jane; Chung, Min-Chun; Jane Yao, Chung-Chen; Huang, Chien-Hsun; Chang, Hao-Hueng; Jeng, Jiiang-Huei; Young, Tai-Horng

    2012-01-01

    The amniotic membrane (AM) has been widely used in the field of tissue engineering because of the favorable biological properties for scaffolding material. However, little is known about the effects of an acellular AM matrix on the osteogenic differentiation of mesenchymal stem cells. In this study, it was found that both basement membrane side and collagenous stroma side of the acellular AM matrix were capable of providing a preferential environment for driving the osteogenic differentiation of human dental apical papilla cells (APCs) with proven stem cell characteristics. Acellular AM matrix potentiated the induction effect of osteogenic supplements (OS) such as ascorbic acid, β-glycerophosphate, and dexamethasone and enhanced the osteogenic differentiation of APCs, as seen by increased core-binding factor alpha 1 (Cbfa-1) phosphorylation, alkaline phosphatase activity, mRNA expression of osteogenic marker genes, and mineralized matrix deposition. Even in the absence of soluble OS, acellular AM matrix also could exert the substrate-induced effect on initiating APCs' differentiation. Especially, the collagenous stroma side was more effective than the basement membrane side. Moreover, the AM-induced effect was significantly inhibited by U0126, an inhibitor of extracellular signaling-regulated kinase 1/2 (ERK1/2) signaling. Taken together, the osteogenic differentiation promoting effect on APCs is AM-specific, which provides potential applications of acellular AM matrix in bone/tooth tissue engineering.

  4. Hair Follicle Morphogenesis in the Treatment of Mouse Full-Thickness Skin Defects Using Composite Human Acellular Amniotic Membrane and Adipose Derived Mesenchymal Stem Cells

    PubMed Central

    Minjuan, Wu; Jun, Xiong; Shiyun, Shao; Sha, Xu; Haitao, Ni

    2016-01-01

    Early repair of skin injury and maximal restoration of the function and appearance have become important targets of clinical treatment. In the present study, we observed the healing process of skin defects in nude mice and structural characteristics of the new skin after transplantation of isolated and cultured adipose derived mesenchymal stem cells (ADMSCs) onto the human acellular amniotic membrane (AAM). The result showed that ADMSCs were closely attached to the surface of AAM and grew well 24 h after seeding. Comparison of the wound healing rate at days 7, 14, and 28 after transplantation showed that ADMSCs seeded on AAM facilitated the healing of full-thickness skin wounds more effectively as compared with either hAM or AAM alone, indicating that ADMSCs participated in skin regeneration. More importantly, we noticed a phenomenon of hair follicle development during the process of skin repair. Composite ADMSCs and AAM not only promoted the healing of the mouse full-thickness defects but also facilitated generation of the appendages of the affected skin, thus promoting restoration of the skin function. Our results provide a new possible therapy idea for the treatment of skin wounds with respect to both anatomical regeneration and functional restoration. PMID:27597871

  5. Hair Follicle Morphogenesis in the Treatment of Mouse Full-Thickness Skin Defects Using Composite Human Acellular Amniotic Membrane and Adipose Derived Mesenchymal Stem Cells

    PubMed Central

    Minjuan, Wu; Jun, Xiong; Shiyun, Shao; Sha, Xu; Haitao, Ni

    2016-01-01

    Early repair of skin injury and maximal restoration of the function and appearance have become important targets of clinical treatment. In the present study, we observed the healing process of skin defects in nude mice and structural characteristics of the new skin after transplantation of isolated and cultured adipose derived mesenchymal stem cells (ADMSCs) onto the human acellular amniotic membrane (AAM). The result showed that ADMSCs were closely attached to the surface of AAM and grew well 24 h after seeding. Comparison of the wound healing rate at days 7, 14, and 28 after transplantation showed that ADMSCs seeded on AAM facilitated the healing of full-thickness skin wounds more effectively as compared with either hAM or AAM alone, indicating that ADMSCs participated in skin regeneration. More importantly, we noticed a phenomenon of hair follicle development during the process of skin repair. Composite ADMSCs and AAM not only promoted the healing of the mouse full-thickness defects but also facilitated generation of the appendages of the affected skin, thus promoting restoration of the skin function. Our results provide a new possible therapy idea for the treatment of skin wounds with respect to both anatomical regeneration and functional restoration.

  6. Hair Follicle Morphogenesis in the Treatment of Mouse Full-Thickness Skin Defects Using Composite Human Acellular Amniotic Membrane and Adipose Derived Mesenchymal Stem Cells.

    PubMed

    Minjuan, Wu; Jun, Xiong; Shiyun, Shao; Sha, Xu; Haitao, Ni; Yue, Wang; Kaihong, Ji

    2016-01-01

    Early repair of skin injury and maximal restoration of the function and appearance have become important targets of clinical treatment. In the present study, we observed the healing process of skin defects in nude mice and structural characteristics of the new skin after transplantation of isolated and cultured adipose derived mesenchymal stem cells (ADMSCs) onto the human acellular amniotic membrane (AAM). The result showed that ADMSCs were closely attached to the surface of AAM and grew well 24 h after seeding. Comparison of the wound healing rate at days 7, 14, and 28 after transplantation showed that ADMSCs seeded on AAM facilitated the healing of full-thickness skin wounds more effectively as compared with either hAM or AAM alone, indicating that ADMSCs participated in skin regeneration. More importantly, we noticed a phenomenon of hair follicle development during the process of skin repair. Composite ADMSCs and AAM not only promoted the healing of the mouse full-thickness defects but also facilitated generation of the appendages of the affected skin, thus promoting restoration of the skin function. Our results provide a new possible therapy idea for the treatment of skin wounds with respect to both anatomical regeneration and functional restoration. PMID:27597871

  7. Metal binding components in human amniotic fluid

    SciTech Connect

    Paterson, P.G.; Zlotkin, S.H.; Sarkar, B. )

    1990-02-26

    Amniotic fluid is a potential source of both nutritionally essential and toxic metals for the fetus. As the binding pattern of these metals in amniotic fluid may be one of the determining factors in their availability to the fetus, the objective of this study was to investigate metal binding in vitro. The binding of six trace metals, Mn(II), Ni(II), Zn(II), Cu(II), Cd(II), and Fe(III), to components of human amniotic fluid was studied by Sephadex G-100 gel filtration at physiological pH, using radioisotopes as tracers and 50 mM TRIS/HCl as the elution buffer. The amniotic fluid was collected at 16-16.5 weeks gestation by amniocentesis and pooled for analysis. Extensive amounts of Fe, Cu, Zn, and Cd and small amounts of Mn and Ni were bound to high molecular weight proteins with elution patterns similar to those seen for the binding of these metals in serum. In addition, large amounts of Fe, Mn, Ni and Cd and small amounts of Zn and Cu were associated with low molecular weight component(s). The identity of these latter components is unknown, but they play an important biological role in amniotic fluid.

  8. Metabolism of vasopressin by human amnion and amniotic fluid

    SciTech Connect

    Claybaugh, J.R.; Uyehara, C.F.T.; Sato, A.K.; Letterie, G.S. )

    1990-02-26

    The authors previously have shown that vasopressin (VP) is metabolized by fetal guinea pig amnion and amniotic fluid into 2 distinct metabolites: M1, produced by amnionic membrane, and M2, produced in amniotic fluid. In this study the authors have examined whether VP can also be metabolized by human amnionic membrane and amniotic fluid collected at term. Tritiated VP (250 pg) was incubated in vitro with either amnionic membrane or amniotic fluid for 2 hours at 37{degrees}C, followed by fractionation by HPLC. After incubation with amnionic membrane, tritiated VP was metabolized to Ml with no production of M2, similar to previous findings with guinea pig amnion. Human amniotic fluid, however, produced both M1 and M2 metabolites, unlike guinea pig amniotic fluid which metabolized VP to only M2 with no M1 production. No other metabolic products other than M1 or M2 were isolated. Also, like the M2 produced in guinea pig amniotic fluid, the M2 metabolite in human amniotic fluid comigrates on HPLC with desglycinamide VP. Thus, metabolic components in the human amniotic sac appear to be similar to those present in the guinea pig. These results indicate that the amniotic compartment may be an important clearance site for fetal VP in humans.

  9. Alternative Sources of Adult Stem Cells: Human Amniotic Membrane

    NASA Astrophysics Data System (ADS)

    Wolbank, Susanne; van Griensven, Martijn; Grillari-Voglauer, Regina; Peterbauer-Scherb, Anja

    Human amniotic membrane is a highly promising cell source for tissue engineering. The cells thereof, human amniotic epithelial cells (hAEC) and human amniotic mesenchymal stromal cells (hAMSC), may be immunoprivileged, they represent an early developmental status, and their application is ethically uncontroversial. Cell banking strategies may use freshly isolated cells or involve in vitro expansion to increase cell numbers. Therefore, we have thoroughly characterized the effect of in vitro cultivation on both phenotype and differentiation potential of hAEC. Moreover, we present different strategies to improve expansion including replacement of animal-derived supplements by human platelet products or the introduction of the catalytic subunit of human telomerase to extend the in vitro lifespan of amniotic cells. Characterization of the resulting cultures includes phenotype, growth characteristics, and differentiation potential, as well as immunogenic and immunomodulatory properties.

  10. Human acellular dermal wound matrix: evidence and experience.

    PubMed

    Kirsner, Robert S; Bohn, Greg; Driver, Vickie R; Mills, Joseph L; Nanney, Lillian B; Williams, Marie L; Wu, Stephanie C

    2015-12-01

    A chronic wound fails to complete an orderly and timely reparative process and places patients at increased risk for wound complications that negatively impact quality of life and require greater health care expenditure. The role of extracellular matrix (ECM) is critical in normal and chronic wound repair. Not only is ECM the largest component of the dermal skin layer, but also ECM proteins provide structure and cell signalling that are necessary for successful tissue repair. Chronic wounds are characterised by their inflammatory and proteolytic environment, which degrades the ECM. Human acellular dermal matrices, which provide an ECM scaffold, therefore, are being used to treat chronic wounds. The ideal human acellular dermal wound matrix (HADWM) would support regenerative healing, providing a structure that could be repopulated by the body's cells. Experienced wound care investigators and clinicians discussed the function of ECM, the evidence related to a specific HADWM (Graftjacket(®) regenerative tissue matrix, Wright Medical Technology, Inc., licensed by KCI USA, Inc., San Antonio, TX), and their clinical experience with this scaffold. This article distills these discussions into an evidence-based and practical overview for treating chronic lower extremity wounds with this HADWM. PMID:24283346

  11. Human acellular dermal wound matrix: evidence and experience.

    PubMed

    Kirsner, Robert S; Bohn, Greg; Driver, Vickie R; Mills, Joseph L; Nanney, Lillian B; Williams, Marie L; Wu, Stephanie C

    2015-12-01

    A chronic wound fails to complete an orderly and timely reparative process and places patients at increased risk for wound complications that negatively impact quality of life and require greater health care expenditure. The role of extracellular matrix (ECM) is critical in normal and chronic wound repair. Not only is ECM the largest component of the dermal skin layer, but also ECM proteins provide structure and cell signalling that are necessary for successful tissue repair. Chronic wounds are characterised by their inflammatory and proteolytic environment, which degrades the ECM. Human acellular dermal matrices, which provide an ECM scaffold, therefore, are being used to treat chronic wounds. The ideal human acellular dermal wound matrix (HADWM) would support regenerative healing, providing a structure that could be repopulated by the body's cells. Experienced wound care investigators and clinicians discussed the function of ECM, the evidence related to a specific HADWM (Graftjacket(®) regenerative tissue matrix, Wright Medical Technology, Inc., licensed by KCI USA, Inc., San Antonio, TX), and their clinical experience with this scaffold. This article distills these discussions into an evidence-based and practical overview for treating chronic lower extremity wounds with this HADWM.

  12. Polychlorinated biphenyls in human amniotic fluid

    SciTech Connect

    Rao, C.V.; Banerji, A.S.

    1988-12-01

    Evidence is available that the PCBs are transferred to the fetus through the placenta. This will pose a grave danger for the developing embryo inside the mothers womb. Since there are not many reports available on the analysis of PCBs in the amniotic fluid, it was thought to analyze some samples of amniotic fluid obtained from random population who do not have record of any previous exposure to PCBs. This paper reports the levels of PCBs in the 26 samples of amniotic fluids samples obtained from maternity hospitals from the women during normal delivery.

  13. Mutagenesis assays of human amniotic fluid

    SciTech Connect

    Everson, R.B.; Milne, K.L.; Warbuton, D.; McClamrock, H.D.; Buchanan, P.D.

    1985-01-01

    Extracts of amniocentesis samples from 144 women were tested for the presence of mutagenic substances using tester strain TA1538 in the Ames Salmonella/mammalian-microsome mutagenicity test. Because the volume of amniotic fluid in these samples was limited (generally less than 10 ml), the authors investigated modifications of this mutagenesis assay that could increase its ability to detect effects from small quantities of test material. Using mutagenicity in samples of urine from smokers as a model, it appeared that improved ability to detect small amounts of mutagen could be obtained by reducing volumes of media and reagents while keeping the amount of test sample constant. Tests of amniotic fluid extracts by this modified procedure showed small increases in revertants, about 50% above dimethylsulfoxide solvent control values. The increases suggest the presence of small amounts of mutagenic material in many of the amniotic fluid samples. At the doses employed, mutagenic activity in these samples was not associated with maternal smoking.

  14. Potential antitumor therapeutic strategies of human amniotic membrane and amniotic fluid-derived stem cells.

    PubMed

    Kang, N-H; Hwang, K-A; Kim, S U; Kim, Y-B; Hyun, S-H; Jeung, E-B; Choi, K-C

    2012-08-01

    As stem cells are capable of self-renewal and can generate differentiated progenies for organ development, they are considered as potential source for regenerative medicine and tissue replacement after injury or disease. Along with this capacity, stem cells have the therapeutic potential for treating human diseases including cancers. According to the origins, stem cells are broadly classified into two types: embryonic stem cells (ESCs) and adult stem cells. In terms of differentiation potential, ESCs are pluripotent and adult stem cells are multipotent. Amnion, which is a membranous sac that contains the fetus and amniotic fluid and functions in protecting the developing embryo during gestation, is another stem cell source. Amnion-derived stem cells are classified as human amniotic membrane-derived epithelial stem cells, human amniotic membrane-derived mesenchymal stem cells and human amniotic fluid-derived stem cells. They are in an intermediate stage between pluripotent ESCs and lineage-restricted adult stem cells, non-tumorigenic, and contribute to low immunogenicity and anti-inflammation. Furthermore, they are easily available and do not cause any controversial issues in their recovery and applications. Not only are amnion-derived stem cells applicable in regenerative medicine, they have anticancer capacity. In non-engineered stem cells transplantation strategies, amnion-derived stem cells effectively target the tumor and suppressed the tumor growth by expressing cytotoxic cytokines. Additionally, they also have a potential as novel delivery vehicles transferring therapeutic genes to the cancer formation sites in gene-directed enzyme/prodrug combination therapy. Owing to their own advantageous properties, amnion-derived stem cells are emerging as a new candidate in anticancer therapy.

  15. Dielectric properties of human placenta, umbilical cord and amniotic fluid

    NASA Astrophysics Data System (ADS)

    Peyman, A.; Gabriel, C.; Benedickter, H. R.; Fröhlich, J.

    2011-04-01

    The dielectric properties of freshly delivered human placenta, umbilical cord and amniotic fluid have been acquired at 37 °C and in the frequency range of 200 MHz-10 GHz. The experimental data were fitted to a Cole-Cole expression. The results show that dielectric properties of the umbilical cord are significantly higher than placenta due to the presence of high water content Wharton's jelly. The results also demonstrate large differences in the dielectric properties of amniotic and cerebrospinal fluids. The data presented can be used in numerical simulations of the exposure of pregnant women to electromagnetic fields.

  16. Effects of Acellular Amniotic Membrane Matrix and Bone Marrow-Derived Mesenchymal Stem Cells in Improving Random Skin Flap Survival in Rats

    PubMed Central

    Chehelcheraghi, Farzaneh; Eimani, Hossein; Homayoonsadraie, Seyed; Torkaman, Giti; Amini, Abdollah; Alavi Majd, Hamid; Shemshadi, Hashem

    2016-01-01

    Background The necrotic skin flap represents a great challenge in plastic and reconstructive surgery. In this study, we evaluated the effect of bioscaffolds, acellular amniotic membranes (AAMs), and bone marrow-derived mesenchymal stem cells (BM-MSCs) on random skin flap (RSF) survival in rats by applying a cell-free extracellular matrix scaffold as a supportive component for the growth and proliferation of BM-MSCs on RSFs. AAM matrix scaffolds were created by incubating AMs in ethylenediaminetetraacetic acid 0.05% at 37°C, and cell scrapers were used. Objectives The aim of the present study was to assess the effect of AAM as a scaffold in TE, and combined with transplanted BM-MSCs, on the survival of RSFs and on the biomechanical parameters of the incision-wound flap margins 7 days after flap elevation. Materials and Methods BM-MSCs and AAMs were transplanted into subcutaneous tissue in the flap area. On the 7th postoperative day, the surviving flap areas were measured using digital imaging software, and the flap tissue was collected for evaluation. Forty rats were randomly divided into four groups of 10 each: group 1 received an AAM injection; group 2 underwent BM-MSC transplantation; group 3 received both AAM injection + BM-MSC transplantation; and group 4 was the control group, receiving only saline. Results The survival area in the AAM/BM-MSC group was significantly higher than in the control group (18.49 ± 1.58 versus 7.51 ± 2.42, P < 0.05). The biomechanical assessment showed no significant differences between the experimental groups and the control group (P > 0.05), and there was no correlation with flap survival. Conclusions Our findings showed that the treatment of flaps with BM-MSC and AAM transplantations significantly promoted flap survival compared to a control group. The viability of the flap was improved by combining BM-MSCs with AAM matrix scaffolds.

  17. Differentiation in human amniotic fluid cell cultures: I: Collagen production.

    PubMed Central

    Priest, R E; Priest, J H; Moinuddin, J F; Keyser, A J

    1977-01-01

    The collagen produced by differentiated cells cultured from human amniotic fluid was characterized in two ways. By chain composition and by 4-hydroxyproline:3-hydroxyproline isomer ratio, the collagen synthesized by F-type (fibroblast) cells was indistinguishable from that made by cultured fetal dermal fibroblasts. The predominant cells in young amniotic fluid cultures, termed AF-type, produced collagen with a lower isomer ratio, resembling that of basement membrane collage. The chain composition, as determined by chromatography on carboxymethyl cellulose, varied for different cultures of the AF-type, but the major pattern was consistent with that of basement membrane collagen. On the basis of these characteristics, F cells are of fibroblast origin, whereas most AF cells are of a different origin either endothelial or epithelial. Other evidence (Megaw et al., 1977) suggests an epithelial origin for AF cells. PMID:881704

  18. One-stage human acellular nerve allograft reconstruction for digital nerve defects

    PubMed Central

    Li, Xue-yuan; Hu, Hao-liang; Fei, Jian-rong; Wang, Xin; Wang, Tian-bing; Zhang, Pei-xun; Chen, Hong

    2015-01-01

    Human acellular nerve allografts have a wide range of donor origin and can effectively avoid nerve injury in the donor area. Very little is known about one-stage reconstruction of digital nerve defects. The present study observed the feasibility and effectiveness of human acellular nerve allograft in the reconstruction of < 5-cm digital nerve defects within 6 hours after injury. A total of 15 cases of nerve injury, combined with nerve defects in 18 digits from the Department of Emergency were enrolled in this study. After debridement, digital nerves were reconstructed using human acellular nerve allografts. The patients were followed up for 6–24 months after reconstruction. Mackinnon-Dellon static two-point discrimination results showed excellent and good rates of 89%. Semmes-Weinstein monofilament test demonstrated that light touch was normal, with an obvious improvement rate of 78%. These findings confirmed that human acellular nerve allograft for one-stage reconstruction of digital nerve defect after hand injury is feasible, which provides a novel trend for peripheral nerve reconstruction. PMID:25788927

  19. Effects of Acellular Amniotic Membrane Matrix and Bone Marrow-Derived Mesenchymal Stem Cells in Improving Random Skin Flap Survival in Rats

    PubMed Central

    Chehelcheraghi, Farzaneh; Eimani, Hossein; Homayoonsadraie, Seyed; Torkaman, Giti; Amini, Abdollah; Alavi Majd, Hamid; Shemshadi, Hashem

    2016-01-01

    Background The necrotic skin flap represents a great challenge in plastic and reconstructive surgery. In this study, we evaluated the effect of bioscaffolds, acellular amniotic membranes (AAMs), and bone marrow-derived mesenchymal stem cells (BM-MSCs) on random skin flap (RSF) survival in rats by applying a cell-free extracellular matrix scaffold as a supportive component for the growth and proliferation of BM-MSCs on RSFs. AAM matrix scaffolds were created by incubating AMs in ethylenediaminetetraacetic acid 0.05% at 37°C, and cell scrapers were used. Objectives The aim of the present study was to assess the effect of AAM as a scaffold in TE, and combined with transplanted BM-MSCs, on the survival of RSFs and on the biomechanical parameters of the incision-wound flap margins 7 days after flap elevation. Materials and Methods BM-MSCs and AAMs were transplanted into subcutaneous tissue in the flap area. On the 7th postoperative day, the surviving flap areas were measured using digital imaging software, and the flap tissue was collected for evaluation. Forty rats were randomly divided into four groups of 10 each: group 1 received an AAM injection; group 2 underwent BM-MSC transplantation; group 3 received both AAM injection + BM-MSC transplantation; and group 4 was the control group, receiving only saline. Results The survival area in the AAM/BM-MSC group was significantly higher than in the control group (18.49 ± 1.58 versus 7.51 ± 2.42, P < 0.05). The biomechanical assessment showed no significant differences between the experimental groups and the control group (P > 0.05), and there was no correlation with flap survival. Conclusions Our findings showed that the treatment of flaps with BM-MSC and AAM transplantations significantly promoted flap survival compared to a control group. The viability of the flap was improved by combining BM-MSCs with AAM matrix scaffolds. PMID:27621924

  20. Human amniotic fluid: a source of stem cells for possible therapeutic use.

    PubMed

    Dziadosz, Margaret; Basch, Ross S; Young, Bruce K

    2016-03-01

    Stem cells are undifferentiated cells with the capacity for differentiation. Amniotic fluid cells have emerged only recently as a possible source of stem cells for clinical purposes. There are no ethical or sampling constraints for the use of amniocentesis as a standard clinical procedure for obtaining an abundant supply of amniotic fluid cells. Amniotic fluid cells of human origin proliferate rapidly and are multipotent with the potential for expansion in vitro to multiple cell lines. Tissue engineering technologies that use amniotic fluid cells are being explored. Amniotic fluid cells may be of clinical benefit for fetal therapies, degenerative disease, and regenerative medicine applications. We present a comprehensive review of the evolution of human amniotic fluid cells as a possible modality for therapeutic use.

  1. Establishment of immortalized human amniotic mesenchymal stem cells.

    PubMed

    Teng, Zan; Yoshida, Toshiko; Okabe, Motonori; Toda, Ayaka; Higuchi, Osamu; Nogami, Makiko; Yoneda, Noriko; Zhou, Kaixuan; Kyo, Satoru; Kiyono, Touru; Nikaido, Toshio

    2013-01-01

    Human amniotic mesenchymal cells (HAM cells) are known to contain somatic stem cells possessing the characteristics of pluripotency. However, little is known about the biology of these somatic cells because isolated HAM cells from amniotic membrane have a limited lifespan. To overcome this problem, we attempted to prolong the lifespan of HAM cells by infecting retrovirus encoding human papillomavirus type16E6 and E7 (HPV16E6E7), bmi-1, and/or human telomerase reverse transcriptase (hTERT) genes and investigated their characteristics as stem cells. We confirmed the immortalization of the four lines of cultured HAM cells for about 1 year. Immortalized human amnion mesenchymal cells (iHAM cells) have continued to proliferate over 200 population doublings (PDs). iHAM cells were positive for CD73, CD90, CD105, and CD44 and negative for CD34, CD14, CD45, and HLA-DR. They expressed stem cell markers such as Oct3/4, Sox2, Nanog, Klf4, SSEA4, c-myc, vimentin, and nestin. They showed adipogenic, osteogenic, and chondrogenic differentiation abilities after induction. These results suggested that immortalized cell lines with characteristics of stem cells can be established. iHAM cells with an extended lifespan can be used to produce good experimental models both in vitro and in vivo.

  2. Biocompatible surgical meshes based on decellularized human amniotic membrane.

    PubMed

    Shi, Peina; Gao, Mengna; Shen, Qiuxia; Hou, Lei; Zhu, Yabin; Wang, Jun

    2015-09-01

    Meshes play important roles to repair human tissue defect. In this work, human amniotic membrane (HAM) was decellularized and explored the efficacy as an implantable biological mesh. Surfactant, hypertonic saline, lipase and DNAase were used individually or collectively to remove all cell components and remain the extracellular matrix. Results of H&E and DAPI staining demonstrated that the method of surfactant and lipase combining with DNAase is the most effective treatment for HAM decellularization. Primary smooth muscle cells were seeded to evaluate the decellularized HAM's (dHAM) in vitro cytocompatibility. The in vivo test was performed via implantation at rabbits' uterus with clinic polypropylene mesh (PP) as the control. The results indicated that dHAM possessed good biocompatibility and will be a potential candidate for biological mesh.

  3. DermACELL: Human Acellular Dermal Matrix Allograft A Case Report.

    PubMed

    Cole, Windy E

    2016-03-01

    Diabetes often causes ulcers on the feet of diabetic patients. A 56-year-old, insulin-dependent, diabetic woman presented to the wound care center with a Wagner grade 3 ulcer of the right heel. She reported a 3-week history of ulceration with moderate drainage and odor and had a history of ulceration and osteomyelitis in the contralateral limb. Rigorous wound care, including hospitalization; surgical incision and drainage; intravenous antibiotic drug therapy; vacuum-assisted therapy; and a new room temperature, sterile, human acellular dermal matrix graft were used to heal the wound, save her limb, and restore her activities of daily living. This case presentation involves alternative treatment of a diabetic foot ulcer with this new acellular dermal matrix, DermACELL. PMID:27031550

  4. A simple alkaline method for decellularizing human amniotic membrane for cell culture.

    PubMed

    Saghizadeh, Mehrnoosh; Winkler, Michael A; Kramerov, Andrei A; Hemmati, David M; Ghiam, Chantelle A; Dimitrijevich, Slobodan D; Sareen, Dhruv; Ornelas, Loren; Ghiasi, Homayon; Brunken, William J; Maguen, Ezra; Rabinowitz, Yaron S; Svendsen, Clive N; Jirsova, Katerina; Ljubimov, Alexander V

    2013-01-01

    Human amniotic membrane is a standard substratum used to culture limbal epithelial stem cells for transplantation to patients with limbal stem cell deficiency. Various methods were developed to decellularize amniotic membrane, because denuded membrane is poorly immunogenic and better supports repopulation by dissociated limbal epithelial cells. Amniotic membrane denuding usually involves treatment with EDTA and/or proteolytic enzymes; in many cases additional mechanical scraping is required. Although ensuring limbal cell proliferation, these methods are not standardized, require relatively long treatment times and can result in membrane damage. We propose to use 0.5 M NaOH to reliably remove amniotic cells from the membrane. This method was used before to lyse cells for DNA isolation and radioactivity counting. Gently rubbing a cotton swab soaked in NaOH over the epithelial side of amniotic membrane leads to nearly complete and easy removal of adherent cells in less than a minute. The denuded membrane is subsequently washed in a neutral buffer. Cell removal was more thorough and uniform than with EDTA, or EDTA plus mechanical scraping with an electric toothbrush, or n-heptanol plus EDTA treatment. NaOH-denuded amniotic membrane did not show any perforations compared with mechanical or thermolysin denuding, and showed excellent preservation of immunoreactivity for major basement membrane components including laminin α2, γ1-γ3 chains, α1/α2 and α6 type IV collagen chains, fibronectin, nidogen-2, and perlecan. Sodium hydroxide treatment was efficient with fresh or cryopreserved (10% dimethyl sulfoxide or 50% glycerol) amniotic membrane. The latter method is a common way of membrane storage for subsequent grafting in the European Union. NaOH-denuded amniotic membrane supported growth of human limbal epithelial cells, immortalized corneal epithelial cells, and induced pluripotent stem cells. This simple, fast and reliable method can be used to standardize

  5. Adaptation of Group A Streptococcus to Human Amniotic Fluid

    PubMed Central

    Sitkiewicz, Izabela; Green, Nicole M.; Guo, Nina; Bongiovanni, Ann M.; Witkin, Steven S.; Musser, James M.

    2010-01-01

    Background For more than 100 years, group A Streptococcus has been identified as a cause of severe and, in many cases, fatal infections of the female urogenital tract. Due to advances in hospital hygiene and the advent of antibiotics, this type of infection has been virtually eradicated. However, within the last three decades there has been an increase in severe intra- and post-partum infections attributed to GAS. Methodology We hypothesized that GAS alters its transcriptome to survive in human amniotic fluid (AF) and cause disease. To identify genes that were up or down regulated in response to growth in AF, GAS was grown in human AF or standard laboratory media (THY) and samples for expression microarray analysis were collected during mid-logarithmic, late-logarithmic, and stationary growth phases. Microarray analysis was performed using a custom Affymetrix chip and normalized hybridization values derived from three biological replicates were collected at each growth point. Ratios of AF/THY above a 2-fold change and P-value <0.05 were considered significant. Principal Findings The majority of changes in the GAS transcriptome involved down regulation of multiple adhesins and virulence factors and activation of the stress response. We observed significant changes in genes involved in the arginine deiminase pathway and in the nucleotide de novo synthesis pathway. Conclusions/Significance Our work provides new insight into how pathogenic bacteria respond to their environment to establish infection and cause disease. PMID:20352104

  6. Comparison of a poly-l-lactide-co-ɛ-caprolactone and human amniotic membrane for urothelium tissue engineering applications

    PubMed Central

    Sartoneva, Reetta; Haimi, Suvi; Miettinen, Susanna; Mannerström, Bettina; Haaparanta, Anne-Marie; Sándor, George K.; Kellomäki, Minna; Suuronen, Riitta; Lahdes-Vasama, Tuija

    2011-01-01

    The reconstructive surgery of urothelial defects, such as severe hypospadias is susceptible to complications. The major problem is the lack of suitable grafting materials. Therefore, finding alternative treatments such as reconstruction of urethra using tissue engineering is essential. The aim of this study was to compare the effects of naturally derived acellular human amniotic membrane (hAM) to synthetic poly-l-lactide-co-ε-caprolactone (PLCL) on human urothelial cell (hUC) viability, proliferation and urothelial differentiation level. The viability of cells was evaluated using live/dead staining and the proliferation was studied using WST-1 measurement. Cytokeratin (CK)7/8 and CK19 were used to confirm that the hUCs maintained their phenotype on different biomaterials. On the PLCL, the cell number significantly increased during the culturing period, in contrast to the hAM, where hUC proliferation was the weakest at 7 and 14 days. In addition, the majority of cells were viable and maintained their phenotype when cultured on PLCL and cell culture plastic, whereas on the hAM, the viability of hUCs decreased with time and the cells did not maintain their phenotype. The PLCL membranes supported the hUC proliferation significantly more than the hAM. These results revealed the significant potential of PLCL membranes in urothelial tissue engineering applications. PMID:21106575

  7. Transparent, resilient human amniotic membrane laminates for corneal transplantation.

    PubMed

    Hariya, Takehiro; Tanaka, Yuji; Yokokura, Shunji; Nakazawa, Toru

    2016-09-01

    This study evaluated a new technique to toughen and optically clarify human amniotic membrane (AM) tissue, which is naturally thin and clouded, and determined the suitability of the altered tissue for corneal transplantation. The technique created a tissue laminate by repeatedly depositing wet layers of AM and dehydrating them, followed by chemical cross-linking to tighten integration at the layer interfaces and within the layers, thereby improving the physical properties of the laminates by increasing light transmittance and mechanical strength. Interestingly, this improvement only occurred in laminates with at least 4 layers. Cross-linking also improved the resistance of the laminates to collagenase degradation, such as occurs in corneal melting. This study also confirmed that the AM tissue was biocompatible by inserting AM monolayers into the corneal stroma of rabbits, and by performing lamellar keratoplasty in rabbits with cross-linked AM laminates. The laminates were sufficiently thick and resilient to need only one set of sutures, whereas in previously described multi-layer AM transplantation technique, each layer required separate sutures. The current findings are a promising advance in the engineering of novel biomaterials and the alteration of existing tissues for medical use. PMID:27267629

  8. Human amniotic fluid stem cell preconditioning improves their regenerative potential.

    PubMed

    Rota, Cinzia; Imberti, Barbara; Pozzobon, Michela; Piccoli, Martina; De Coppi, Paolo; Atala, Anthony; Gagliardini, Elena; Xinaris, Christodoulos; Benedetti, Valentina; Fabricio, Aline S C; Squarcina, Elisa; Abbate, Mauro; Benigni, Ariela; Remuzzi, Giuseppe; Morigi, Marina

    2012-07-20

    Human amniotic fluid stem (hAFS) cells, a novel class of broadly multipotent stem cells that share characteristics of both embryonic and adult stem cells, have been regarded as promising candidate for cell therapy. Taking advantage by the well-established murine model of acute kidney injury (AKI), we studied the proregenerative effect of hAFS cells in immunodeficient mice injected with the nephrotoxic drug cisplatin. Infusion of hAFS cells in cisplatin mice improved renal function and limited tubular damage, although not to control level, and prolonged animal survival. Human AFS cells engrafted injured kidney predominantly in peritubular region without acquiring tubular epithelial markers. Human AFS cells exerted antiapoptotic effect, activated Akt, and stimulated proliferation of tubular cells possibly via local release of factors, including interleukin-6, vascular endothelial growth factor, and stromal cell-derived factor-1, which we documented in vitro to be produced by hAFS cells. The therapeutic potential of hAFS cells was enhanced by cell pretreatment with glial cell line-derived neurotrophic factor (GDNF), which markedly ameliorated renal function and tubular injury by increasing stem cell homing to the tubulointerstitial compartment. By in vitro studies, GDNF increased hAFS cell production of growth factors, motility, and expression of receptors involved in cell homing and survival. These findings indicate that hAFS cells can promote functional recovery and contribute to renal regeneration in AKI mice via local production of mitogenic and prosurvival factors. The effects of hAFS cells can be remarkably enhanced by GDNF preconditioning.

  9. Human Amniotic Fluid Stem Cell Preconditioning Improves Their Regenerative Potential

    PubMed Central

    Rota, Cinzia; Imberti, Barbara; Pozzobon, Michela; Piccoli, Martina; De Coppi, Paolo; Atala, Anthony; Gagliardini, Elena; Xinaris, Christodoulos; Benedetti, Valentina; Fabricio, Aline S.C.; Squarcina, Elisa; Abbate, Mauro; Benigni, Ariela; Remuzzi, Giuseppe

    2012-01-01

    Human amniotic fluid stem (hAFS) cells, a novel class of broadly multipotent stem cells that share characteristics of both embryonic and adult stem cells, have been regarded as promising candidate for cell therapy. Taking advantage by the well-established murine model of acute kidney injury (AKI), we studied the proregenerative effect of hAFS cells in immunodeficient mice injected with the nephrotoxic drug cisplatin. Infusion of hAFS cells in cisplatin mice improved renal function and limited tubular damage, although not to control level, and prolonged animal survival. Human AFS cells engrafted injured kidney predominantly in peritubular region without acquiring tubular epithelial markers. Human AFS cells exerted antiapoptotic effect, activated Akt, and stimulated proliferation of tubular cells possibly via local release of factors, including interleukin-6, vascular endothelial growth factor, and stromal cell–derived factor-1, which we documented in vitro to be produced by hAFS cells. The therapeutic potential of hAFS cells was enhanced by cell pretreatment with glial cell line–derived neurotrophic factor (GDNF), which markedly ameliorated renal function and tubular injury by increasing stem cell homing to the tubulointerstitial compartment. By in vitro studies, GDNF increased hAFS cell production of growth factors, motility, and expression of receptors involved in cell homing and survival. These findings indicate that hAFS cells can promote functional recovery and contribute to renal regeneration in AKI mice via local production of mitogenic and prosurvival factors. The effects of hAFS cells can be remarkably enhanced by GDNF preconditioning. PMID:22066606

  10. Amniotic Fluid

    PubMed Central

    Smith, Heather C.; Muglia, Louis J.; Morrow, Ardythe L.

    2014-01-01

    Our aim was to review the use of high-dimensional biology techniques, specifically transcriptomics, proteomics, and metabolomics, in amniotic fluid to elucidate the mechanisms behind preterm birth or assessment of fetal development. We performed a comprehensive MEDLINE literature search on the use of transcriptomic, proteomic, and metabolomic technologies for amniotic fluid analysis. All abstracts were reviewed for pertinence to preterm birth or fetal maturation in human subjects. Nineteen articles qualified for inclusion. Most articles described the discovery of biomarker candidates, but few larger, multicenter replication or validation studies have been done. We conclude that the use of high-dimensional systems biology techniques to analyze amniotic fluid has significant potential to elucidate the mechanisms of preterm birth and fetal maturation. However, further multicenter collaborative efforts are needed to replicate and validate candidate biomarkers before they can become useful tools for clinical practice. Ideally, amniotic fluid biomarkers should be translated to a noninvasive test performed in maternal serum or urine. PMID:23599373

  11. Characteristics of human amniotic fluid mesenchymal stem cells and their tropism to human ovarian cancer.

    PubMed

    Li, Liru; Wang, Dejun; Zhou, Jun; Cheng, Yan; Liang, Tian; Zhang, Guangmei

    2015-01-01

    The mesenchymal stem cells (MSCs) derived from amniotic fluid (AF) have become an attractive stem cells source for cell-based therapy because they can be harvested at low cost and avoid ethical disputes. In human research, stem cells derived from AF gradually became a hot research direction for disease treatment, specifically for their plasticity, their reduced immunogenicity and their tumor tropism regardless of the tumor size, location and source. Our work aimed to obtain and characterize human amniotic fluid mesenchymal stem cells (AFMSCs) and detect their ovarian cancer tropsim in nude mice model. Ten milliliters of twenty independent amniotic fluid samples were collected from 16-20 week pregnant women who underwent amniocentesis for fetal genetic determination in routine prenatal diagnosis in the first affiliated hospital of Harbin medical university. We successfully isolated the AFMSCs from thirteen of twenty amniotic fluid samples. AFMSCs presented a fibroblastic-like morphology during the culture. Flow cytometry analyses showed that the cells were positive for specific stem cell markers CD73,CD90, CD105, CD166 and HLA-ABC (MHC class I), but negative for CD 45,CD40, CD34, CD14 and HLA-DR (MHC class II). RT-PCR results showed that the AFMSCs expressed stem cell marker OCT4. AFMSCs could differentiate into bone cells, fat cells and chondrocytes under certain conditions. AFMSCs had the high motility to migrate to ovarian cancer site but didn't have the tumorigenicity. This study enhances the possibility of AFMSCs as drug carrier in human cell-based therapy. Meanwhile, the research emphasis in the future can also put in targeting therapy of ovarian cancer.

  12. The role of amniotic fluid in force transfer during human birth

    NASA Astrophysics Data System (ADS)

    Baumer, Alexa; Lehn, Andrea; Leftwich, Megan

    2013-11-01

    This study seeks to understand the fundamental fluid dynamic processes involved in human birth. We begin by examining the importance of amniotic fluid. This is done using two experimental techniques that approximate the laboring human uterus to different degrees of anatomical correctness. The first, in which a latex uterus is filled with fluid and a solid fetus is extracted, investigates the importance of both amniotic fluid properties and fetal position in the force required to remove a fetus. The second experiment simplifies the geometry of birth even more. In this case, a solid cylindrical rod is pulled through a highly flexible outer tube. The force to pull the inner cylinder as a function of the gap fluid properties is measured. By carefully controlling the fluid properties of the experiment, the study will provide further insight into the roles of amniotic fluid in human birth.

  13. Neovascularization in a mouse model via stem cells derived from human fetal amniotic membranes.

    PubMed

    Kim, Hwi Gon; Choi, Ook Hwan

    2011-03-01

    In this study, we evaluated the effect of culture-expanded mesenchymal stem cells (MSCs), derived from amniotic membranes, on neovascularization and blood flow, in an animal model of limb ischemia in immune-deficient mice. MSCs were cultured from human amniotic membranes by collagenase digestion. Human amniotic mesenchymal stem cells (hAMSCs) were administered intramuscularly at three different sites of the ischemic leg whose femoral vessels were ligated. After 4 weeks of culture, a population of homogeneous mesenchymal cells was isolated from the human amniotic membranes after confluence was reached. We performed three different groups of mice model [controls, hAMSCs, conditioned media from the hAMSCs (hAMSCs-CM)]. The blood flow recovery in the hindlimb ischemia model was significantly higher in the hAMSC-transplanted group than in the control group. Moreover, hAMSCs-CM significantly improved the cutaneous blood flow. The histological examination showed that red fluorescence (CM-DiI)-labeled hAMSCs was detected in the interstitial tissues between the muscle fibers 2 weeks after transplantation. The results of this study showed that hAMSCs may be an attractive, alternative source of progenitor or stem cells for basic research as well as clinical applications.

  14. Effect of the Human Amniotic Membrane on Liver Regeneration in Rats

    PubMed Central

    Sipahi, Mesut; Şahin, Sevinç; Arslan, Ergin; Börekci, Hasan; Metin, Bayram; Cantürk, Nuh Zafer

    2015-01-01

    Introduction. Operations are performed for broader liver surgery indications for a better understanding of hepatic anatomy/physiology and developments in operation technology. Surgery can cure some patients with liver metastasis of some tumors. Nevertheless, postoperative liver failure is the most feared complication causing mortality in patients who have undergone excision of a large liver mass. The human amniotic membrane has regenerative effects. Thus, we investigated the effects of the human amniotic membrane on regeneration of the resected liver. Methods. Twenty female Wistar albino rats were divided into control and experimental groups and underwent a 70% hepatectomy. The human amniotic membrane was placed over the residual liver in the experimental group. Relative liver weight, histopathological features, and biochemical parameters were assessed on postoperative day 3. Results. Total protein and albumin levels were significantly lower in the experimental group than in the control group. No difference in relative liver weight was observed between the groups. Hepatocyte mitotic count was significantly higher in the experimental group than in the control group. Hepatic steatosis was detected in the experimental group. Conclusion. Applying the amniotic membrane to residual liver adversely affected liver regeneration. However, mesenchymal stem cell research has the potential to accelerate liver regeneration investigations. PMID:26457000

  15. Three-dimensional Reconstruction of the Microstructure of Human Acellular Nerve Allograft.

    PubMed

    Zhu, Shuang; Zhu, Qingtang; Liu, Xiaolin; Yang, Weihong; Jian, Yutao; Zhou, Xiang; He, Bo; Gu, Liqiang; Yan, Liwei; Lin, Tao; Xiang, Jianping; Qi, Jian

    2016-01-01

    The exact inner 3D microstructure of the human peripheral nerve has been a mystery for decades. Therefore, it has been difficult to solve several problems regarding peripheral nerve injury and repair. We used high-resolution X-ray computed microtomography (microCT) to scan a freeze-dried human acellular nerve allograft (hANA). The microCT images were then used to reconstruct a 3D digital model, which was used to print a 3D resin model of the nerve graft. The 3D digital model of the hANA allowed visualization of all planes. The magnified 3D resin model clearly showed the nerve bundles and basement membrane tubes of the hANA. Scanning electron microscopy (SEM) was used to analyse the microstructure of the hANA. Compared to the SEM images, the microCT image clearly demonstrated the microstructure of the hANA cross section at a resolution of up to 1.2 μm. The 3D digital model of the hANA facilitates a clear and easy understanding of peripheral nerve microstructure. Furthermore, the enlarged 3D resin model duplicates the unique inner structure of each individual hANA. This is a crucial step towards achieving 3D printing of a hANA or nerve that can be used as a nerve graft. PMID:27476584

  16. Three-dimensional Reconstruction of the Microstructure of Human Acellular Nerve Allograft.

    PubMed

    Zhu, Shuang; Zhu, Qingtang; Liu, Xiaolin; Yang, Weihong; Jian, Yutao; Zhou, Xiang; He, Bo; Gu, Liqiang; Yan, Liwei; Lin, Tao; Xiang, Jianping; Qi, Jian

    2016-01-01

    The exact inner 3D microstructure of the human peripheral nerve has been a mystery for decades. Therefore, it has been difficult to solve several problems regarding peripheral nerve injury and repair. We used high-resolution X-ray computed microtomography (microCT) to scan a freeze-dried human acellular nerve allograft (hANA). The microCT images were then used to reconstruct a 3D digital model, which was used to print a 3D resin model of the nerve graft. The 3D digital model of the hANA allowed visualization of all planes. The magnified 3D resin model clearly showed the nerve bundles and basement membrane tubes of the hANA. Scanning electron microscopy (SEM) was used to analyse the microstructure of the hANA. Compared to the SEM images, the microCT image clearly demonstrated the microstructure of the hANA cross section at a resolution of up to 1.2 μm. The 3D digital model of the hANA facilitates a clear and easy understanding of peripheral nerve microstructure. Furthermore, the enlarged 3D resin model duplicates the unique inner structure of each individual hANA. This is a crucial step towards achieving 3D printing of a hANA or nerve that can be used as a nerve graft.

  17. Three-dimensional Reconstruction of the Microstructure of Human Acellular Nerve Allograft

    PubMed Central

    Zhu, Shuang; Zhu, Qingtang; Liu, Xiaolin; Yang, Weihong; Jian, Yutao; Zhou, Xiang; He, Bo; Gu, Liqiang; Yan, Liwei; Lin, Tao; Xiang, Jianping; Qi, Jian

    2016-01-01

    The exact inner 3D microstructure of the human peripheral nerve has been a mystery for decades. Therefore, it has been difficult to solve several problems regarding peripheral nerve injury and repair. We used high-resolution X-ray computed microtomography (microCT) to scan a freeze-dried human acellular nerve allograft (hANA). The microCT images were then used to reconstruct a 3D digital model, which was used to print a 3D resin model of the nerve graft. The 3D digital model of the hANA allowed visualization of all planes. The magnified 3D resin model clearly showed the nerve bundles and basement membrane tubes of the hANA. Scanning electron microscopy (SEM) was used to analyse the microstructure of the hANA. Compared to the SEM images, the microCT image clearly demonstrated the microstructure of the hANA cross section at a resolution of up to 1.2 μm. The 3D digital model of the hANA facilitates a clear and easy understanding of peripheral nerve microstructure. Furthermore, the enlarged 3D resin model duplicates the unique inner structure of each individual hANA. This is a crucial step towards achieving 3D printing of a hANA or nerve that can be used as a nerve graft. PMID:27476584

  18. Quantitative mapping of intracellular cations in the human amniotic membrane

    NASA Astrophysics Data System (ADS)

    Moretto, Ph.; Llabador, Y.; Simonoff, M.; Razafindrabe, L.; Bara, M.; Guiet-Bara, A.

    1993-05-01

    The effect of magnesium and taurine on the permeability of cell membranes to monovalent cations has been investigated using the Bordeaux nuclear microprobe. PIXE and RBS techniques have been used to provide quantitative measurements and ion distributions in the isolated amniotic membrane. This physiological model for cellular exchanges allowed us to reveal the distribution of most elements involved in cellular pathways and the modifications under different experimental conditions of incubation in physiological fluids. The PIXE microanalysis provided an original viewpoint on these mechanisms. Following this first study, the amnion compact lamina was found to play a role which was not, up to now, taken into account in the interpretation of electrophysiological experimentations. The release of some ionic species, such as K +, from the epithelial cells, during immersion in isotonic fluids, could have been hitherto underestimated.

  19. Use of peracetic acid to sterilize human donor skin for production of acellular dermal matrices for clinical use.

    PubMed

    Huang, Qizhi; Dawson, Rebecca A; Pegg, David E; Kearney, John N; Macneil, Sheila

    2004-01-01

    We previously reported methods for sterilizing human skin for clinical use. In a comparison of gamma-irradiation, glycerol, and ethylene oxide, sterilization with ethylene oxide after treatment with glycerol provided the most satisfactory dermis in terms of structure and its ability to produce reconstructed skin with many of the characteristics of normal skin. However, the use of ethylene oxide is becoming less common in the United Kingdom due to concerns about its possible genotoxicity. The aim of this study was to evaluate peracetic acid as an alternative sterilizing agent. Skin sterilized with peracetic acid was compared with skin sterilized using glycerol alone or glycerol with ethylene oxide. The effect of subsequently storing peracetic acid sterilized skin in glycerol or propylene glycol was also examined. Acellular dermal matrices were produced after removal of the epidermis and cells in the dermis, processed for histological and ultrastructural analysis, and the biological function was evaluated by reconstitution with keratinocytes and fibroblasts. Results showed that sterilized acellular matrices retained the integrity of dermal structure and major components of the basement membrane. There were no overall significant differences in the ability of these matrices to form reconstructed skin, but peracetic acid alone gave a lower histologic score than when combined with glycerol or propylene glycol. We conclude that peracetic acid sterilization followed by preservation in glycerol or propylene glycol offers a convenient alternative protocol for processing of human skin. It is suggested that this sterile acellular dermis may be suitable for clinical use.

  20. Effects of Gamma Irradiation on Bacterial Microflora Associated with Human Amniotic Membrane

    PubMed Central

    Binte Atique, Fahmida; Ahmed, Kazi Tahsin; Asaduzzaman, S. M.; Hasan, Kazi Nadim

    2013-01-01

    Human amniotic membrane is considered a promising allograft material for the treatment of ocular surface reconstruction, burns, and other skin defects. In order to avoid the transmission of any diseases, grafts should be perfectly sterile. Twenty-five amniotic sacs were collected to determine the microbiological quality of human amniotic membrane, to analyze the radiation sensitivity pattern of the microorganism, and to detect the radiation decimal reduction dose (D10) values. All the samples were found to be contaminated, and the bioburden was ranged from 3.4 × 102 to 1.2 × 105 cfu/g. Initially, a total fifty bacterial isolates were characterized according to their cultural, morphological, and biochemical characteristics and then tested for the radiation sensitivity in an incremental series of radiation doses from 1 to 10 KGy. The results depict gradual decline in bioburden with incline of radiation doses. Staphylococcus spp. were the most frequently isolated bacterial contaminant in tissue samples (44%). The D10 values of the bacterial isolates were ranged from 0.6 to 1.27 KGy. Streptococcus spp. were found to be the highest radioresistant strain with the radiation sterilization dose (RSD) of 11.4 KGy for a bioburden level of 1000. To compare the differences, D10 values were also calculated by graphical evaluations of the data with two of the representative isolates of each bacterial species which showed no significant variations. Findings of this study indicate that lower radiation dose is quite satisfactory for the sterilization of amniotic membrane grafts. Therefore, these findings would be helpful to predict the efficacy of radiation doses for the processing of amniotic membrane for various purposes. PMID:24063009

  1. Glycerolized Reticular Dermis as a New Human Acellular Dermal Matrix: An Exploratory Study.

    PubMed

    Ferrando, Pietro Maria; Balmativola, Davide; Cambieri, Irene; Scalzo, Maria Stella; Bergallo, Massimiliano; Annaratone, Laura; Casarin, Stefania; Fumagalli, Mara; Stella, Maurizio; Sapino, Anna; Castagnoli, Carlotta

    2016-01-01

    Human Acellular Dermal Matrices (HADM) are employed in various reconstructive surgery procedures as scaffolds for autologous tissue regeneration. The aim of this project was to develop a new type of HADM for clinical use, composed of glycerolized reticular dermis decellularized through incubation and tilting in Dulbecco's Modified Eagle's Medium (DMEM). This manufacturing method was compared with a decellularization procedure already described in the literature, based on the use of sodium hydroxide (NaOH), on samples from 28 donors. Cell viability was assessed using an MTT assay and microbiological monitoring was performed on all samples processed after each step. Two surgeons evaluated the biomechanical characteristics of grafts of increasing thickness. The effects of the different decellularization protocols were assessed by means of histological examination and immunohistochemistry, and residual DNA after decellularization was quantified using a real-time TaqMan MGB probe. Finally, we compared the results of DMEM based decellularization protocol on reticular dermis derived samples with the results of the same protocol applied on papillary dermis derived grafts. Our experimental results indicated that the use of glycerolized reticular dermis after 5 weeks of treatment with DMEM results in an HADM with good handling and biocompatibility properties. PMID:26918526

  2. Glycerolized Reticular Dermis as a New Human Acellular Dermal Matrix: An Exploratory Study

    PubMed Central

    Ferrando, Pietro Maria; Balmativola, Davide; Cambieri, Irene; Scalzo, Maria Stella; Bergallo, Massimiliano; Annaratone, Laura; Casarin, Stefania; Fumagalli, Mara; Stella, Maurizio; Sapino, Anna; Castagnoli, Carlotta

    2016-01-01

    Human Acellular Dermal Matrices (HADM) are employed in various reconstructive surgery procedures as scaffolds for autologous tissue regeneration. The aim of this project was to develop a new type of HADM for clinical use, composed of glycerolized reticular dermis decellularized through incubation and tilting in Dulbecco’s Modified Eagle’s Medium (DMEM). This manufacturing method was compared with a decellularization procedure already described in the literature, based on the use of sodium hydroxide (NaOH), on samples from 28 donors. Cell viability was assessed using an MTT assay and microbiological monitoring was performed on all samples processed after each step. Two surgeons evaluated the biomechanical characteristics of grafts of increasing thickness. The effects of the different decellularization protocols were assessed by means of histological examination and immunohistochemistry, and residual DNA after decellularization was quantified using a real-time TaqMan MGB probe. Finally, we compared the results of DMEM based decellularization protocol on reticular dermis derived samples with the results of the same protocol applied on papillary dermis derived grafts. Our experimental results indicated that the use of glycerolized reticular dermis after 5 weeks of treatment with DMEM results in an HADM with good handling and biocompatibility properties. PMID:26918526

  3. Decellularized human amniotic membrane: more is needed for an efficient dressing for protection of burns against antibiotic-resistant bacteria isolated from burn patients.

    PubMed

    Gholipourmalekabadi, M; Bandehpour, M; Mozafari, M; Hashemi, A; Ghanbarian, H; Sameni, M; Salimi, M; Gholami, M; Samadikuchaksaraei, A

    2015-11-01

    Human amniotic membranes (HAMs) have attracted the attention of burn surgeons for decades due to favorable properties such as their antibacterial activity and promising support of cell proliferation. On the other hand, as a major implication in the health of burn patients, the prevalence of bacteria resistant to multiple antibiotics is increasing due to overuse of antibiotics. The aim of this study was to investigate whether HAMs (both fresh and acellular) are an effective antibacterial agent against antibiotic-resistant bacteria isolated from burn patients. Therefore, a HAM was decellularized and tested for its antibacterial activity. Decellularization of the tissue was confirmed by hematoxylin and eosin (H&E) and 4,6-diamidino-2-phenylindole (DAPI) staining. In addition, the cyto-biocompatibility of the acellular HAM was proven by the cell viability test (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide, MTT) and scanning electron microscopy (SEM). The resistant bacteria were isolated from burns, identified, and tested for their susceptibility to antibiotics using both the antibiogram and polymerase chain reaction (PCR) techniques. Among the isolated bacteria, three blaIMP gene-positive Pseudomonas aeruginosa strains were chosen for their high resistance to the tested antibiotics. The antibacterial activity of the HAM was also tested for Klebsiella pneumoniae (American Type Culture Collection (ATCC) 700603) as a resistant ATCC bacterium; Staphylococcus aureus (mecA positive); and three standard strains of ATCC bacteria including Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27833), and S. aureus (ATCC 25923). Antibacterial assay revealed that only the latter three bacteria were susceptible to the HAM. All the data obtained from this study suggest that an alternative strategy is required to complement HAM grafting in order to fully protect burns from nosocomial infections.

  4. Human amniotic membrane as an alternative source of stem cells for regenerative medicine.

    PubMed

    Díaz-Prado, Silvia; Muiños-López, Emma; Hermida-Gómez, Tamara; Cicione, Claudia; Rendal-Vázquez, M Esther; Fuentes-Boquete, Isaac; de Toro, Francisco J; Blanco, Francisco J

    2011-03-01

    The human amniotic membrane (HAM) is a highly abundant and readily available tissue. This amniotic tissue has considerable advantageous characteristics to be considered as an attractive material in the field of regenerative medicine. It has low immunogenicity, anti-inflammatory properties and their cells can be isolated without the sacrifice of human embryos. Since it is discarded post-partum it may be useful for regenerative medicine and cell therapy. Amniotic membranes have already been used extensively as biologic dressings in ophthalmic, abdominal and plastic surgery. HAM contains two cell types, from different embryological origins, which display some characteristic properties of stem cells. Human amnion epithelial cells (hAECs) are derived from the embryonic ectoderm, while human amnion mesenchymal stromal cells (hAMSCs) are derived from the embryonic mesoderm. Both populations have similar immunophenotype and multipotential for in vitro differentiation into the major mesodermal lineages, however they differ in cell yield. Therefore, HAM has been proposed as a good candidate to be used in cell therapy or regenerative medicine to treat damaged or diseased tissues.

  5. [Human amniotic epithelium (HAE) as a possible source of stem cells (SC)].

    PubMed

    García-López, Guadalupe; García-Castro, Irma Lydia; Avila-González, Daniela; Molina-Hernández, Anayansi; Flores-Herrera, Héctor; Merchant-Larios, Horacio; Díaz-Martínez, Fabián

    2015-01-01

    There have been major recent advances in the field of developmental biology due to the investigation on stem cells (SC). Stem cells are characterized by their capacity of auto-renewal and differentiation to different cellular phenotypes. Based on the developmental stage, they can be classified into two different types: embryonic SCs and adult SCs. It has been widely reported that several problems need to be resolved before their possible clinical applications. As a result, fetal membranes have been suggested as an alternative source of SCs. In the human amniotic epithelium, the presence of markers of pluripotent SC´s has been reported, and its capacity as a feeder layer for expansion of different SC types. Also, fetal membranes are a discarded product after delivery, and thus there are not any ethical issues related to its use. In conclusion, the human amniotic epithelium can be a strong candidate for regenerative medicine.

  6. Isolation, Characterization, Cryopreservation of Human Amniotic Stem Cells and Differentiation to Osteogenic and Adipogenic Cells

    PubMed Central

    Gholizadeh-Ghaleh Aziz, Shiva; Pashaei-Asl, Fatima; Fardyazar, Zahra; Pashaiasl, Maryam

    2016-01-01

    Human stem cells and progenitor cells can be used to treat cancer and replace dysfunctional cells within a tissue or organ. The objective of this study was to identify the appropriate cells type in regenerative medicine and targeted therapy. As an alternative to embryonic and bone marrow stem cells, we examined human amniotic fluid stem cells (hAFSCs), one of the potential source of multipotent stem cells isolated from both cell pellet (using single-stage method), and supernatant of human amniotic fluid. Source of isolation and unique property of the cells emphasize that these cells are one of the promising new tools in therapeutic field. Double sources for isolation and availability of the left over samples in diagnostic laboratory at the same time have less legal and ethical concerns compared with embryonic stem cell studies. Cells were isolated, cultured for 18th passage for 6 months and characterized using qPCR and flow cytometry. Cells showed good proliferative ability in culture condition. The cells successfully differentiated into the adipogenic and osteogenic lineages. Based on these findings, amniotic fluid can be considered as an appropriate and convenient source of human amniotic fluid stem cells. These cells provide potential tools for therapeutic applications in the field of regenerative medicine. To get a better understanding of crosstalk between Oct4/NANOG with osteogenesis and adipogenesis, we used network analysis based on Common Targets algorithm and Common Regulators algorithm as well as subnetwork discovery based on gene set enrichment. Network analysis highlighted the possible role of MIR 302A and MIR let-7g. We demonstrated the high expression of MIR 302A and low expression of MIR let7g in hAFSCs by qPCR. PMID:27434028

  7. Human amniotic membrane as an intestinal patch for neomucosal growth in the rabbit model.

    PubMed

    Barlas, M; Gökçora, H; Erekul, S; Dindar, H; Yücesan, S

    1992-05-01

    This experiment was carried out as a preliminary study, an attempt to grow new intestinal mucosa on human amniotic membrane in the terminal ileum in 37 rabbits. After ketamin sulfate anesthesia at laparatomy, 5-cm ileal defects were patched with human amniotic membrane (5 x 2 cm). These patched intestines were investigated on the first postoperative day and the 2nd, 5th, 10th, and 20th weeks corresponding to 4, 5, 5, 10, and 10 rabbits, respectively. Only three rabbits died in the early postoperative period. There was no evidence of intestinal obstruction or dilatation with barium meal. Microscopically, the neomucosa consisted of a thin layer of columnar epithelial cells at 2 weeks with more maturity of the villi and less irregularity and branching by 20 weeks. All patches were covered with neomucosa commencing at 2 weeks and covering the whole patch area by 20 weeks. This technique's advantages are the large size and the ease of the availability of the human amniotic membrane for neonates at risk without jeopardizing the neonates tissues. It is hoped that this method might be considered when neonatal material is scarce.

  8. Recellularizing of human acellular dermal matrices imaged by high-definition optical coherence tomography.

    PubMed

    Boone, Marc A L M; Draye, Jean Pierre; Verween, Gunther; Aiti, Annalisa; Pirnay, Jean-Paul; Verbeken, Gilbert; De Vos, Daniel; Rose, Thomas; Jennes, Serge; Jemec, Gregor B E; Del Marmol, Veronique

    2015-05-01

    High-definition optical coherence tomography (HD-OCT) permits real-time 3D imaging of the impact of selected agents on human skin allografts. The real-time 3D HD-OCT assessment of (i) the impact on morphological and cellular characteristics of the processing of human acellular dermal matrices (HADMs) and (ii) repopulation of HADMs in vitro by human fibroblasts and remodelling of the extracellular matrix by these cells. Four different skin decellularization methods, Dispase II/Triton X-100, Dispase II/SDS (sodium dodecyl sulphate), NaCl/Triton X-100 and NaCl/SDS, were analysed by HD-OCT. HD-OCT features of epidermal removal, dermo-epidermal junction (DEJ) integrity, cellularity and dermal architecture were correlated with reflectance confocal microscopy (RCM), histopathology and immunohistochemistry. Human adult dermal fibroblasts were in vitro seeded on the NaCl/Triton X-100 processed HADMs, cultured up to 19 days and evaluated by HD-OCT in comparison with MTT proliferation test and histology. Epidermis was effectively removed by all treatments. DEJ was best preserved after NaCl/Triton X-100 treatment. Dispase II/SDS treatment seemed to remove all cellular debris in comparison with NaCl/Triton X-100 but disturbed the DEJ severely. The dermal micro-architectural structure and vascular spaces of (sub)papillary dermis were best preserved with the NaCl/Triton X-100. The impact on the 3D structure and vascular holes was detrimental with Dispase II/SDS. Elastic fibre fragmentation was only observed after Dispase II incubation. HD-OCT showed that NaCl/Triton X-100 processed matrices permitted in vitro repopulation by human dermal fibroblasts (confirmed by MTT test and histology) and underwent remodelling upon increasing incubation time. Care must be taken in choosing the appropriate processing steps to maintain selected properties of the extracellular matrix in HADMs. Processing HADMs with NaCl/Triton X-100 permits in vitro the proliferation and remodelling activity of

  9. Chondrogenesis of Human Infrapatellar Fat Pad Stem Cells on Acellular Dermal Matrix.

    PubMed

    Ye, Ken; Traianedes, Kathy; Choong, Peter F M; Myers, Damian E

    2016-01-01

    Acellular dermal matrix (ADM) has been in clinical use for decades in numerous surgical applications. The ability for ADM to promote cellular repopulation, revascularisation and tissue regeneration is well documented. Adipose stem cells have the ability to differentiate into mesenchymal tissue types, including bone and cartilage. The aim of this study was to investigate the potential interaction between ADM and adipose stem cells in vitro using TGFβ3 and BMP6. Human infrapatellar fat pad-derived adipose stem cells (IPFP-ASC) were cultured with ADM derived from rat dermis in chondrogenic (TGFβ3 and BMP6) medium in vitro for 2 and 4 weeks. Histology, qPCR, and immunohistochemistry were performed to assess for markers of chondrogenesis (collagen Type II, SOX9 and proteoglycans). At 4 weeks, cell-scaffold constructs displayed cellular changes consistent with chondrogenesis, with evidence of stratification of cell layers and development of a hyaline-like cartilage layer superficially, which stained positively for collagen Type II and proteoglycans. Significant cell-matrix interaction was seen between the cartilage layer and the ADM itself with seamless integration between each layer. Real time qPCR showed significantly increased COL2A1, SOX9, and ACAN gene expression over 4 weeks when compared to control. COL1A2 gene expression remained unchanged over 4 weeks. We believe that the principles that make ADM versatile and successful for tissue regeneration are applicable to cartilage regeneration. This study demonstrates in vitro the ability for IPFP-ASCs to undergo chondrogenesis, infiltrate, and interact with ADM. These outcomes serve as a platform for in vivo modelling of ADM for cartilage repair. PMID:26858950

  10. Chondrogenesis of Human Infrapatellar Fat Pad Stem Cells on Acellular Dermal Matrix

    PubMed Central

    Ye, Ken; Traianedes, Kathy; Choong, Peter F. M.; Myers, Damian E.

    2016-01-01

    Acellular dermal matrix (ADM) has been in clinical use for decades in numerous surgical applications. The ability for ADM to promote cellular repopulation, revascularisation and tissue regeneration is well documented. Adipose stem cells have the ability to differentiate into mesenchymal tissue types, including bone and cartilage. The aim of this study was to investigate the potential interaction between ADM and adipose stem cells in vitro using TGFβ3 and BMP6. Human infrapatellar fat pad-derived adipose stem cells (IPFP-ASC) were cultured with ADM derived from rat dermis in chondrogenic (TGFβ3 and BMP6) medium in vitro for 2 and 4 weeks. Histology, qPCR, and immunohistochemistry were performed to assess for markers of chondrogenesis (collagen Type II, SOX9 and proteoglycans). At 4 weeks, cell-scaffold constructs displayed cellular changes consistent with chondrogenesis, with evidence of stratification of cell layers and development of a hyaline-like cartilage layer superficially, which stained positively for collagen Type II and proteoglycans. Significant cell–matrix interaction was seen between the cartilage layer and the ADM itself with seamless integration between each layer. Real time qPCR showed significantly increased COL2A1, SOX9, and ACAN gene expression over 4 weeks when compared to control. COL1A2 gene expression remained unchanged over 4 weeks. We believe that the principles that make ADM versatile and successful for tissue regeneration are applicable to cartilage regeneration. This study demonstrates in vitro the ability for IPFP-ASCs to undergo chondrogenesis, infiltrate, and interact with ADM. These outcomes serve as a platform for in vivo modelling of ADM for cartilage repair. PMID:26858950

  11. Chondrogenesis of Human Infrapatellar Fat Pad Stem Cells on Acellular Dermal Matrix.

    PubMed

    Ye, Ken; Traianedes, Kathy; Choong, Peter F M; Myers, Damian E

    2016-01-01

    Acellular dermal matrix (ADM) has been in clinical use for decades in numerous surgical applications. The ability for ADM to promote cellular repopulation, revascularisation and tissue regeneration is well documented. Adipose stem cells have the ability to differentiate into mesenchymal tissue types, including bone and cartilage. The aim of this study was to investigate the potential interaction between ADM and adipose stem cells in vitro using TGFβ3 and BMP6. Human infrapatellar fat pad-derived adipose stem cells (IPFP-ASC) were cultured with ADM derived from rat dermis in chondrogenic (TGFβ3 and BMP6) medium in vitro for 2 and 4 weeks. Histology, qPCR, and immunohistochemistry were performed to assess for markers of chondrogenesis (collagen Type II, SOX9 and proteoglycans). At 4 weeks, cell-scaffold constructs displayed cellular changes consistent with chondrogenesis, with evidence of stratification of cell layers and development of a hyaline-like cartilage layer superficially, which stained positively for collagen Type II and proteoglycans. Significant cell-matrix interaction was seen between the cartilage layer and the ADM itself with seamless integration between each layer. Real time qPCR showed significantly increased COL2A1, SOX9, and ACAN gene expression over 4 weeks when compared to control. COL1A2 gene expression remained unchanged over 4 weeks. We believe that the principles that make ADM versatile and successful for tissue regeneration are applicable to cartilage regeneration. This study demonstrates in vitro the ability for IPFP-ASCs to undergo chondrogenesis, infiltrate, and interact with ADM. These outcomes serve as a platform for in vivo modelling of ADM for cartilage repair.

  12. [Differentiation of human amniotic fluid stem cells into cardiomyocytes through embryonic body formation].

    PubMed

    Wang, Han; Chen, Shuai; Cheng, Xiang; Dou, Zhongying; Wang, Huayan

    2008-09-01

    To isolate human amniotic fluid stem cells (hASCs) and induce hASCs into cardiomyocytes after forming the embryonic bodies. We cultivated hASCs isolated from the amniotic fluid continually for over 42 passages. The biological characteristics of hASCs were detected by immunocytochemistry, RT-PCR and flow cytometer, hASCs at 10-15th passage were suspension cultured to form embryonic bodies that were induced to cardiomyocytes. Fibroblastoid-type hASCs were obtained. Immunocytochemistry, RT-PCR and flow cytometry analysis demonstrated that hASCs were positive for some specific makers of the embryonic stem cell. hASCs could form embryonic bodies that were alkaline-phosphatase positive and expressed fgf5, zeta-globin and alpha-fetoprotein. The embryonic bodies could differentiate into cardiomyocytes showing alpha-actin positive and Tbx5, Nkx2.5, GATA4 and alpha-MHC positive. We conclued that hASCs obtained from human amniotic fluid could differentiate into cardiomyocytes through the formation of embryonic bodies. PMID:19160841

  13. [Differentiation of human amniotic fluid stem cells into cardiomyocytes through embryonic body formation].

    PubMed

    Wang, Han; Chen, Shuai; Cheng, Xiang; Dou, Zhongying; Wang, Huayan

    2008-09-01

    To isolate human amniotic fluid stem cells (hASCs) and induce hASCs into cardiomyocytes after forming the embryonic bodies. We cultivated hASCs isolated from the amniotic fluid continually for over 42 passages. The biological characteristics of hASCs were detected by immunocytochemistry, RT-PCR and flow cytometer, hASCs at 10-15th passage were suspension cultured to form embryonic bodies that were induced to cardiomyocytes. Fibroblastoid-type hASCs were obtained. Immunocytochemistry, RT-PCR and flow cytometry analysis demonstrated that hASCs were positive for some specific makers of the embryonic stem cell. hASCs could form embryonic bodies that were alkaline-phosphatase positive and expressed fgf5, zeta-globin and alpha-fetoprotein. The embryonic bodies could differentiate into cardiomyocytes showing alpha-actin positive and Tbx5, Nkx2.5, GATA4 and alpha-MHC positive. We conclued that hASCs obtained from human amniotic fluid could differentiate into cardiomyocytes through the formation of embryonic bodies.

  14. Hepatic differentiation of human pluripotent stem cells on human liver progenitor HepaRG-derived acellular matrix.

    PubMed

    Kanninen, Liisa K; Porola, Pauliina; Niklander, Johanna; Malinen, Melina M; Corlu, Anne; Guguen-Guillouzo, Christiane; Urtti, Arto; Yliperttula, Marjo L; Lou, Yan-Ru

    2016-02-15

    Human hepatocytes are extensively needed in drug discovery and development. Stem cell-derived hepatocytes are expected to be an improved and continuous model of human liver to study drug candidates. Generation of endoderm-derived hepatocytes from human pluripotent stem cells (hPSCs), including human embryonic stem cells and induced pluripotent stem cells, is a complex, challenging process requiring specific signals from soluble factors and insoluble matrices at each developmental stage. In this study, we used human liver progenitor HepaRG-derived acellular matrix (ACM) as a hepatic progenitor-specific matrix to induce hepatic commitment of hPSC-derived definitive endoderm (DE) cells. The DE cells showed much better attachment to the HepaRG ACM than other matrices tested and then differentiated towards hepatic cells, which expressed hepatocyte-specific makers. We demonstrate that Matrigel overlay induced hepatocyte phenotype and inhibited biliary epithelial differentiation in two hPSC lines studied. In conclusion, our study demonstrates that the HepaRG ACM, a hepatic progenitor-specific matrix, plays an important role in the hepatic differentiation of hPSCs.

  15. Immunosuppressive activity of human amniotic fluid of normal and abnormal pregnancies.

    PubMed

    Shohat, B; Faktor, J M

    1988-01-01

    Twenty specimens of amniotic fluid (AF) obtained between week 16 and 18 of gestation from normal pregnant women and six specimens from pregnant women in which trisomia of chromosome 21 was found were tested for immunosuppressive activity. Incubation of normal human donor lymphocytes with 0.2-1 mL of AF from normal pregnant women for one hour at 37 degrees C was sufficient for induction of significant inhibition of the ability of these cells to induce a local xenogeneic graft-versus-host reaction (GVHR) as well as inhibition of E and E-active rosette formation, the GVHR being the most sensitive test. On the other hand, amniotic fluid obtained from the six pregnant women in which trisomia of chromosome 21 was found showed no inhibitory activity in either the E or E-active rosette formation, nor in the local xenogeneic graft-versus-host reaction. AF from all the women tested was found to have no effect on phenotype expression of the lymphocytes, as tested by the monoclonal antibodies OKT4+ and OKT8+, nor on B-lymphocytes, as tested by surface immunoglobulins. No correlation was found between the alpha-fetoprotein levels in the sera of those women and the immunosuppressive activity. These findings indicate that genetic defects of the conceptus are not limited to the embryo but may affect the composition of immunosuppressive components present in normal amniotic fluid.

  16. Corneal haze induced by excimer laser photoablation in rabbits is reduced by preserved human amniotic membrane graft

    NASA Astrophysics Data System (ADS)

    Wang, Ming X.; Gray, Trevor; Prabhasawat, Pinnita; Ma, Xiong; Culbertson, William; Forster, Richard; Hanna, Khalil; Tseng, Scheffer C. G.

    1998-06-01

    We conducted a study to determine if preserved human amniotic membrane can reduce corneal haze induced by excimer laser photoablation. Excimer photoablation was performed bilaterally on 40 New Zealand white rabbits with a 6 mm ablation zone and 120 micrometer depth (PTK) using the VISX Star. One eye was randomly covered with a preserved human amniotic membrane and secured using four interrupted 10 - 0 nylon sutures; the other eye served as control. The amniotic membranes were removed at one week, and the corneal haze was graded with a slit-lamp biomicroscopy by three masked corneal specialists (WC, KH and RF) biweekly for the ensuing 12 weeks. Histology and in situ TUNEL staining (for fragmented DNA as an index for apoptosis) was performed at days 1, 3 and 7 and at 12 weeks. One week after excimer photoablation, the amniotic membrane-covered corneas showed more anterior stromal edema, which resolved at the second week. A consistent grading of organized reticular corneal haze was noted among the three masked observers. Such corneal haze peaked at the seventh week in both groups. The amniotic membrane-covered group showed statistically significant less corneal haze (0.50 plus or minus 0.15) than the control groups (1.25 plus or minus 0.35) (p less than 0.001). The amniotic membrane-covered corneas had less inflammatory response at days 1 and 3, showing nearly nil DNA fragmentation on keratocytes on the ablated anterior stromal and less stromal fibroblast activation. There is less altered epithelial cell morphology and less epithelial hyperplasia at 1 week in these amniotic membrane-treated eyes. We concluded from this study that amniotic membrane matrix is effective in reducing corneal haze induced by excimer photoablation in rabbits and may have clinical applications.

  17. Isolation of c-Kit+ human amniotic fluid stem cells from second trimester.

    PubMed

    Pozzobon, Michela; Piccoli, Martina; Schiavo, Andrea Alex; Atala, Anthony; De Coppi, Paolo

    2013-01-01

    Amniotic fluid-derived stem (AFS) cells have been described as an appealing source of stem cells because of their (1) fetal, non-embryonic origin, (2) easy access during pregnancy overcoming the ethical issues related both to the use of human embryonic cells and to the postnatal tissue biopsy with donor site morbidity, and (3) their undemanding ability to be expanded. We and others have demonstrated the broad differentiation potential and here we describe the established protocol we developed to obtain c-Kit+ human AFS cells, starting from second trimester amniocentesis samples.

  18. Functional Human Podocytes Generated in Organoids from Amniotic Fluid Stem Cells.

    PubMed

    Xinaris, Christodoulos; Benedetti, Valentina; Novelli, Rubina; Abbate, Mauro; Rizzo, Paola; Conti, Sara; Tomasoni, Susanna; Corna, Daniela; Pozzobon, Michela; Cavallotti, Daniela; Yokoo, Takashi; Morigi, Marina; Benigni, Ariela; Remuzzi, Giuseppe

    2016-05-01

    Generating kidney organoids using human stem cells could offer promising prospects for research and therapeutic purposes. However, no cell-based strategy has generated nephrons displaying an intact three-dimensional epithelial filtering barrier. Here, we generated organoids using murine embryonic kidney cells, and documented that these tissues recapitulated the complex three-dimensional filtering structure of glomerular slits in vivo and accomplished selective glomerular filtration and tubular reabsorption. Exploiting this technology, we mixed human amniotic fluid stem cells with mouse embryonic kidney cells to establish three-dimensional chimeric organoids that engrafted in vivo and grew to form vascularized glomeruli and tubular structures. Human cells contributed to the formation of glomerular structures, differentiated into podocytes with slit diaphragms, and internalized exogenously infused BSA, thus attaining in vivo degrees of specialization and function unprecedented for donor stem cells. In conclusion, human amniotic fluid stem cell chimeric organoids may offer new paths for studying renal development and human podocyte disease, and for facilitating drug discovery and translational research.

  19. Acellular porcine corneal matrix as a carrier scaffold for cultivating human corneal epithelial cells and fibroblasts in vitro

    PubMed Central

    Zhang, Ju; Zhang, Can-Wei; Du, Li-Qun; Wu, Xin-Yi

    2016-01-01

    AIM To investigate the feasibility of corneal anterior lamellar reconstruction with human corneal epithelial cells and fibroblasts, and an acellular porcine cornea matrix (APCM) in vitro. METHODS The scaffold was prepared from fresh porcine corneas which were treated with 0.5% sodium dodecyl sulfate (SDS) solution and the complete removal of corneal cells was confirmed by hematoxylin-eosin (HE) staining and 4′, 6-diamidino-2-phenylindole (DAPI) staining. Human corneal fibroblasts and epithelial cells were cultured with leaching liquid extracted from APCM, and then cell proliferative ability was evaluated by MTT assay. To construct a human corneal anterior lamellar replacement, corneal fibroblasts were injected into the APCM and cultured for 3d, followed by culturing corneal epithelial cells on the stroma construction surface for another 10d. The corneal replacement was analyzed by HE staining, and immunofluorescence staining. RESULTS Histological examination indicated that there were no cells in the APCM by HE staining, and DAPI staining did not detect any residual DNA. The leaching liquid from APCM had little influence on the proliferation ability of human corneal fibroblasts and epithelial cells. At 10d, a continuous 3 to 5 layers of human corneal epithelial cells covering the surface of the APCM was observed, and the injected corneal fibroblasts distributed within the scaffold. The phenotype of the construction was similar to normal human corneas, with high expression of cytokeratin 12 in the epithelial cell layer and high expression of vimentin in the stroma. CONCLUSION Corneal anterior lamellar replacement can be reconstructed in vitro by cultivating human corneal epithelial cells and fibroblasts with an acellular porcine cornea matrix. This laid the foundation for the further transplantation in vivo. PMID:26949602

  20. Isolation of Human Neural Stem Cells from the Amniotic Fluid with Diagnosed Neural Tube Defects.

    PubMed

    Chang, Yu-Jen; Su, Hong-Lin; Hsu, Lee-Feng; Huang, Po-Jui; Wang, Tzu-Hao; Cheng, Fu-Chou; Hsu, Li-Wen; Tsai, Ming-Song; Chen, Chih-Ping; Chang, Yao-Lung; Chao, An-Shine; Hwang, Shiaw-Min

    2015-08-01

    Human neural stem cells (NSCs) are particularly valuable for the study of neurogenesis process and have a therapeutic potential in treating neurodegenerative disorders. However, current progress in the use of human NSCs is limited due to the available NSC sources and the complicated isolation and culture techniques. In this study, we describe an efficient method to isolate and propagate human NSCs from the amniotic fluid with diagnosed neural tube defects (NTDs), specifically, anencephaly. These amniotic fluid-derived NSCs (AF-NSCs) formed neurospheres and underwent long-term expansion in vitro. In addition, these cells showed normal karyotypes and telomerase activity and expressed NSC-specific markers, including Nestin, Sox2, Musashi-1, and the ATP-binding cassette G2 (ABCG2). AF-NSCs displayed typical morphological patterns and expressed specific markers that were consistent with neurons, astrocytes, oligodendrocytes, and dopaminergic neurons after proper induction conditions. Furthermore, grafted AF-NSCs improved the physiological functions in a rat stroke model. The ability to isolate and bank human NSCs from this novel source provides a unique opportunity for translational studies of neurological disorders. PMID:25923707

  1. Amniotic Fluid Stem Cells: A Novel Source for Modeling of Human Genetic Diseases

    PubMed Central

    Antonucci, Ivana; Provenzano, Martina; Rodrigues, Melissa; Pantalone, Andrea; Salini, Vincenzo; Ballerini, Patrizia; Borlongan, Cesar V.; Stuppia, Liborio

    2016-01-01

    In recent years, great interest has been devoted to the use of Induced Pluripotent Stem cells (iPS) for modeling of human genetic diseases, due to the possibility of reprogramming somatic cells of affected patients into pluripotent cells, enabling differentiation into several cell types, and allowing investigations into the molecular mechanisms of the disease. However, the protocol of iPS generation still suffers from technical limitations, showing low efficiency, being expensive and time consuming. Amniotic Fluid Stem cells (AFS) represent a potential alternative novel source of stem cells for modeling of human genetic diseases. In fact, by means of prenatal diagnosis, a number of fetuses affected by chromosomal or Mendelian diseases can be identified, and the amniotic fluid collected for genetic testing can be used, after diagnosis, for the isolation, culture and differentiation of AFS cells. This can provide a useful stem cell model for the investigation of the molecular basis of the diagnosed disease without the necessity of producing iPS, since AFS cells show some features of pluripotency and are able to differentiate in cells derived from all three germ layers “in vitro”. In this article, we describe the potential benefits provided by using AFS cells in the modeling of human genetic diseases. PMID:27110774

  2. Human amniotic epithelial cells as feeder layer to derive and maintain human embryonic stem cells from poor-quality embryos.

    PubMed

    Ávila-González, Daniela; Vega-Hernández, Eva; Regalado-Hernández, Juan Carlos; De la Jara-Díaz, Julio Francisco; García-Castro, Irma Lydia; Molina-Hernández, Anayansi; Moreno-Verduzco, Elsa Romelia; Razo-Aguilera, Guadalupe; Flores-Herrera, Héctor; Portillo, Wendy; Díaz-Martínez, Néstor Emmanuel; García-López, Guadalupe; Díaz, Néstor Fabián

    2015-09-01

    Data from the literature suggest that human embryonic stem cell (hESC) lines used in research do not genetically represent all human populations. The derivation of hESC through conventional methods involve the destruction of viable human embryos, as well the use of mouse embryonic fibroblasts as a feeder layer, which has several drawbacks. We obtained the hESC line (Amicqui-1) from poor-quality (PQ) embryos derived and maintained on human amniotic epithelial cells (hAEC). This line displays a battery of markers of pluripotency and we demonstrated the capacity of these cells to produce derivates of the three germ layers.

  3. The cell mediated and humoral immune response to vaccination with acellular and whole cell pertussis vaccine in adult humans.

    PubMed

    Petersen, J W; Ibsen, P H; Bentzon, M W; Capiau, C; Heron, I

    1991-10-01

    The cell mediated immune response (CMI) against pertussis antigens following vaccination with the traditional Danish whole cell pertussis vaccine (WC-P) and the Japanese acellular pertussis vaccine (A-PV) JNIH-3 was studied in four adult human volunteers. Vaccination with the A-PV induced an in vitro proliferative response of peripheral blood lymphocytes to pertussis toxin (PT) subunits S2-S4, S3-S4 and S5 and the filamentous hemagglutinin (FHA), and a better serological response to native PT, detoxified PT (dPT) and FHA than the WC-PV. The induced CMI and serological response were followed over a period of 17 weeks, and were not seen to decline during this period. Further, an in vitro proliferative response to Bordetella pertussis agglutinogen 2 and 3 were demonstrated using lymphocytes from recently and not-so-recently pertussis-vaccinated adults. PMID:1797049

  4. Evaluation of repair in duodenal perforation with human amniotic membrane: An animal model (dog)

    PubMed Central

    Ghahramani, Leila; Jahromi, Ali Bagherpour; Dehghani, Mohammad Reza; Ashraf, Mohammad Javad; Rahimikazerooni, Salar; Rezaianzadeh, Abbas; Safarpour, Ali Reza; Hosseini, Seyed Vahid

    2014-01-01

    Background: There is a growing tendency toward application of human amniotic membrane (HAM) as a biologic substitute in various tissue injuries where a significant tissue loss is a matter of concern. In gastrointestinal injuries especially duodenal ones, some potential limitations in current surgical techniques contribute to not fully acceptable healing outcomes. Thus, this study was carried out to assess repair with HAM patch for duodenal defect in comparison with simple duodenoraphy in an animal model (dog). Materials and Methods: A total of 15 male German shepherd dogs weighing 23-27 kg were randomly divided into two groups. Group A with 10 dogs, which were a candidate for duodenal repair by amniotic membrane patch and Group B consisted of 5 dogs perform simple duodenorraphy. A precise control was made to match all conditions except surgical technique. Macroscopic and microscopic features of the healed duodenal lumen in both groups were recorded. Results: Gross evaluation revealed no difference in luminal diameter in both groups. Statistical analysis of duodenal diameter between both groups after operation also showed no significant difference (Pv = 0.789). Histological assessment indicated less inflammation with better wound healing in Group A. Conclusion: It seems that repairing duodenal wall defect with HAM would result in better histological outcomes compared with what is seen in simple duodenoraphy in animal models. However, there is no significant difference regarding surgical findings. PMID:24804187

  5. AFM studies of cellular mechanics during osteogenic differentiation of human amniotic fluid-derived stem cells.

    PubMed

    Chen, Qian; Xiao, Pan; Chen, Jia-Nan; Cai, Ji-Ye; Cai, Xiao-Fang; Ding, Hui; Pan, Yun-Long

    2010-01-01

    Amniotic fluid-derived stem cells (AFSCs) are becoming an important source of cells for regenerative medicine given with apparent advantages of accessibility, renewal capacity and multipotentiality. In this study, the mechanical properties of human amniotic fluid-derived stem cells (hAFSCs), such as the average Young's modulus, were determined by atomic force microscopy (3.97 ± 0.53 kPa for hAFSCs vs. 1.52 ± 0.63 kPa for fully differentiated osteoblasts). These differences in cell elasticity result primarily from differential actin cytoskeleton organization in these two cell types. Furthermore, ultrastructures, nanostructural details on the surface of cell, were visualized by atomic force microscopy (AFM). It was clearly shown that surface of osteoblasts were covered by mineralized particles, and the histogram of particles size showed that most of the particles on the surface of osteoblasts distributed from 200 to 400 nm in diameter, while the diameter of hAFSCs particles ranged from 100 to 200 nm. In contrast, there were some dips on the surface of hAFSCs, and particles were smaller than that of osteoblasts. Additionally, as osteogenic differentiation of hAFSCs progressed, more and more stress fibers were replaced by a thinner actin network which is characteristic of mature osteoblasts. These results can improve our understanding of the mechanical properties of hAFSCs during osteogenic differentiation. AFM can be used as a powerful tool for detecting ultrastructures and mechanical properties.

  6. The development of a radioimmunoassay for reverse triiodothyronine sulfate in human serum and amniotic fluid

    SciTech Connect

    Wu, Sing-Yung ); Huang, Wen-Sheng; Chen, Wei-Lian ); Polk, D.; Reviczky, A.; Williams, J. III; Chopra, I.J.; Fisher, D.A. )

    1993-06-01

    Sulfated iodothyronines including T[sub 4]-sulfate (T[sub 4]S) and T[sub 3]-sulfate (T[sub 3]S) have been identified in human serum and amniotic fluid. Little is know, however, about the existence of sulfate conjugation of reverse T[sub 3] (rT[sub 3]S) in man. In this report, the authors employed a novel, sensitive, and specific rT[sub 3]S RIA to address this question. The rabbit antiserum to rT[sub 3]S was highly specific; T[sub 4], T[sub 3], rT[sub 3], and 3,3'-T[sub 2] showed less than 0.002% cross-reaction with the antiserum. Only T[sub 4]S and T[sub 3]S cross-reacted significantly (0.3% and 0.01%, respectively); other analogs cross-reacted less than 0.0001%. The detection threshold of the RIA was 14 pmol/L (1.0 ng/dL). The mean serum rT[sub 3]S concentration (pmol/L) was 40 in euthyroid subjects. Values were similar in hypothyroid patients (38) and pregnant women (52) but significantly (P < 0.01) elevated to 176 in hyperthyroid patient, 74 in patients with nonthyroid illnesses, and 684 in cord sera of newborns. Serum rT[sub 3]S increased significantly in hyperthyroid patients 1 day after administration of 1 g sodium ipodate orally. Reverse T[sub 3]S was detected consistently in amniotic fluid at 14 to 22 weeks of gestation and showed a marked rise 1-3 weeks after intraamniotic administration of 500-1000 [mu]g T[sub 4]. The various data suggest that : (1) rT[sub 3]S is a normal component of human serum and amniotic fluid; (2) it is derived from metabolism of T[sub 4] or rT[sub 3]; (3) circulating rT[sub 3]S increases in hyperthyroidism and in circumstances where type I 5'-monodeiodinating activity is low, e.g. nonthyroid illnesses, fetal life, and after administration of ipodate. 20 refs., 4 figs.

  7. Acellular human glans extracellular matrix as a scaffold for tissue engineering: in vitro cell support and biocompatibility

    PubMed Central

    Egydio, Fernanda M.; Freitas, Luiz G.; Sayeg, Kleber; Laks, Marcus; Oliveira, Andréia S.; Almeida, Fernando G.

    2015-01-01

    ABSTRACT Objectives: Diseases of the genitourinary tract can lead to significant damage. Current reconstructive techniques are limited by tissue availability and compatibility. This study aims to assess if the decellularized human glans can be used as a biomaterial for penile reconstruction. Materials and Methods: Samples of the glans matrices were descellularized. We evaluate the presence of collagen type I and III, and elastic fibers. Biocompatibility assays were performed to assess the cytotoxic and non-cytotoxic interactions between the acellular matrix and 3T3 cells. The matrices were seeded with mesenchymal stem cells and were assessed for viability and integration of these cells. Biomechanical tests in native tissue, descellularized matrix and seeded matrix were performed to characterize their biomechanical properties. Results: The tissue architecture of the decellularized matrix of human glans was preserved as well as the maintenance of the biomechanical and biological properties. The analyzes of glans seeded with mesenchymal stem cells revealed the integration of these cells to the matrices, and its viability during two weeks “in vitro”. Conclusion: The decellularization process did not alter the biological and biomechanical characteristics of the human glans. When these matrices were seeded they were able to maintain the cells integrity and vitality. PMID:26689526

  8. Human transgene-free amniotic-fluid-derived induced pluripotent stem cells for autologous cell therapy.

    PubMed

    Jiang, Guihua; Di Bernardo, Julie; Maiden, Michael M; Villa-Diaz, Luis G; Mabrouk, Omar S; Krebsbach, Paul H; O'Shea, K Sue; Kunisaki, Shaun M

    2014-11-01

    The establishment of a reliable prenatal source of autologous, transgene-free progenitor cells has enormous potential in the development of regenerative-medicine-based therapies for infants born with devastating birth defects. Here, we show that a largely CD117-negative population of human amniotic fluid mesenchymal stromal cells (AF-MSCs) obtained from fetuses with or without prenatally diagnosed anomalies are readily abundant and have limited baseline differentiation potential when compared with bone-marrow-derived MSCs and other somatic cell types. Nonetheless, the AF-MSCs could be easily reprogrammed into induced pluripotent stem cells (iPSCs) using nonintegrating Sendai viral vectors encoding for OCT4, SOX2, KLF4, and cMYC. The iPSCs were virtually indistinguishable from human embryonic stem cells in multiple assays and could be used to generate a relatively homogeneous population of neural progenitors, expressing PAX6, SOX2, SOX3, Musashi-1, and PSA-NCAM, for potential use in neurologic diseases. Further, these neural progenitors showed engraftment potential in vivo and were capable of differentiating into mature neurons and astrocytes in vitro. This study demonstrates the usefulness of AF-MSCs as an excellent source for the generation of human transgene-free iPSCs ideally suited for autologous perinatal regenerative medicine applications. PMID:25014361

  9. Adenovirus E1A/E1B Transformed Amniotic Fluid Cells Support Human Cytomegalovirus Replication

    PubMed Central

    Krömmelbein, Natascha; Wiebusch, Lüder; Schiedner, Gudrun; Büscher, Nicole; Sauer, Caroline; Florin, Luise; Sehn, Elisabeth; Wolfrum, Uwe; Plachter, Bodo

    2016-01-01

    The human cytomegalovirus (HCMV) replicates to high titers in primary human fibroblast cell cultures. A variety of primary human cells and some tumor-derived cell lines do also support permissive HCMV replication, yet at low levels. Cell lines established by transfection of the transforming functions of adenoviruses have been notoriously resistant to HCMV replication and progeny production. Here, we provide first-time evidence that a permanent cell line immortalized by adenovirus type 5 E1A and E1B (CAP) is supporting the full HCMV replication cycle and is releasing infectious progeny. The CAP cell line had previously been established from amniotic fluid cells which were likely derived from membranes of the developing fetus. These cells can be grown under serum-free conditions. HCMV efficiently penetrated CAP cells, expressed its immediate-early proteins and dispersed restrictive PML-bodies. Viral DNA replication was initiated and viral progeny became detectable by electron microscopy in CAP cells. Furthermore, infectious virus was released from CAP cells, yet to lower levels compared to fibroblasts. Subviral dense bodies were also secreted from CAP cells. The results show that E1A/E1B expression in transformed cells is not generally repressive to HCMV replication and that CAP cells may be a good substrate for dense body based vaccine production. PMID:26848680

  10. Decreased Laminin Expression by Human Lung Epithelial Cells and Fibroblasts Cultured in Acellular Lung Scaffolds from Aged Mice

    PubMed Central

    Godin, Lindsay M.; Sandri, Brian J.; Wagner, Darcy E.; Meyer, Carolyn M.; Price, Andrew P.; Akinnola, Ifeolu; Weiss, Daniel J.; Panoskaltsis-Mortari, Angela

    2016-01-01

    The lung changes functionally and structurally with aging. However, age-related effects on the extracellular matrix (ECM) and corresponding effects on lung cell behavior are not well understood. We hypothesized that ECM from aged animals would induce aging-related phenotypic changes in healthy inoculated cells. Decellularized whole organ scaffolds provide a powerful model for examining how ECM cues affect cell phenotype. The effects of age on ECM composition in both native and decellularized mouse lungs were assessed as was the effect of young vs old acellular ECM on human bronchial epithelial cells (hBECs) and lung fibroblasts (hLFs). Native aged (1 year) lungs demonstrated decreased expression of laminins α3 and α4, elastin and fibronectin, and elevated collagen, compared to young (3 week) lungs. Proteomic analyses of decellularized ECM demonstrated similar findings, and decellularized aged lung ECM contained less diversity in structural proteins compared to young ECM. When seeded in old ECM, hBECs and hLFs demonstrated lower gene expression of laminins α3 and α4, respectively, as compared to young ECM, paralleling the laminin deficiency of aged ECM. ECM changes appear to be important factors in potentiating aging-related phenotypes and may provide clues to mechanisms that allow for aging-related lung diseases. PMID:26954258

  11. Stem cells from human amniotic fluid exert immunoregulatory function via secreted indoleamine 2,3-dioxygenase1.

    PubMed

    Romani, Rita; Pirisinu, Irene; Calvitti, Mario; Pallotta, Maria Teresa; Gargaro, Marco; Bistoni, Giovanni; Vacca, Carmine; Di Michele, Alessandro; Orabona, Ciriana; Rosati, Jessica; Pirro, Matteo; Giovagnoli, Stefano; Matino, Davide; Prontera, Paolo; Rosi, Gabriella; Grohmann, Ursula; Talesa, Vincenzo N; Donti, Emilio; Puccetti, Paolo; Fallarino, Francesca

    2015-07-01

    Although human amniotic fluid does contain different populations of foetal-derived stem cells, scanty information is available on the stemness and the potential immunomodulatory activity of in vitro expanded, amniotic fluid stem cells. By means of a methodology unrequiring immune selection, we isolated and characterized different stem cell types from second-trimester human amniotic fluid samples (human amniotic fluid stem cells, HASCs). Of those populations, one was characterized by a fast doubling time, and cells were thus designated as fHASCs. Cells maintained their original phenotype under prolonged in vitro passaging, and they were able to originate embryoid bodies. Moreover, fHASCs exhibited regulatory properties when treated with interferon (IFN)-γ, including induction of the immunomodulatory enzyme indoleamine 2,3-dioxygenase 1 (IDO1). On coculture with human peripheral blood mononuclear cells, IFN-γ-treated fHASCs caused significantly decreased T-cell proliferation and increased frequency in CD4(+)  CD25(+)  FOXP3(+) regulatory T cells. Both effects required an intact IDO1 function and were cell contact-independent. An unprecedented finding in our study was that purified vesicles from IFN-γ-treated fHASCs abundantly expressed the functional IDO1 protein, and those vesicles were endowed with an fHASC-like regulatory function. In vivo, fHASCs were capable of immunoregulatory function, promoting allograft survival in a mouse model of allogeneic skin transplantation. This was concurrent with the expansion of CD4(+)  CD25(+)  Foxp3(+) T cells in graft-draining lymph nodes from recipient mice. Thus fHASCs, or vesicles thereof, may represent a novel opportunity for immunoregulatory maneuvers both in vitro and in vivo.

  12. Stem cells from human amniotic fluid exert immunoregulatory function via secreted indoleamine 2,3-dioxygenase1

    PubMed Central

    Romani, Rita; Pirisinu, Irene; Calvitti, Mario; Pallotta, Maria Teresa; Gargaro, Marco; Bistoni, Giovanni; Vacca, Carmine; Di Michele, Alessandro; Orabona, Ciriana; Rosati, Jessica; Pirro, Matteo; Giovagnoli, Stefano; Matino, Davide; Prontera, Paolo; Rosi, Gabriella; Grohmann, Ursula; Talesa, Vincenzo N; Donti, Emilio; Puccetti, Paolo; Fallarino, Francesca

    2015-01-01

    Although human amniotic fluid does contain different populations of foetal-derived stem cells, scanty information is available on the stemness and the potential immunomodulatory activity of in vitro expanded, amniotic fluid stem cells. By means of a methodology unrequiring immune selection, we isolated and characterized different stem cell types from second-trimester human amniotic fluid samples (human amniotic fluid stem cells, HASCs). Of those populations, one was characterized by a fast doubling time, and cells were thus designated as fHASCs. Cells maintained their original phenotype under prolonged in vitro passaging, and they were able to originate embryoid bodies. Moreover, fHASCs exhibited regulatory properties when treated with interferon (IFN)-γ, including induction of the immunomodulatory enzyme indoleamine 2,3-dioxygenase 1 (IDO1). On coculture with human peripheral blood mononuclear cells, IFN-γ–treated fHASCs caused significantly decreased T-cell proliferation and increased frequency in CD4+ CD25+ FOXP3+ regulatory T cells. Both effects required an intact IDO1 function and were cell contact-independent. An unprecedented finding in our study was that purified vesicles from IFN-γ–treated fHASCs abundantly expressed the functional IDO1 protein, and those vesicles were endowed with an fHASC-like regulatory function. In vivo, fHASCs were capable of immunoregulatory function, promoting allograft survival in a mouse model of allogeneic skin transplantation. This was concurrent with the expansion of CD4+ CD25+ Foxp3+ T cells in graft-draining lymph nodes from recipient mice. Thus fHASCs, or vesicles thereof, may represent a novel opportunity for immunoregulatory maneuvers both in vitro and in vivo. PMID:25783564

  13. Cerebroside Sulfatase Activity in Cultivated Human Skin Fibroblasts and Amniotic Fluid Cells

    ERIC Educational Resources Information Center

    Booth, Carol W.; And Others

    1975-01-01

    Prenatal monitoring for metachromatic leukodystrophy (a fatal inherited metabolic disorder) suggested that the determination of levels of cerebroside sulfatase in the amniotic fluid helped in the prenatal detection of this disorder. (DB)

  14. A Comparison of Culture Characteristics between Human Amniotic Mesenchymal Stem Cells and Dental Stem Cells.

    PubMed

    Yusoff, Nurul Hidayat; Alshehadat, Saaid Ayesh; Azlina, Ahmad; Kannan, Thirumulu Ponnuraj; Hamid, Suzina Sheikh Abdul

    2015-04-01

    In the past decade, the field of stem cell biology is of major interest among researchers due to its broad therapeutic potential. Stem cells are a class of undifferentiated cells that are able to differentiate into specialised cell types. Stem cells can be classified into two main types: adult stem cells (adult tissues) and embryonic stem cells (embryos formed during the blastocyst phase of embryological development). This review will discuss two types of adult mesenchymal stem cells, dental stem cells and amniotic stem cells, with respect to their differentiation lineages, passage numbers and animal model studies. Amniotic stem cells have a greater number of differentiation lineages than dental stem cells. On the contrary, dental stem cells showed the highest number of passages compared to amniotic stem cells. For tissue regeneration based on animal studies, amniotic stem cells showed the shortest time to regenerate in comparison with dental stem cells.

  15. Human amniotic membrane transplantation: Different modalities of its use in ophthalmology

    PubMed Central

    Malhotra, Chintan; Jain, Arun K

    2014-01-01

    The amniotic membrane (AM) is the inner layer of the fetal membranes and consist of 3 different layers: the epithelium, basement membrane and stroma which further consists of three contiguous but distinct layers: the inner compact layer, middle fibroblast layer and the outermost spongy layer. The AM has been shown to have anti-inflammatory, anti-fibrotic, anti-angiogenic as well as anti-microbial properties. Also because of its transparent structure, lack of immunogenicity and the ability to provide an excellent substrate for growth, migration and adhesion of epithelial corneal and conjunctival cells, it is being used increasingly for ocular surface reconstruction in a variety of ocular pathologies including corneal disorders associated with limbal stem cell deficiency, surgeries for conjunctival reconstruction, as a carrier for ex vivo expansion of limbal epithelial cells, glaucoma surgeries and sceral melts and perforations. However indiscriminate use of human AM needs to be discouraged as complications though infrequent can occur. These include risk of transmission of bacterial, viral or fungal infections to the recipient if the donors are not adequately screened for communicable diseases, if the membrane is not processed under sterile conditions or if storage is improper. Optimal outcomes can be achieved only with meticulous case selection. This review explores the ever expanding ophthalmological indications for the use of human AM. PMID:25032100

  16. Nukbone® promotes proliferation and osteoblastic differentiation of mesenchymal stem cells from human amniotic membrane

    SciTech Connect

    Rodríguez-Fuentes, Nayeli; Rodríguez-Hernández, Ana G.; Enríquez-Jiménez, Juana; Alcántara-Quintana, Luz E.; Fuentes-Mera, Lizeth; Piña-Barba, María C.; Zepeda-Rodríguez, Armando; and others

    2013-05-10

    Highlights: •Nukbone showed to be a good scaffold for adhesion, proliferation and differentiation of stem cells. •Nukbone induced osteoblastic differentiation of human mesenchymal stem cells. •Results showed that Nukbone offer an excellent option for bone tissue regeneration due to properties. -- Abstract: Bovine bone matrix Nukbone® (NKB) is an osseous tissue-engineering biomaterial that retains its mineral and organic phases and its natural bone topography and has been used as a xenoimplant for bone regeneration in clinics. There are not studies regarding its influence of the NKB in the behavior of cells during the repairing processes. The aim of this research is to demonstrate that NKB has an osteoinductive effect in human mesenchymal stem cells from amniotic membrane (AM-hMSCs). Results indicated that NKB favors the AM-hMSCs adhesion and proliferation up to 7 days in culture as shown by the scanning electron microscopy and proliferation measures using an alamarBlue assay. Furthermore, as demonstrated by reverse transcriptase polymerase chain reaction, it was detected that two gene expression markers of osteoblastic differentiation: the core binding factor and osteocalcin were higher for AM-hMSCs co-cultured with NKB in comparison with cultivated cells in absence of the biomaterial. As the results indicate, NKB possess the capability for inducing successfully the osteoblastic differentiation of AM-hMSC, so that, NKB is an excellent xenoimplant option for repairing bone tissue defects.

  17. Nuclear Nox4 Role in Stemness Power of Human Amniotic Fluid Stem Cells

    PubMed Central

    Maraldi, Tullia; Guida, Marianna; Zavatti, Manuela; Resca, Elisa; Bertoni, Laura; La Sala, Giovanni B.; De Pol, Anto

    2015-01-01

    Human amniotic fluid stem cells (AFSC) are an attractive source for cell therapy due to their multilineage differentiation potential and accessibility advantages. However the clinical application of human stem cells largely depends on their capacity to expand in vitro, since there is an extensive donor-to-donor heterogeneity. Reactive oxygen species (ROS) and cellular oxidative stress are involved in many physiological and pathophysiological processes of stem cells, including pluripotency, proliferation, differentiation, and stress resistance. The mode of action of ROS is also dependent on the localization of their target molecules. Thus, the modifications induced by ROS can be separated depending on the cellular compartments they affect. NAD(P)H oxidase family, particularly Nox4, has been known to produce ROS in the nucleus. In the present study we show that Nox4 nuclear expression (nNox4) depends on the donor and it correlates with the expression of transcription factors involved in stemness regulation, such as Oct4, SSEA-4, and Sox2. Moreover nNox4 is linked with the nuclear localization of redox sensitive transcription factors, as Nrf2 and NF-κB, and with the differentiation potential. Taken together, these results suggest that nNox4 regulation may have important effects in stem cell capability through modulation of transcription factors and DNA damage. PMID:26273418

  18. Osteogenic Differentiation of Human Amniotic Fluid Mesenchymal Stem Cells Is Determined by Epigenetic Changes

    PubMed Central

    2016-01-01

    Osteogenic differentiation of human amniotic fluid derived mesenchymal stem cells (AF-MSCs) has been widely studied in vitro and in vivo as a potential tool for regenerative medicine and tissue engineering. While most of the studies analyze changes in transcriptional profile during differentiation to date there is not much information regarding epigenetic changes in AF-MSCs during differentiation. The aim of our study was to evaluate epigenetic changes during osteogenic differentiation of AF-MS cells. Isolated AF-MSCs were characterized morphologically and osteogenic differentiation was confirmed by cell staining and determining expression of alkaline phosphatase and osteopontin by RT-qPCR. Variation in gene expression levels of pluripotency markers and specific microRNAs were also evaluated. Analysis of epigenetic changes revealed that levels of chromatin modifying enzymes such as Polycomb repressive complex 2 (PRC2) proteins (EZH2 and SUZ12), DNMT1, HDAC1, and HDAC2 were reduced after osteogenic differentiation of AF-MSCs. We demonstrated that the level of specific histone markers keeping active state of chromatin (H3K4me3, H3K9Ac, and others) increased and markers of repressed state of chromatin (H3K27me3) decreased. Our results show that osteogenic differentiation of AF-MSCs is conducted by various epigenetic alterations resulting in global chromatin remodeling and provide insights for further epigenetic investigations in human AF-MSCs.

  19. Human amniotic membrane-derived stromal cells (hAMSC) interact depending on breast cancer cell type through secreted molecules.

    PubMed

    Kim, Sun-Hee; Bang, So Hee; Kang, So Yeong; Park, Ki Dae; Eom, Jun Ho; Oh, Il Ung; Yoo, Si Hyung; Kim, Chan-Wha; Baek, Sun Young

    2015-02-01

    Human amniotic membrane-derived stromal cells (hAMSC) are candidates for cell-based therapies. We examined the characteristics of hAMSC including the interaction between hAMSC and breast cancer cells, MCF-7, and MDA-MB-231. Human amniotic membrane-derived stromal cells showed typical MSC properties, including fibroblast-like morphology, surface antigen expression, and mesodermal differentiation. To investigate cell-cell interaction via secreted molecules, we cultured breast cancer cells in hAMSC-conditioned medium (hAMSC-CM) and analyzed their proliferation, migration, and secretome profiles. MCF-7 and MDA-MB-231 cells exposed to hAMSC-CM showed increased proliferation and migration. However, in hAMSC-CM, MCF-7 cells proliferated significantly faster than MDA-MB-231 cells. When cultured in hAMSC-CM, MCF-7 cells migrated faster than MDA-MB-231 cells. Two cell types showed different profiles of secreted factors. MCF-7 cells expressed much amounts of IL-8, GRO, and MCP-1 in hAMSC-CM. Human amniotic membrane-derived stromal cells interact with breast cancer cells through secreted molecules. Factors secreted by hAMSCs promote the proliferation and migration of MCF-7 breast cancer cells. For much safe cell-based therapies using hAMSC, it is necessary to study carefully about interaction between hAMSC and cancer cells.

  20. Quantitative analysis of human parvovirus B19 DNA in maternal and fetal serum, and amniotic fluid during an early stage of pregnancy.

    PubMed

    Ishikawa, Aki; Yoto, Yuko; Asakura, Hirofumi; Tsutsumi, Hiroyuki

    2015-04-01

    Simple and accurate diagnosis of vertical transmission of human parvovirus B19 (B19V) infection remains an important issue in pregnancy. There are few reports on quantitative analysis of B19V in amniotic fluids. Quantitative estimation of B19V DNA in amniotic fluids was comparerd with those in maternal or fetal serum obtained at an early stage of pregnancy with possible mother-to-fetus transmission. All pregnant women contracted B19V infection between 13 to 14 weeks gestation. The B19V DNA amount in 3 maternal serum and amniotic fluid sample pairs collected between 16 to 27 weeks gestation was quantified by a real-time polymerase chain reaction assay. Serum from 2 fetuses was included. The B19V DNA concentrations in maternal sera and amniotic fluids ranged from 10(4) to 10(5) copies/ml and from 10(7) to 10(8) copies/ml, respectively. The B19V DNA in the amniotic fluids concentration coincided with those of each fetal serum. The concentrations in amniotic fluids are 100 to 5,000 times higher than in those of maternal sera, and corresponded to the matching fetal serum. Amniotic fluids may substitute for the fetal sera in terms of quantitative estimation of fetal B19V infection at an early stage of pregnancy.

  1. Human amniotic fluid derived mesenchymal stem cells cause an anti-cancer effect on breast cancer cell line in vitro.

    PubMed

    Ghafarzadeh, M; Eatemadi, A; Fakhravar, Z

    2016-01-01

    Human amniotic fluid stem cells (hAFSCs) have the ability to self-renew, and multipotent differentiation into three germ layer cells. We obtained 5 ml amniotic fluid from ten 16-20 week pregnant women undergoing amniocentesis. hAFSCs were isolated from all samples, co-cultured with T47D breast cancer cell line and characterized using flow cytometry and RT-PCR. After 3, 4 and 5 days, T47D and HSFCs viability were evaluated with MTT assay. After 5 days of co-culture T47D cells viability were decreased. Our findings showed that hAFSCs can release soluble factors in cell culture, causing an efficient anticancer effect. PMID:27262812

  2. Amniotic fluid

    MedlinePlus

    ... baby is born), or gestational diabetes . Too little amniotic fluid is known as oligohydramnios. This condition may occur with late pregnancies, ruptured membranes, placental dysfunction , or fetal abnormalities. Abnormal amounts of ...

  3. Eicosanomic profiling reveals dominance of the epoxygenase pathway in human amniotic fluid at term in spontaneous labor

    PubMed Central

    Maddipati, Krishna Rao; Romero, Roberto; Chaiworapongsa, Tinnakorn; Zhou, Sen-Lin; Xu, Zhonghui; Tarca, Adi L.; Kusanovic, Juan Pedro; Munoz, Hernan; Honn, Kenneth V.

    2014-01-01

    Lipid mediators play an important role in reproductive biology, especially, in parturition. Enhanced biosynthesis of eicosanoids, such as prostaglandin E2 (PGE2) and PGF2α, precedes the onset of labor as a result of increased expression of inducible cyclooxygenase 2 (COX-2) in placental tissues. Metabolism of arachidonic acid results in bioactive lipid mediators beyond prostaglandins that could significantly influence myometrial activity. Therefore, an unbiased lipidomic approach was used to profile the arachidonic acid metabolome of amniotic fluid. In this study, liquid chromatography–mass spectrometry was used for the first time to quantitate these metabolites in human amniotic fluid by comparing patients at midtrimester, at term but not in labor, and at term and in spontaneous labor. In addition to exposing novel aspects of COX pathway metabolism, this lipidomic study revealed a dramatic increase in epoxygenase- and lipoxygenase-pathway-derived lipid mediators in spontaneous labor with remarkable product selectivity. Despite their recognition as anti-inflammatory lipid mediators and regulators of ion channels, little is known about the epoxygenase pathway in labor. Epoxygenase pathway metabolites are established regulators of vascular homeostasis in cardiovascular and renal physiology. Their presence as the dominant lipid mediators in spontaneous labor at term portends a yet undiscovered physiological function in parturition.—Maddipati, K. R., Romero, R., Chaiworapongsa, T., Zhou, S.-L., Xu, Z., Tarca, A. L., Kusanovic, J. P., Munoz, H., Honn, K. V. Eicosanomic profiling reveals dominance of the epoxygenase pathway in human amniotic fluid at term in spontaneous labor. PMID:25059230

  4. Human Amniotic Fluid Mesenchymal Stem Cells from Second- and Third-Trimester Amniocentesis: Differentiation Potential, Molecular Signature, and Proteome Analysis

    PubMed Central

    Savickiene, Jurate; Treigyte, Grazina; Baronaite, Sandra; Valiuliene, Giedre; Kaupinis, Algirdas; Valius, Mindaugas; Arlauskiene, Audrone; Navakauskiene, Ruta

    2015-01-01

    Human amniotic fluid stem cells have become an attractive stem cell source for potential applications in regenerative medicine and tissue engineering. The aim of this study was to characterize amniotic fluid-derived mesenchymal stem cells (AF-MSCs) from second- and third-trimester of gestation. Using two-stage protocol, MSCs were successfully cultured and exhibited typical stem cell morphological, specific cell surface, and pluripotency markers characteristics. AF-MSCs differentiated into adipocytes, osteocytes, chondrocytes, myocytes, and neuronal cells, as determined by morphological changes, cell staining, and RT-qPCR showing the tissue-specific gene presence for differentiated cell lineages. Using SYNAPT G2 High Definition Mass Spectrometry technique approach, we performed for the first time the comparative proteomic analysis between undifferentiated AF-MSCs from late trimester of gestation and differentiated into myogenic, adipogenic, osteogenic, and neurogenic lineages. The analysis of the functional and expression patterns of 250 high abundance proteins selected from more than 1400 demonstrated the similar proteome of cultured and differentiated AF-MSCs but the unique changes in their expression profile during cell differentiation that may help the identification of key markers in differentiated cells. Our results provide evidence that human amniotic fluid of second- and third-trimester contains stem cells with multilineage potential and may be attractive source for clinical applications. PMID:26351462

  5. Osteogenic Differentiation of Human Amniotic Epithelial Cells and Its Application in Alveolar Defect Restoration

    PubMed Central

    Jiawen, Si; Jianjun, Zhang; Jiewen, Dai; Dedong, Yu; Hongbo, Yu; Jun, Shi; Xudong, Wang; Shen, Steve G.F.

    2014-01-01

    The present study investigated the detailed in vitro osteogenic differentiation process and in vivo bone regenerative property of human amniotic epithelial cells (hAECs). The in vitro osteogenic differentiation process of hAECs was evaluated by biochemical staining, real-time polymerase chain reaction, and immunofluorescence. Next, β-tricalcium phosphate (β-TCP) scaffolds alone or loaded with hAECs were implanted into the alveolar defects of rats. Micro-computed tomography evaluation and histologic studies were conducted. Our results validated the in vitro osteogenic capacity of hAECs by upregulation of Runx2, osterix, alkaline phosphatase, collagen I, and osteopontin, with positive biochemical staining for osteoblasts. An epithelial-mesenchymal transformation process might be involved in the osteogenic differentiation of hAECs by increased expression of transforming growth factor-β1. Our data also demonstrated that in vivo implantation of hAECs loaded on β-TCP scaffolds, not only improved bone regeneration by direct participation, but also reduced the early host immune response to the scaffolds. The presented data indicate that hAECs possess proper osteogenic differentiation potential and a modulatory influence on the early tissue remodeling process, making these cells a potential source of progenitor cells for clinical restoration of the alveolar defect. PMID:25368378

  6. Induction of metallothionein and stomatin by interleukin-6 and glucocorticoids in a human amniotic cell line.

    PubMed

    Snyers, L; Content, J

    1994-07-15

    Interleukin 6 (IL-6) is an important mediator of various kinds of inflammatory and immune responses. The human amniotic cell line UAC has an increased number of IL-6 receptors after treatment by glucocorticoids. To find a possible activity of IL-6 on these cells, a cDNA library of IL-6- and dexamethasone-treated cells was screened with cDNA probes from both induced and non-induced cells. Two cDNAs showed a differential hybridization signal. The first one corresponds to metallothionein, a group of small cysteine-rich proteins thought to participate in the metabolism and storage of zinc and to protect cells against oxidative damage. A second cDNA corresponds to the recently cloned cDNA of band 7 integral membrane protein also called stomatin. In hereditary stomatocytosis, absence of this protein in erythrocyte membranes is associated with high Na+ and low K+ intracellular concentrations [Stewart, G. W., Hepworth-Jones, B. E., Keen, J. N., Dash, B. C. J., Argent, A. C. & Casimir, C. M. (1992) Blood 79, 1593-1601]. In UAC cells both metallothionein and stomatin are induced by dexamethasone and IL-6 in a more than additive manner. Western blot analysis shows that stomatin protein is induced in a similar way as its mRNA. IL-6 and dexamethasone induce a state of resistance against hydrogen peroxide toxicity in UAC cells. Metallothionein induction might be partly responsible for this cytoprotection against oxidative stress.

  7. Therapeutic potential of human amniotic membrane-derived mesenchymal stem cells in APP transgenic mice

    PubMed Central

    Jiao, Hongliang; Shi, Ke; Zhang, Weijie; Yang, Liang; Yang, Lu; Guan, Fangxia; Yang, Bo

    2016-01-01

    Growing evidence indicates that the presence of extensive oxidative stress plays an essential role in the initiation and progression of Alzheimer's disease (AD). Amyloid-β (Aβ) aggregation is involved in the elevation of oxidative stress, contributing to mitochondrial dysfunction and lipid peroxidation. In the present study, human placenta amniotic membrane-derived mesenchymal stem cells (hAMMSCs) were intravenously injected into C57BL/6J-APP transgenic mice. hAMMSCs significantly ameliorated spatial learning and memory function, and were associated with a decreased amount of amyloid plaques of the brain. The correlation of oxidative stress with Aβ levels was lower in the hAMMSCs-injected group than in the phosphate-buffered saline (PBS)-injected group, as indicated by the increased level of antioxidative enzymes and the decreased level of lipid peroxidation product. The glutathione (GSH) level and ratio of GSH to glutathione disulfide were higher in the hAMMSC group than in the PBS group. The superoxide dismutase activity and malonaldehyde level were improved significantly as the level of Aβ decreased, but there was no such trend in the PBS group. As a result, our findings represent evidence that hAMMSC treatment might improve the pathology of AD and memory function through the regulation of oxidative stress. PMID:27588134

  8. An ultrastructural analysis of isolated basement membranes in the acellular renal cortex: a comparison study of human and laboratory animals.

    PubMed

    Carlson, E C; Kenney, M C

    1982-02-01

    Freshly harvested kidneys from New Zealand white rabbits, Sprague-Dawley white rats, rhesus monkeys, and transplant-quality human kidneys were used in this study. Minced renal cortical tissue blocks (less than 2 mm3) were treated with 1 mM EDTA, 3% Triton X-100, 0.025% DNAse, and 4% sodium deoxycholate in an effort to remove all cellular elements and leave the extracellular matrix (ECM) intact. These preparations showed remarkable structural preservation and all components of the ECM, including basement membranes (BMs), maintained their in vivo histoarchitectural relationships. By light microscopy, at least four major BM types were recognizable, including Bowman's capsular BM (BCBM), tubular BM (TBM), glomerular BM (GBM), and peritubular capillary BM (PTCBM). Scanning electron microscopy demonstrated that, despite the lack of supporting interstitium, GBMs in human, monkey, and rat (and rabbit to a lesser degree) exhibit intrinsic structural rigidity such that their convoluted spheroidal shapes are maintained following cell removal. Transmission electron microscopy showed that major BM types are morphologically heterogeneous and vary markedly within and between species. Randomized measurements showed that isolated BM thicknesses (lamina densa only) compared favorably with those reported in cellular preparations. Mean thicknesses of GBMs were within normal ranges in all species with or without power transformations to reduce right-sided skew of distribution curves. In all species, thickness of BCBM greater than TBM greater than GBM greater than PTCBM. The striking morphologic heterogeneity of major BM types demonstrated in the acellular renal cortex is not surprising in view of recent biochemical analyses that show that BMs derived from different sources are compositionally disparate. We conclude that BMs should be evaluated and characterized individually and that morphologic definition of isolated BMs necessary prior to further analysis. PMID:7062344

  9. Lectin-based analysis of fucose and sialic acid expressions on human amniotic IgA during normal pregnancy.

    PubMed

    Orczyk-Pawiłowicz, Magdalena; Augustyniak, Daria; Hirnle, Lidia; Kątnik-Prastowska, Iwona

    2013-08-01

    The sugar moiety of IgA is known to provide a link between the innate and adaptive immune systems. Terminally located glycotopes on IgA are potential ligands engaged in the interactions which may modulate the biological activities of IgA. In the present work the expressions of Maackia amurensis (MAA), Sambucus nigra (SNA), Lens culinaris (LCA), Tetragonolobus purpureus (LTA), and Ulex europaeus (UEA) reactive glycotopes on maternal plasma and amniotic IgA were evaluated in relation to the progression of a normal human pregnancy, from the 2nd trimester, throughout the 3rd trimester, perinatal period, post-date pregnancy and delivery, by lectin-IgA-ELISA, using specific biotinylated lectins. The amniotic and maternal plasma IgA concentrations and a degree of SNA and LCA reactivity of maternal plasma IgA were almost unaltered during the normal pregnancy. The amniotic IgA from the 2nd trimester was decorated by MAA-, SNA-reactive and LCA-, LTA-, and UEA-reactive glycotopes. At the turn of the 2nd and 3rd trimesters the expression of MAA-, SNA-, LTA-, and UEA-reactive glycotopes, except for LCA-reactive, increased and remained almost at unaltered levels throughout the perinatal period and delivery. However, in the post-date pregnancy the expression of LCA-, LTA-, and UEA-reactive and SNA-reactive glycotopes were significantly higher. The unique fucosylated and sialylated glycovariants of amniotic IgA associated with the progression of the normal pregnancy may illustrate a general importance of carbohydrate-lectin receptor interactions in the control and modulation of biological events to ensuring homeostasis during pregnancy, protection and well-being of fetus.

  10. In vivo effects of human adipose-derived stem cells reseeding on acellular bovine pericardium in nude mice

    PubMed Central

    Dai, Miao; Xu, Peirong; Hou, Min; Teng, Yincheng; Feng, Jie

    2015-01-01

    Tissue-engineered biologic products may be a viable option in the reconstruction of pelvic organ prolapse (POP). This study was based on the hypothesis that human adipose-derived stem cells (hASCs) are viable in acellular bovine pericardium (ABP), when reseeded by two different techniques, and thus, aid in the reconstruction. To investigate the reseeding of hASCs on ABP grafts by using non-invasive bioluminescence imaging (BLI), and to identify the effective hASCs–scaffold combinations that enabled regeneration. Thirty female athymic nude mice were randomly divided into three groups: In the VIVO group, ABPs were implanted in the subcutaneous pockets and enhanced green fluorescent protein luciferase (eGFP·Luc)-hASCs (1 × 106 cells/50 µL) were injected on the ABP at the same time. In the VITRO group, the mice were implanted with grafts that ABP were co-cultured with eGFP·Luc-hASCs in vitro. The BLANK group mice were implanted with ABP only. The eGFP·Luc-hASCs reseeded on ABP were analyzed by BLI, histology, and immunohistochemistry. The eGFP·Luc-hASCs reseeded on ABP could be visualized at 12 weeks in vivo. Histology revealed that the VIVO group displayed the highest cell ingrowths, small vessels, and percent of collagen content per unit area. Desmin and α-smooth muscle actin were positive at the same site in the VIVO group cells. However, few smooth muscles were observed in the VITRO and BLANK groups. These results suggest that hASCs reseeded on ABP in vivo during surgery may further enhance the properties of ABP and may promote regeneration at the recipient site, resulting in a promising treatment option for POP. PMID:26253192

  11. In vitro differentiation of human amniotic epithelial cells into insulin-producing 3D spheroids.

    PubMed

    Okere, Bernard; Alviano, Francesco; Costa, Roberta; Quaglino, Daniela; Ricci, Francesca; Dominici, Massimo; Paolucci, Paolo; Bonsi, Laura; Iughetti, Lorenzo

    2015-09-01

    Regenerative medicine and stem cell therapy may represent the solution for the treatment of non-curable human diseases such as type 1 diabetes. In this context of growing demand for functional and safe stem cells, human amniotic epithelial cells (hAECs) from term placenta have attracted increasing interest for their wide availability, stem cell properties, and differentiation plasticity, which make them a promising tool for stem cell-based therapeutic applications. We initially assayed the stemness characteristics of hAECs in serum-free conditions. Subsequently we developed a culture procedure on extracellular matrix for the formation of three-dimensional (3D) spheroids. Finally, we tested the immunomodulation and differentiation potential of hAEC spheroids: the presence of pancreatic endocrine hormones was revealed with transmission electron microscopy and immunofluorescence analyses; the release of C-peptide in hyperglycemic conditions was assayed with ELISA. The serum-free culture conditions we applied proved to maintain the basic stemness characteristics of hAECs. We also demonstrated that 3D spheroids formed by hAECs in extracellular matrix can be induced to differentiate into insulin-producing cells. Finally, we proved that control and induced cells equally inhibit the proliferation of activated mononuclear cells. The results of this study highlight the properties of amnion derived epithelial cells as promising and abundant source for cell-based therapies. In particular we are the first group to show the in vitro pancreatic induction of hAECs cultured on extracellular matrix in a 3D fashion. We accordingly propose the outcomes of this study as a novel contribution to the development of future cell replacement therapies involving placenta-derived cells.

  12. Human amniotic epithelial cells are reprogrammed more efficiently by induced pluripotency than adult fibroblasts.

    PubMed

    Easley, Charles A; Miki, Toshio; Castro, Carlos A; Ozolek, John A; Minervini, Crescenzio F; Ben-Yehudah, Ahmi; Schatten, Gerald P

    2012-06-01

    Cellular reprogramming from adult somatic cells into an embryonic cell-like state, termed induced pluripotency, has been achieved in several cell types. However, the ability to reprogram human amniotic epithelial cells (hAECs), an abundant cell source derived from discarded placental tissue, has only recently been investigated. Here we show that not only are hAECs easily reprogrammed into induced pluripotent stem cells (AE-iPSCs), but hAECs reprogram faster and more efficiently than adult and neonatal somatic dermal fibroblasts. Furthermore, AE-iPSCs express higher levels of NANOG and OCT4 compared to human foreskin fibroblast iPSCs (HFF1-iPSCs) and express decreased levels of genes associated with differentiation, including NEUROD1 and SOX17, markers of neuronal differentiation. To elucidate the mechanism behind the higher reprogramming efficiency of hAECs, we analyzed global DNA methylation, global histone acetylation, and the mitochondrial DNA A3243G point mutation. Whereas hAECs show no differences in global histone acetylation or mitochondrial point mutation accumulation compared to adult and neonatal dermal fibroblasts, hAECs demonstrate a decreased global DNA methylation compared to dermal fibroblasts. Likewise, quantitative gene expression analyses show that hAECs endogenously express OCT4, SOX2, KLF4, and c-MYC, all four factors used in cellular reprogramming. Thus, hAECs represent an ideal cell type for testing novel approaches for generating clinically viable iPSCs and offer significant advantages over postnatal cells that more likely may be contaminated by environmental exposures and infectious agents. PMID:22686477

  13. Human Lung Cancer Cells Grown on Acellular Rat Lung Matrix Create Perfusable Tumor Nodules

    PubMed Central

    Mishra, Dhruva K.; Thrall, Michael J.; Baird, Brandi N.; Ott, Harald C.; Blackmon, Shanda H.; Kurie, Jonathan M.; Kim, Min P.

    2015-01-01

    Background Extracellular matrix allows lung cancer to form its shape and grow. Recent studies on organ reengineering for orthotopic transplantation have provided a new avenue for isolating purified native matrix to use for growing cells. Whether human lung cancer cells grown in a decellularized rat lung matrix would create perfusable human lung cancer nodules was tested. Methods Rat lungs were harvested and native cells were removed using sodium dodecyl sulfate and Triton X-100 in a decellularization chamber to create a decellularized rat lung matrix. Human A549, H460, or H1299 lung cancer cells were placed into the decellularized rat lung matrix and grown in a customized bioreactor with perfusion of oxygenated media for 7 to 14 days. Results Decellularized rat lung matrix showed preservation of matrix architecture devoid of all rat cells. All three human lung cancer cell lines grown in the bioreactor developed tumor nodules with intact vasculature. Moreover, the lung cancer cells developed a pattern of growth similar to the original human lung cancer. Conclusions Overall, this study shows that human lung cancer cells form perfusable tumor nodules in a customized bioreactor on a decellularized rat lung matrix created by a customized decellularization chamber. The lung cancer cells grown in the matrix had features similar to the original human lung cancer. This ex vivo model can be used potentially to gain a deeper understanding of the biologic processes involved in human lung cancer. PMID:22385822

  14. Human Acellular Dermis versus Submuscular Tissue Expander Breast Reconstruction: A Multivariate Analysis of Short-Term Complications

    PubMed Central

    Davila, Armando A.; Seth, Akhil K.; Wang, Edward; Hanwright, Philip; Bilimoria, Karl; Fine, Neil

    2013-01-01

    Background Acellular dermal matrix (ADM) allografts and their putative benefits have been increasingly described in prosthesis based breast reconstruction. There have been a myriad of analyses outlining ADM complication profiles, but few large-scale, multi-institutional studies exploring these outcomes. In this study, complication rates of acellular dermis-assisted tissue expander breast reconstruction were compared with traditional submuscular methods by evaluation of the American College of Surgeon's National Surgical Quality Improvement Program (NSQIP) registry. Methods Patients who underwent immediate tissue expander breast reconstruction from 2006-2010 were identified using surgical procedure codes. Two hundred forty tracked variables from over 250 participating sites were extracted for patients undergoing acellular dermis-assisted versus submuscular tissue expander reconstruction. Thirty-day postoperative outcomes and captured risk factors for complications were compared between the two groups. Results A total of 9,159 patients underwent tissue expander breast reconstruction; 1,717 using acellular dermis and 7,442 with submuscular expander placement. Total complications and reconstruction related complications were similar in both cohorts (5.5% vs. 5.3%, P=0.68 and 4.7% vs. 4.3%, P=0.39, respectively). Multivariate logistic regression revealed body mass index and smoking as independent risk factors for reconstructive complications in both cohorts (P<0.01). Conclusions The NSQIP database provides large-scale, multi-institutional, independent outcomes for acellular dermis and submuscular breast reconstruction. Both thirty-day complication profiles and risk factors for post operative morbidity are similar between these two reconstructive approaches. PMID:23362476

  15. Orthopedic applications of acellular human dermal allograft for shoulder and elbow surgery.

    PubMed

    Acevedo, Daniel C; Shore, Brett; Mirzayan, Raffy

    2015-07-01

    Shoulder and elbow tendon injuries are some of the most challenging problems to treat surgically. Tendon repairs in the upper extremity can be complicated by poor tendon quality and, often times, poor healing. Extracellular matrices, such as human dermal allografts, have been used to augment tendon repairs in shoulder and elbow surgery. The indications and surgical techniques regarding the use of human dermal allograft continue to evolve. This article reviews the basic science, rationale for use, and surgical applications of human dermal allograft in shoulder and elbow tendon injuries.

  16. Human Acellular Dermal Matrix Paired With Silver-zinc Coupled Electroceutical Dressing Results in Rapid Healing of Complicated Diabetic Wounds of Mixed Etiology: A Novel Case Series.

    PubMed

    Cole, Windy

    2016-07-01

    Patients with diabetes are well known for having difficult-to-close wounds. When additional factors are added, such as gouty tophi or tumors, the difficulty is compounded and conventional care often fails to heal the wound. In this case series, an innovative wound modality that combined a human acellular dermal matrix with a silver-zinc coupled electroceutical wound dressing was used in 3 particularly difficult and complex cases. In all 3 cases, this alternative treatment provided full healing within 6 weeks in wounds that conventional care had been unable to close in up to 2 years. PMID:27428719

  17. An epidermal stem cells niche microenvironment created by engineered human amniotic membrane.

    PubMed

    Ji, Shi-zhao; Xiao, Shi-chu; Luo, Peng-fei; Huang, Guo-feng; Wang, Guang-yi; Zhu, Shi-hui; Wu, Min-juan; Xia, Zhao-fan

    2011-11-01

    How to amplify epidermal stem cells (ESCs) rapidly is a challenging crux in skin tissue engineering research. The present study describes the preparation of 3D micronized (300-600 μm) amniotic membrane (mAM) by means of repeated freeze-thawing cycles to deplete cell components and homogenized with a macrohomogenizer in liquid nitrogen. This newly prepared mAM not only possessed the characteristics of a microcarrier but completely retained the basement membrane structure and abundant active substances such as NGF, HGF, KGF, bFGF, TGF-β1 and EGF in the AM matrix. The result showed that mAM combined with rotary cell culture system (RCCS) was able to amplify ESCs quickly. The relative cell viability at day 7 and 14 was significantly higher than that of the conventional 2D plate culture (326 ± 28% and 535 ± 47% versus 232 ± 21% and 307 ± 32%, P < 0.05). In addition, the new method was able to prevent cell differentiation effectively and retain the characteristics of stem cells. When mAM loaded with ESCs (ESC-mAM) was further transplanted to full-thickness skin defects in nude mice, ESCs survived well and formed a new epidermis. Four weeks after transplantation, papilla-like structures were observed, and collagen fibers were well and regularly arranged in the newly formed dermal layer. In conclusion, the mAM as a novel natural microcarrier possesses an intact basement membrane structure and bioactivities. It not only provides the microenvironment similar to the stem cell niche within the human body favorable for ex vivo culture and amplification of ESCs but can be used as the dermal scaffold in constructing a skin substitute containing ESCs for the repair of full-thickness skin defects.

  18. The human amniotic fluid stem cell secretome effectively counteracts doxorubicin-induced cardiotoxicity

    PubMed Central

    Lazzarini, Edoardo; Balbi, Carolina; Altieri, Paola; Pfeffer, Ulrich; Gambini, Elisa; Canepa, Marco; Varesio, Luigi; Bosco, Maria Carla; Coviello, Domenico; Pompilio, Giulio; Brunelli, Claudio; Cancedda, Ranieri; Ameri, Pietro; Bollini, Sveva

    2016-01-01

    The anthracycline doxorubicin (Dox) is widely used in oncology, but it may cause a cardiomyopathy with bleak prognosis that cannot be effectively prevented. The secretome of human amniotic fluid-derived stem cells (hAFS) has previously been demonstrated to significantly reduce ischemic cardiac damage. Here it is shown that, following hypoxic preconditioning, hAFS conditioned medium (hAFS-CM) antagonizes senescence and apoptosis of cardiomyocytes and cardiac progenitor cells, two major features of Dox cardiotoxicity. Mechanistic studies with mouse neonatal ventricular cardiomyocytes (mNVCM) reveal that hAFS-CM inhibition of Dox-elicited senescence and apoptosis is associated with decreased DNA damage, nuclear translocation of NF-kB, and upregulation of the NF-kB controlled genes, Il6 and Cxcl1, promoting mNVCM survival. Furthermore, hAFS-CM induces expression of the efflux transporter, Abcb1b, and Dox extrusion from mNVCM. The PI3K/Akt signaling cascade, upstream of NF-kB, is potently activated by hAFS-CM and pre-treatment with a PI3K inhibitor abrogates NF-kB accumulation into the nucleus, modulation of Il6, Cxcl1 and Abcb1b, and prevention of Dox-initiated senescence and apoptosis in response to hAFS-CM. These results support the concept that hAFS are a valuable source of cardioprotective factors and lay the foundations for the development of a stem cell-based paracrine treatment of chemotherapy-related cardiotoxicity. PMID:27444332

  19. An epidermal stem cells niche microenvironment created by engineered human amniotic membrane.

    PubMed

    Ji, Shi-zhao; Xiao, Shi-chu; Luo, Peng-fei; Huang, Guo-feng; Wang, Guang-yi; Zhu, Shi-hui; Wu, Min-juan; Xia, Zhao-fan

    2011-11-01

    How to amplify epidermal stem cells (ESCs) rapidly is a challenging crux in skin tissue engineering research. The present study describes the preparation of 3D micronized (300-600 μm) amniotic membrane (mAM) by means of repeated freeze-thawing cycles to deplete cell components and homogenized with a macrohomogenizer in liquid nitrogen. This newly prepared mAM not only possessed the characteristics of a microcarrier but completely retained the basement membrane structure and abundant active substances such as NGF, HGF, KGF, bFGF, TGF-β1 and EGF in the AM matrix. The result showed that mAM combined with rotary cell culture system (RCCS) was able to amplify ESCs quickly. The relative cell viability at day 7 and 14 was significantly higher than that of the conventional 2D plate culture (326 ± 28% and 535 ± 47% versus 232 ± 21% and 307 ± 32%, P < 0.05). In addition, the new method was able to prevent cell differentiation effectively and retain the characteristics of stem cells. When mAM loaded with ESCs (ESC-mAM) was further transplanted to full-thickness skin defects in nude mice, ESCs survived well and formed a new epidermis. Four weeks after transplantation, papilla-like structures were observed, and collagen fibers were well and regularly arranged in the newly formed dermal layer. In conclusion, the mAM as a novel natural microcarrier possesses an intact basement membrane structure and bioactivities. It not only provides the microenvironment similar to the stem cell niche within the human body favorable for ex vivo culture and amplification of ESCs but can be used as the dermal scaffold in constructing a skin substitute containing ESCs for the repair of full-thickness skin defects. PMID:21803416

  20. The human amniotic fluid stem cell secretome effectively counteracts doxorubicin-induced cardiotoxicity.

    PubMed

    Lazzarini, Edoardo; Balbi, Carolina; Altieri, Paola; Pfeffer, Ulrich; Gambini, Elisa; Canepa, Marco; Varesio, Luigi; Bosco, Maria Carla; Coviello, Domenico; Pompilio, Giulio; Brunelli, Claudio; Cancedda, Ranieri; Ameri, Pietro; Bollini, Sveva

    2016-01-01

    The anthracycline doxorubicin (Dox) is widely used in oncology, but it may cause a cardiomyopathy with bleak prognosis that cannot be effectively prevented. The secretome of human amniotic fluid-derived stem cells (hAFS) has previously been demonstrated to significantly reduce ischemic cardiac damage. Here it is shown that, following hypoxic preconditioning, hAFS conditioned medium (hAFS-CM) antagonizes senescence and apoptosis of cardiomyocytes and cardiac progenitor cells, two major features of Dox cardiotoxicity. Mechanistic studies with mouse neonatal ventricular cardiomyocytes (mNVCM) reveal that hAFS-CM inhibition of Dox-elicited senescence and apoptosis is associated with decreased DNA damage, nuclear translocation of NF-kB, and upregulation of the NF-kB controlled genes, Il6 and Cxcl1, promoting mNVCM survival. Furthermore, hAFS-CM induces expression of the efflux transporter, Abcb1b, and Dox extrusion from mNVCM. The PI3K/Akt signaling cascade, upstream of NF-kB, is potently activated by hAFS-CM and pre-treatment with a PI3K inhibitor abrogates NF-kB accumulation into the nucleus, modulation of Il6, Cxcl1 and Abcb1b, and prevention of Dox-initiated senescence and apoptosis in response to hAFS-CM. These results support the concept that hAFS are a valuable source of cardioprotective factors and lay the foundations for the development of a stem cell-based paracrine treatment of chemotherapy-related cardiotoxicity. PMID:27444332

  1. Acellular human heart matrix: A critical step toward whole heart grafts.

    PubMed

    Sánchez, Pedro L; Fernández-Santos, M Eugenia; Costanza, Salvatore; Climent, Andreu M; Moscoso, Isabel; Gonzalez-Nicolas, M Angeles; Sanz-Ruiz, Ricardo; Rodríguez, Hugo; Kren, Stefan M; Garrido, Gregorio; Escalante, Jose L; Bermejo, Javier; Elizaga, Jaime; Menarguez, Javier; Yotti, Raquel; Pérez del Villar, Candelas; Espinosa, M Angeles; Guillem, María S; Willerson, James T; Bernad, Antonio; Matesanz, Rafael; Taylor, Doris A; Fernández-Avilés, Francisco

    2015-08-01

    The best definitive treatment option for end-stage heart failure currently is transplantation, which is limited by donor availability and immunorejection. Generating an autologous bioartificial heart could overcome these limitations. Here, we have decellularized a human heart, preserving its 3-dimensional architecture and vascularity, and recellularized the decellularized extracellular matrix (dECM). We decellularized 39 human hearts with sodium-dodecyl-sulfate for 4-8 days. Cell removal and architectural integrity were determined anatomically, functionally, and histologically. To assess cytocompatibility, we cultured human cardiac-progenitor cells (hCPC), bone-marrow mesenchymal cells (hMSCs), human endothelial cells (HUVECs), and H9c1 and HL-1 cardiomyocytes in vitro on dECM ventricles up to 21 days. Cell survival, gene expression, organization and/or electrical coupling were analyzed and compared to conventional 2-dimensional cultures. Decellularization removed cells but preserved the 3-dimensional cardiac macro and microstructure and the native vascular network in a perfusable state. Cell survival was observed on dECM for 21 days. hCPCs and hMSCs expressed cardiocyte genes but did not adopt cardiocyte morphology or organization; HUVECs formed a lining of endocardium and vasculature; differentiated cardiomyocytes organized into nascent muscle bundles and displayed mature calcium dynamics and electrical coupling in recellularized dECM. In summary, decellularization of human hearts provides a biocompatible scaffold that retains 3-dimensional architecture and vascularity and that can be recellularized with parenchymal and vascular cells. dECM promotes cardiocyte gene expression in stem cells and organizes existing cardiomyocytes into nascent muscle showing electrical coupling. These findings represent a first step toward manufacturing human heart grafts or matrix components for treating cardiovascular disease.

  2. The use of human amniotic fluid mesenchymal stem cells as the feeder layer to establish human embryonic stem cell lines.

    PubMed

    Soong, Yung-Kwei; Huang, Shang-Yu; Yeh, Chiu-Hsiang; Wang, Tzu-Hao; Chang, Kuo-Hsuan; Cheng, Po-Jen; Shaw, S W Steven

    2015-12-01

    Human embryonic stem cells (hESCs) are pluripotent cells that have the potential to differentiate into the three germ layers and possibly all tissues of the human body. To fulfil the clinical potentials for cell-based therapy, banks of hESC lines that express different combinations of the major histocompatibility genes should be established, preferably without exposing such cells to animal cells and proteins. In this study, we tested human amniotic fluid mesenchymal stem cells (AFMSCs) as feeder cells to support the growth of hESCs. Our results indicated that mitomycin-treated AFMSCs were able to support the newly established hESC lines CGLK-1 and CGLK-2. The hESC colonies cultured on AFMSCs expressed alkaline phosphatase (ALK-P), SSEA-4, TRA-1-60, TRA-1-81, Oct-4, Nanog and Sox-2, which are markers for undifferentiated hESCs. Chromosomal analyses of both hESC lines, CGLK-1 and CGLK-2, which were cultured on AFMSC feeders for 22 and 14 passages, respectively, were confirmed to be normal karyotypes (46, XX). The ability of AFMSCs as feeder cells to maintain the undifferentiated growth and pluripotency of hESCs was confirmed by in vivo formation of teratomas derived on AFMSC hESCs in severe combined immune-compromised mice. The use of AFMSCs for feeder cells to culture hESCs has several advantages, in that AFMSCs are not tumourigenic and can be expanded extensively with a short doubling time.

  3. Cryopreserved Human Amniotic Membrane and A Bioinspired Underwater Adhesive To Seal And Promote Healing Of Iatrogenic Fetal Membrane Defect Sites

    PubMed Central

    Papanna, Ramesha; Mann, Lovepreet K; Tseng, Scheffer C.G.; Stewart, Russell J; Kaur, Sarbjit S; Swindle, M Michael; Kyriakides, Themis R; Tatevian, Nina; Moise, Kenneth J

    2015-01-01

    Introduction We investigated the ability of cryopreserved human amniotic membrane (hAM) scaffold sealed with an underwater adhesive, bio-inspired by marine sandcastle worms to promote healing of iatrogenic fetal membrane defects in a pregnant swine model. Methods Twelve Yucatan miniature pigs underwent laparotomy under general anesthesia at 70 days gestation (term = 114 days). The gestational sacs were assigned to uninstrumented (n=24) and instrumented with 12 Fr trocar, which was further randomized into four different arms-no hAM patch, (n=22), hAM patch secured with suture (n=16), hAM patch with no suture (n=14), and hAM patch secured with adhesive (n=9). The animals were euthanized 20 days after the procedure. Gross and histological examination of the entry site was performed for fetal membrane healing. Results There were no differences in fetal survival, amniotic fluid levels, or dye-leakage from the amniotic cavity between the groups. The fetal membranes spontaneously healed in instrumented sacs without hAM patches. In sacs with hAM patches secured with sutures, the patch was incorporated into the swine fetal membranes. In sacs with hAM patches without sutures, 100% of the patches were displaced from the defect site, whereas in sacs with hAM patches secured with adhesive 55% of the patches remained in place and showed complete healing (p=0.04). Discussion In contrast to humans, swine fetal membranes heal spontaneously after an iatrogenic injury and thus not an adequate model. hAM patches became incorporated into the defect site by cellular ingrowth from the fetal membranes. The bioinspired adhesive adhered the hAM patches within the defect site. PMID:26059341

  4. Markers of Pluripotency in Human Amniotic Epithelial Cells and Their Differentiation to Progenitor of Cortical Neurons.

    PubMed

    García-Castro, Irma Lydia; García-López, Guadalupe; Ávila-González, Daniela; Flores-Herrera, Héctor; Molina-Hernández, Anayansi; Portillo, Wendy; Ramón-Gallegos, Eva; Díaz, Néstor Fabián

    2015-01-01

    Human pluripotent stem cells (hPSC) have promise for regenerative medicine due to their auto-renovation and differentiation capacities. Nevertheless, there are several ethical and methodological issues about these cells that have not been resolved. Human amniotic epithelial cells (hAEC) have been proposed as source of pluripotent stem cells. Several groups have studied hAEC but have reported inconsistencies about their pluripotency properties. The aim of the present study was the in vitro characterization of hAEC collected from a Mexican population in order to identify transcription factors involved in the pluripotency circuitry and to determine their epigenetic state. Finally, we evaluated if these cells differentiate to cortical progenitors. We analyzed qualitatively and quantitatively the expression of the transcription factors of pluripotency (OCT4, SOX2, NANOG, KLF4 and REX1) by RT-PCR and RT-qPCR in hAEC. Also, we determined the presence of OCT4, SOX2, NANOG, SSEA3, SSEA4, TRA-1-60, E-cadherin, KLF4, TFE3 as well as the proliferation and epigenetic state by immunocytochemistry of the cells. Finally, hAEC were differentiated towards cortical progenitors using a protocol of two stages. Here we show that hAEC, obtained from a Mexican population and cultured in vitro (P0-P3), maintained the expression of several markers strongly involved in pluripotency maintenance (OCT4, SOX2, NANOG, TFE3, KLF4, SSEA3, SSEA4, TRA-1-60 and E-cadherin). Finally, when hAEC were treated with growth factors and small molecules, they expressed markers characteristic of cortical progenitors (TBR2, OTX2, NeuN and β-III-tubulin). Our results demonstrated that hAEC express naïve pluripotent markers (KLF4, REX1 and TFE3) as well as the cortical neuron phenotype after differentiation. This highlights the need for further investigation of hAEC as a possible source of hPSC.

  5. Markers of Pluripotency in Human Amniotic Epithelial Cells and Their Differentiation to Progenitor of Cortical Neurons

    PubMed Central

    García-Castro, Irma Lydia; García-López, Guadalupe; Ávila-González, Daniela; Flores-Herrera, Héctor; Molina-Hernández, Anayansi; Portillo, Wendy; Ramón-Gallegos, Eva; Díaz, Néstor Fabián

    2015-01-01

    Human pluripotent stem cells (hPSC) have promise for regenerative medicine due to their auto-renovation and differentiation capacities. Nevertheless, there are several ethical and methodological issues about these cells that have not been resolved. Human amniotic epithelial cells (hAEC) have been proposed as source of pluripotent stem cells. Several groups have studied hAEC but have reported inconsistencies about their pluripotency properties. The aim of the present study was the in vitro characterization of hAEC collected from a Mexican population in order to identify transcription factors involved in the pluripotency circuitry and to determine their epigenetic state. Finally, we evaluated if these cells differentiate to cortical progenitors. We analyzed qualitatively and quantitatively the expression of the transcription factors of pluripotency (OCT4, SOX2, NANOG, KLF4 and REX1) by RT-PCR and RT-qPCR in hAEC. Also, we determined the presence of OCT4, SOX2, NANOG, SSEA3, SSEA4, TRA-1-60, E-cadherin, KLF4, TFE3 as well as the proliferation and epigenetic state by immunocytochemistry of the cells. Finally, hAEC were differentiated towards cortical progenitors using a protocol of two stages. Here we show that hAEC, obtained from a Mexican population and cultured in vitro (P0-P3), maintained the expression of several markers strongly involved in pluripotency maintenance (OCT4, SOX2, NANOG, TFE3, KLF4, SSEA3, SSEA4, TRA-1-60 and E-cadherin). Finally, when hAEC were treated with growth factors and small molecules, they expressed markers characteristic of cortical progenitors (TBR2, OTX2, NeuN and β-III-tubulin). Our results demonstrated that hAEC express naïve pluripotent markers (KLF4, REX1 and TFE3) as well as the cortical neuron phenotype after differentiation. This highlights the need for further investigation of hAEC as a possible source of hPSC. PMID:26720151

  6. Socket Preservation Therapy with Acellular Dermal Matrix and Mineralized Bone Allograft After Tooth Extraction in Humans: A Clinical and Histomorphometric Study.

    PubMed

    Fernandes, Patricia Garani; Muglia, Valdir Antonio; Reino, Danilo Maeda; Maia, Luciana Prado; de Moraes Grisi, Marcio Fernando; de Souza, Sergio Luís; Taba, Mario; Palioto, Daniela Bazan; de Almeida, Adriana G; Novaes, Arthur Belém

    2016-01-01

    The aim of this study was to analyze through clinical and histomorphometric parameters the use of acellular dermal matrix (ADM) with or without mineralized bone allograft (AB) on bone formation in human alveoli after a 6- to 8-month healing period. A total of 19 patients in need of extraction of the maxillary anterior teeth were selected and randomly assigned to the test group (ADM plus AB) or to the control group (ADM only). Clinical and histomorphometric measurements and histologic analysis were recorded 6 to 8 months after ridge preservation procedures. Clinical parameters and amount of mineralized and nonmineralized tissue were measured and analyzed. In the clinical measurements, the test group showed reduced bone loss in the buccopalatal dimension after 6 to 8 months (intragroup analysis P < .01). Histologic findings showed higher percentages of mineralized tissue and lower percentages of nonmineralized tissue in the test group when compared with the control group (P < .05). In this randomized controlled clinical and histomorphometric study in humans, acellular dermal matrix in association with mineralized bone allograft reduced alveolar bone loss in the anterior maxillae both in height and width after a follow-up period of 6 to 8 months. PMID:26901306

  7. Human amniotic epithelial cell niche enhances the functional properties of human corneal endothelial cells via inhibiting P53-survivin-mitochondria axis.

    PubMed

    Sha, Xiangyin; Liu, Zhiping; Song, Li; Wang, Zhonghao; Liang, Xuanwei

    2013-11-01

    The particular microenvironment or niche plays an important role in determining the fate of stem cells and adult cells. The objective of this study was to explore the potential role of the niche of human amniotic epithelial cells in enhancing the functional properties of human corneal endothelial cells (HCECs). The HCECs were cultured in different media, including corneal endothelium medium (CEM), 20% human amniotic epithelial cell culture medium (20% HAEC-Me), and 20% human amniotic epithelial cell-conditioned medium (20% HAEC-CM). We observed that the HCECs cultured in the 20% HAEC-CM had an increased proliferative capacity, higher colony-forming efficiency (CFE), fewer apoptotic cells, and similar cell-junction formation capabilities and pump functionality compared with primary HCECs. Compared with CEM and 20% HAEC-Me, the 20% HAEC-CM system enhanced the functional properties of HCECs by reducing the generation of reactive oxygen species (ROS), maintaining the membrane potential (Δψm) at higher levels, reducing the expression of the p53 protein, and increasing the level of survivin protein expression. This study may shed light on the expansion of HCECs and the clinical applications of these cells in regenerative medicine, especially in corneal tissue engineering.

  8. Characterization of mesenchymal stem cell subpopulations from human amniotic membrane with dissimilar osteoblastic potential.

    PubMed

    Leyva-Leyva, Margarita; Barrera, Lourdes; López-Camarillo, César; Arriaga-Pizano, Lourdes; Orozco-Hoyuela, Gabriel; Carrillo-Casas, Erika M; Calderón-Pérez, Jaime; López-Díaz, Annia; Hernandez-Aguilar, Felipe; González-Ramírez, Ricardo; Kawa, Simón; Chimal-Monroy, Jesús; Fuentes-Mera, Lizeth

    2013-04-15

    Human fetal mesenchymal stem cells can be isolated from the amniotic membrane (AM-hMSCs) by enzymatic digestion. The biological properties of this cell population have been characterized; however, few studies have focused on the presence of stem cell subpopulations and their differentiation potential. The aim of the present study was to isolate homogeneous AM-hMSC subpopulations based on the coexpression of surface markers. In addition, we aimed to characterize stem cell subpopulations through the detection of typical stem cell markers and its differentiation potential. In this study, fluorescence-activated cell sorting (FACS) was used to positively select for the surface markers CD44, CD73, and CD105. Two subpopulations were isolated: CD44+ / CD73+ / CD105+ (CD105+), and CD44+ / CD73+ / CD105- (CD105-). To characterize the cell subpopulations, the expression of pluripotency-associated markers was analyzed by reverse transcriptase-polymerase chain reaction and immunofluorescence. Our results showed positive expression of SOX2, SOX3, PAX6, OCT3/4, and NANOG in the CD105+ and CD105(-) cell subpopulations. In contrast, we did not detect expression of SSEA4 or FOXD3 in either subpopulation. Immunophenotypes, such as mesenchymal and hematopoietic markers, were studied by FACS analyses. Our data revealed the expression of the CD49a, CD49d, CD29, integrin α9β1, CD44, CD73, and CD105 antigens in both subpopulations. In contrast, CD90, CD45, CD34, CD14, and HLA-DR expression was not detected. The ability of both subpopulations to differentiate into osteoblasts, adipocytes, and chondrocytes was evidenced using Alizarin red, Oil-Red, and Alcian blue staining, respectively. Furthermore, neuronal differentiation was demonstrated by the expression of GFAP and NEURO-D. Interestingly, we observed a dissimilar osteoblastic differentiation potential between the subpopulations. CD105- cells showed stronger expression of secreted protein acidic and rich in cysteine (SPARC) and

  9. Multiple systemic transplantations of human amniotic mesenchymal stem cells exert therapeutic effects in an ALS mouse model.

    PubMed

    Sun, Haitao; Hou, Zongliu; Yang, Huaqiang; Meng, Mingyao; Li, Peng; Zou, Qingjian; Yang, Lujun; Chen, Yuxin; Chai, Huihui; Zhong, Huilin; Yang, Zara Zhuyun; Zhao, Jing; Lai, Liangxue; Jiang, Xiaodan; Xiao, Zhicheng

    2014-09-01

    Amyotrophic lateral sclerosis (ALS) is an adult-onset progressive neurodegenerative disease involving degeneration of motor neurons in the central nervous system. Stem cell treatment is a potential therapy for this fatal disorder. The human amniotic membrane (HAM), an extremely rich and easily accessible tissue, has been proposed as an attractive material in cellular therapy and regenerative medicine because of its advantageous characteristics. In the present study, we evaluate the long-term effects of a cellular treatment by intravenous administration of human amniotic mesenchymal stem cells (hAMSCs) derived from HAM into a hSOD1(G93A) mouse model. The mice received systemic administration of hAMSCs or phosphate-buffered saline (PBS) at the onset, progression and symptomatic stages of the disease. hAMSCs were detected in the spinal cord at the final stage of the disease, in the form of isolates or clusters and were negative for β-tubulin III and GFAP. Compared with the treatment with PBS, multiple hAMSC transplantations significantly retarded disease progression, extended survival, improved motor function, prevented motor neuron loss and decreased neuroinflammation in mice. These findings demonstrate that hAMSC transplantation is a promising cellular treatment for ALS.

  10. Transplantation with cultured stem cells derived from the human amniotic membrane for corneal alkali burns: an experimental study.

    PubMed

    Zeng, Wei; Li, Yanwei; Zeng, Guangwei; Yang, Bo; Zhu, Yu

    2014-01-01

    Amniotic membranes (AM) have been used in a wide range of clinical applications. We successfully extracted mesenchymal stem cells (MSCs) from human AM, but little is known about the use and efficacy of human amniotic membrane-derived mesenchymal stem cells (hAM-dMSCs) for the treatment of alkali burns. We utilized hAM-dMSCs transplantation, AM grafting, and their combined use in the treatment of alkali burns. An experimental model in rabbits was devised to analyze the use of these techniques with immunocytochemistry and ELISA. The survival and migration of hAM-dMSCs labeled by SPION in the host were assessed with Prussian blue staining. Compared with the control group, the treated groups demonstrated faster reconstruction of the corneal epithelium, and lower levels of corneal opacification and neovascularization within corneal alkali burns. Furthermore, dark blue-stained particles were detected in the limbus corneae at day 28. These results demonstrated the ability of hAM-dMSCs to enhance epithelial healing and reduce corneal opacification and neovascularization in corneal alkali wounds.

  11. A new method to repair recto-vaginal fistula: Use of human amniotic membrane in an animal model

    PubMed Central

    Roshanravan, Reza; Ghahramani, Leila; Hosseinzadeh, Massood; Mohammadipour, Mastoureh; Moslemi, Sam; Rezaianzadeh, Abbas; Safarpour, Ali Reza; Rahimikazerooni, Salar; Hosseini, Seyed Vahid

    2014-01-01

    Background: Recto-vaginal fistula is primarily one of the co-morbidities of vaginal delivery. These patients suffer from persistent malodor vaginal discharge. Various surgical techniques have been employed by surgeons in the course of time. This is the first trial of applying Human Amniotic Membrane (HAM) as a bio-prosthesis in repairing recto-vaginal fistula. Materials and Methods: In a prospective animal study, 8 mixed-breed female dogs weighing 23-27 kg with the age of 12-18 months were selected. They were randomly divided into two groups for standard recto-vaginal fistula repair and fistula repair with human amniotic membrane. The Kruskal-Wallis and Mann Whitney tests were performed to indicate statistical differences. Results: After 6 weeks, fistulas were evaluated both grossly and microscopically. In gross examination, there were no difference between the two groups and healing of fistula seemed to have been occurred in all dogs expect for one which had a persistent patent fistulous tract. Microscopic healing was scored according to epithelialization, collagenization inflammation, ulcer and necrosis of samples. Healing score was significantly higher in the HAM group than the standard group (P = 0.029). Conclusion: Our findings revealed that using HAM as a bio-prosthesis to repair recto-vaginal fistula would result in better surgical and histological outcomes comparing to simple repair. PMID:24804188

  12. Eicosanomic profiling reveals dominance of the epoxygenase pathway in human amniotic fluid at term in spontaneous labor.

    PubMed

    Maddipati, Krishna Rao; Romero, Roberto; Chaiworapongsa, Tinnakorn; Zhou, Sen-Lin; Xu, Zhonghui; Tarca, Adi L; Kusanovic, Juan Pedro; Munoz, Hernan; Honn, Kenneth V

    2014-11-01

    Lipid mediators play an important role in reproductive biology, especially, in parturition. Enhanced biosynthesis of eicosanoids, such as prostaglandin E2 (PGE2) and PGF2α, precedes the onset of labor as a result of increased expression of inducible cyclooxygenase 2 (COX-2) in placental tissues. Metabolism of arachidonic acid results in bioactive lipid mediators beyond prostaglandins that could significantly influence myometrial activity. Therefore, an unbiased lipidomic approach was used to profile the arachidonic acid metabolome of amniotic fluid. In this study, liquid chromatography-mass spectrometry was used for the first time to quantitate these metabolites in human amniotic fluid by comparing patients at midtrimester, at term but not in labor, and at term and in spontaneous labor. In addition to exposing novel aspects of COX pathway metabolism, this lipidomic study revealed a dramatic increase in epoxygenase- and lipoxygenase-pathway-derived lipid mediators in spontaneous labor with remarkable product selectivity. Despite their recognition as anti-inflammatory lipid mediators and regulators of ion channels, little is known about the epoxygenase pathway in labor. Epoxygenase pathway metabolites are established regulators of vascular homeostasis in cardiovascular and renal physiology. Their presence as the dominant lipid mediators in spontaneous labor at term portends a yet undiscovered physiological function in parturition.

  13. Human amniotic fluid for fertilization and culture of human embryos: results of clinical trials in human in vitro fertilization (IVF) programs.

    PubMed

    Gianaroli, L; Trounson, A; King, C; Ferraretti, A; Chiappazzo, L; Bafaro, G

    1989-08-01

    We report the outcome of clinical trials carried out in two IVF programs, comparing the use of human amniotic fluid (HAF) as a complete medium to Whittingham's T6 medium containing human serum (T6 + 10% HS) for egg incubation, insemination, embryo culture, and embryo transfer. There were no significant differences in the clinical trials between HAF used alone as a complete medium and T6 + 10% HS in fertilization rates of eggs, cleavage rates of embryos up to 48 hours in culture, pregnancy success rates after embryo replacement or the outcome of pregnancies. There was no advantage in using T6 + 10% HS for fertilization of eggs and HAF as a complete medium for embryo culture and transfer in any of the parameters examined. We conclude that HAF does not meet the complete requirements of human eggs and embryos in vitro and further developments of culture media are required to obtain embryo development equivalent to that in vivo.

  14. Metabolomics of Human Amniotic Fluid and Maternal Plasma during Normal Pregnancy.

    PubMed

    Orczyk-Pawilowicz, Magdalena; Jawien, Ewa; Deja, Stanislaw; Hirnle, Lidia; Zabek, Adam; Mlynarz, Piotr

    2016-01-01

    Metabolic profiles of amniotic fluid and maternal blood are sources of valuable information about fetus development and can be potentially useful in diagnosis of pregnancy disorders. In this study, we applied 1H NMR-based metabolic profiling to track metabolic changes occurring in amniotic fluid (AF) and plasma (PL) of healthy mothers over the course of pregnancy. AF and PL samples were collected in the 2nd (T2) and 3rd (T3) trimester, prolonged pregnancy (PP) until time of delivery (TD). A multivariate data analysis of both biofluids reviled a metabolic switch-like transition between 2nd and 3rd trimester, which was followed by metabolic stabilization throughout the rest of pregnancy probably reflecting the stabilization of fetal maturation and development. The differences were further tested using univariate statistics at α = 0.001. In plasma the progression from T2 to T3 was related to increasing levels of glycerol, choline and ketone bodies (3-hydroxybutyrate and acetoacetate) while pyruvate concentration was significantly decreased. In amniotic fluid, T2 to T3 transition was associated with decreasing levels of glucose, carnitine, amino acids (valine, leucine, isoleucine, alanine, methionine, tyrosine, and phenylalanine) and increasing levels of creatinine, succinate, pyruvate, choline, N,N-dimethylglycine and urocanate. Lactate to pyruvate ratio was decreased in AF and conversely increased in PL. The results of our study, show that metabolomics profiling can be used to better understand physiological changes of the complex interdependencies of the mother, the placenta and the fetus during pregnancy. In the future, these results might be a useful reference point for analysis of complicated pregnancies. PMID:27070784

  15. Metabolomics of Human Amniotic Fluid and Maternal Plasma during Normal Pregnancy.

    PubMed

    Orczyk-Pawilowicz, Magdalena; Jawien, Ewa; Deja, Stanislaw; Hirnle, Lidia; Zabek, Adam; Mlynarz, Piotr

    2016-01-01

    Metabolic profiles of amniotic fluid and maternal blood are sources of valuable information about fetus development and can be potentially useful in diagnosis of pregnancy disorders. In this study, we applied 1H NMR-based metabolic profiling to track metabolic changes occurring in amniotic fluid (AF) and plasma (PL) of healthy mothers over the course of pregnancy. AF and PL samples were collected in the 2nd (T2) and 3rd (T3) trimester, prolonged pregnancy (PP) until time of delivery (TD). A multivariate data analysis of both biofluids reviled a metabolic switch-like transition between 2nd and 3rd trimester, which was followed by metabolic stabilization throughout the rest of pregnancy probably reflecting the stabilization of fetal maturation and development. The differences were further tested using univariate statistics at α = 0.001. In plasma the progression from T2 to T3 was related to increasing levels of glycerol, choline and ketone bodies (3-hydroxybutyrate and acetoacetate) while pyruvate concentration was significantly decreased. In amniotic fluid, T2 to T3 transition was associated with decreasing levels of glucose, carnitine, amino acids (valine, leucine, isoleucine, alanine, methionine, tyrosine, and phenylalanine) and increasing levels of creatinine, succinate, pyruvate, choline, N,N-dimethylglycine and urocanate. Lactate to pyruvate ratio was decreased in AF and conversely increased in PL. The results of our study, show that metabolomics profiling can be used to better understand physiological changes of the complex interdependencies of the mother, the placenta and the fetus during pregnancy. In the future, these results might be a useful reference point for analysis of complicated pregnancies.

  16. The effects of cryopreservation on angiogenesis modulation activity of human amniotic membrane.

    PubMed

    Yazdanpanah, Ghasem; Paeini-Vayghan, Ghodsieh; Asadi, Samira; Niknejad, Hassan

    2015-12-01

    Amniotic membrane (AM), as the innermost layer of placenta, has side dependent effects on the angiogenesis. Cryopreservation is a necessary process to avoid the challenging problems of fresh tissues; a procedure which makes the AM ready-to-use. Since the cryopreservation can influence the AM characteristics for experimental and clinical purposes, in this study the effects of cryopreservation were evaluated on angiogenesis modulation activity of the AM compared to fresh tissues in an animal model. The AM was implanted mesenchymal side up or epithelial side up in a rat dorsal skinfold chamber. The length and number of branches of formed capillaries were measured via intravital microscopy after 7 days. The amount of IL-8 (interleukin-8) and TIMP-2 (Tissue Inhibitor of Matrix Metalloproteinase-2) as two factors in amniotic cells which have great impacts on angiogenesis were evaluated using ELISA assay. The epithelial surface of cryopreserved AM had inhibitory effects on vessel formation. The cryopreserved amniotic mesenchymal side increased the vessel length and sprout. The result of cryopreserved AM on angiogenesis was similar to that of fresh tissues. The levels of IL-8 and TIMP-2 in cryopreserved samples were significantly less than fresh AMs which shows that angio-modulatory properties are not limited to the effects of amnion epithelial and mesenchymal stem cells and the other components such as extracellular matrix may contribute in angio-modulatory effects. These promising results show that inducing and inhibitory effects of the AM, which make it an appropriate candidate for different clinical situations, were maintained after cryopreservation.

  17. Metabolomics of Human Amniotic Fluid and Maternal Plasma during Normal Pregnancy

    PubMed Central

    Deja, Stanislaw; Hirnle, Lidia; Zabek, Adam; Mlynarz, Piotr

    2016-01-01

    Metabolic profiles of amniotic fluid and maternal blood are sources of valuable information about fetus development and can be potentially useful in diagnosis of pregnancy disorders. In this study, we applied 1H NMR-based metabolic profiling to track metabolic changes occurring in amniotic fluid (AF) and plasma (PL) of healthy mothers over the course of pregnancy. AF and PL samples were collected in the 2nd (T2) and 3rd (T3) trimester, prolonged pregnancy (PP) until time of delivery (TD). A multivariate data analysis of both biofluids reviled a metabolic switch-like transition between 2nd and 3rd trimester, which was followed by metabolic stabilization throughout the rest of pregnancy probably reflecting the stabilization of fetal maturation and development. The differences were further tested using univariate statistics at α = 0.001. In plasma the progression from T2 to T3 was related to increasing levels of glycerol, choline and ketone bodies (3-hydroxybutyrate and acetoacetate) while pyruvate concentration was significantly decreased. In amniotic fluid, T2 to T3 transition was associated with decreasing levels of glucose, carnitine, amino acids (valine, leucine, isoleucine, alanine, methionine, tyrosine, and phenylalanine) and increasing levels of creatinine, succinate, pyruvate, choline, N,N-dimethylglycine and urocanate. Lactate to pyruvate ratio was decreased in AF and conversely increased in PL. The results of our study, show that metabolomics profiling can be used to better understand physiological changes of the complex interdependencies of the mother, the placenta and the fetus during pregnancy. In the future, these results might be a useful reference point for analysis of complicated pregnancies. PMID:27070784

  18. Surgical Outcomes of Deep Superior Sulcus Augmentation Using Acellular Human Dermal Matrix in Anophthalmic or Phthisis Socket.

    PubMed

    Cho, Won-Kyung; Jung, Su-Kyung; Paik, Ji-Sun; Yang, Suk-Woo

    2016-07-01

    Patients with anophthalmic or phthisis socket suffer from cosmetic problems. To resolve those problems, the authors present the surgical outcomes of deep superior sulcus (DSS) augmentation using acellular dermal matrix in patients with anophthalmic or phthisis socket. The authors retrospectively reviewed anophthalmic or phthisis patients who underwent surgery for DSS augmentation using acellular dermal matrix. To evaluate surgical outcomes, the authors focused on 3 aspects: the possibility of wearing contact prosthesis, the degree of correction of the DSS, and any surgical complications. The degree of correction of DSS was classified as excellent: restoration of superior sulcus enough to remove sunken sulcus shadow; fair: gain of correction effect but sunken shadow remained; or fail: no effect of correction at all. Ten eyes of 10 patients were included. There was a mean 21.3 ± 37.1-month period from evisceration or enucleation to the operation for DSS augmentation. All patients could wear contact prosthesis after the operation (100%). The degree of correction was excellent in 8 patients (80%) and fair in 2. Three of 10 (30%) showed complications: eyelid entropion, upper eyelid multiple creases, and spontaneous wound dehiscence followed by inflammation after stitch removal. Uneven skin surface and paresthesia in the forehead area of the affected eye may be observed after surgery. The overall surgical outcomes were favorable, showing an excellent degree of correction of DSS and low surgical complication rates. This procedure is effective for patients who have DSS in the absence or atrophy of the eyeball.

  19. Surgical Outcomes of Deep Superior Sulcus Augmentation Using Acellular Human Dermal Matrix in Anophthalmic or Phthisis Socket.

    PubMed

    Cho, Won-Kyung; Jung, Su-Kyung; Paik, Ji-Sun; Yang, Suk-Woo

    2016-07-01

    Patients with anophthalmic or phthisis socket suffer from cosmetic problems. To resolve those problems, the authors present the surgical outcomes of deep superior sulcus (DSS) augmentation using acellular dermal matrix in patients with anophthalmic or phthisis socket. The authors retrospectively reviewed anophthalmic or phthisis patients who underwent surgery for DSS augmentation using acellular dermal matrix. To evaluate surgical outcomes, the authors focused on 3 aspects: the possibility of wearing contact prosthesis, the degree of correction of the DSS, and any surgical complications. The degree of correction of DSS was classified as excellent: restoration of superior sulcus enough to remove sunken sulcus shadow; fair: gain of correction effect but sunken shadow remained; or fail: no effect of correction at all. Ten eyes of 10 patients were included. There was a mean 21.3 ± 37.1-month period from evisceration or enucleation to the operation for DSS augmentation. All patients could wear contact prosthesis after the operation (100%). The degree of correction was excellent in 8 patients (80%) and fair in 2. Three of 10 (30%) showed complications: eyelid entropion, upper eyelid multiple creases, and spontaneous wound dehiscence followed by inflammation after stitch removal. Uneven skin surface and paresthesia in the forehead area of the affected eye may be observed after surgery. The overall surgical outcomes were favorable, showing an excellent degree of correction of DSS and low surgical complication rates. This procedure is effective for patients who have DSS in the absence or atrophy of the eyeball. PMID:27258711

  20. Gender-Typed Play and Amniotic Testosterone

    ERIC Educational Resources Information Center

    Knickmeyer, Rebecca Christine; Wheelwright, Sally; Taylor, Kevin; Raggatt, Peter; Hackett, Gerald; Baron-Cohen, Simon

    2005-01-01

    Sex differences in play are apparent in a number of mammalian species, including humans. Prenatal testosterone may contribute to these differences. The authors report the first attempt to correlate gender-typed play in a normative sample of humans with measurements of amniotic testosterone (aT). Testosterone was measured in the amniotic fluid of…

  1. Human amniotic fluid stem cells as a model for functional studies of genes involved in human genetic diseases or oncogenesis.

    PubMed

    Rosner, Margit; Dolznig, Helmut; Schipany, Katharina; Mikula, Mario; Brandau, Oliver; Hengstschläger, Markus

    2011-09-01

    Besides their putative usage for therapies, stem cells are a promising tool for functional studies of genes involved in human genetic diseases or oncogenesis. For this purpose induced pluripotent stem (iPS) cells can be derived from patients harbouring specific mutations. In contrast to adult stem cells, iPS cells are pluripotent and can efficiently be grown in culture. However, iPS cells are modulated due to the ectopic induction of pluripotency, harbour other somatic mutations accumulated during the life span of the source cells, exhibit only imperfectly cleared epigenetic memory of the source cell, and are often genomically instable. In addition, iPS cells from patients only allow the investigation of mutations, which are not prenatally lethal. Embryonic stem (ES) cells have a high proliferation and differentiation potential, but raise ethical issues. Human embryos, which are not transferred in the course of in vitro fertilization, because of preimplantation genetic diagnosis of a genetic defect, are still rarely donated for the establishment of ES cell lines. In addition, their usage for studies on gene functions for oncogenesis is hampered by the fact the ES cells are already tumorigenic per se. In 2003 amniotic fluid stem (AFS) cells have been discovered, which meanwhile have been demonstrated to harbour the potential to differentiate into cells of all three germ layers. Monoclonal human AFS cell lines derived from amniocenteses have a high proliferative potential, are genomically stable and are not associated with ethical controversies. Worldwide amniocenteses are performed for routine human genetic diagnosis. We here discuss how generation and banking of monoclonal human AFS cell lines with specific chromosomal aberrations or monogenic disease mutations would allow to study the functional consequences of disease causing mutations. In addition, recently a protocol for efficient and highly reproducible siRNA-mediated long-term knockdown of endogenous gene

  2. In vitro assessment of biodurability: acellular systems.

    PubMed Central

    de Meringo, A; Morscheidt, C; Thélohan, S; Tiesler, H

    1994-01-01

    The assessment of biodurability of man-made vitreous fibers is essential to the limitation of health hazards associated with human exposure to environments in which respirable fibers are present. In vitro acellular systems provide effective test methods of measuring fiber solubility provided care is taken to select the most suitable solvent and test conditions for the specific fiber type and dimension. PMID:7882955

  3. In vitro differentiation of human amniotic fluid-derived cells: augmentation towards a neuronal dopaminergic phenotype.

    PubMed

    Pfeiffer, Shona; McLaughlin, David

    2010-09-01

    Amniotic fluid is known to yield a number of cell types which are multipotent, ethically derived, genetically stable, easily grown, expanded and possess favourable immunogenicity, which has resulted in an increasing interest for use in various neuronal disorders such as Parkinson's disease. The neuronal potential of cells derived from the adherent fraction of amniotic fluid, routinely taken by amniocentesis, are least explored. The aim of the present study was to investigate the capacity of these cells for neuronal and dopaminergic differentiation using in vitro differentiation protocols with canonical inductive factors not previously tested. To do this, samples derived from multiple donors were grown under four conditions: standard serum-containing media, NB (neurobasal) media designed specifically for propagation and maintenance of neuronal cells, NB media with addition of retinoic acid and BDNF (brain-derived neurotrophic factor) for NI (neuronal induction), and NB media with addition of FGF8 (fibroblast growth factor-8) and Shh (sonic hedgehog) after NI. Our results showed the presence of multiple neuronal markers after growth in serum-containing medium [TUJ1, MAP2, NF-M, TH (tyrosine hydroxylase)], which was significantly up-regulated after serum withdrawal in NB medium alone with induction of NeuN (neuronal nuclei) and NSE (neuron-specific enolase). NI and DA.I (dopaminergic induction) was accompanied by further increases in expression and a distinct transition to a sustained neuronal morphology. Western blot analysis confirmed increasing TH expression and NURR1, expressed in base serum-containing media, found to be down-regulated after induction. In conclusion, these cells possess a highly favourable base neuronal and dopaminergic prepotential, which may easily be accentuated by standard induction protocols. PMID:20388119

  4. Magnetic Resonance Imaging of Human-Derived Amniotic Membrane Stem Cells Using PEGylated Superparamagnetic Iron Oxide Nanoparticles

    PubMed Central

    Naseroleslami, Maryam; Parivar, Kazem; Khoei, Samideh; Aboutaleb, Nahid

    2016-01-01

    Objective The label and detection of cells injected into target tissues is an area of focus for researchers. Iron oxide nanoparticles can be used to label cells as they have special characteristics. The purpose of this study is to examine the effects of iron oxide nanoparticles on human-derived amniotic membrane stem cell (hAMCs) survival and to investigate the magnetic properties of these nanoparticles with increased contrast in magnetic resonance imaging (MRI). Materials and Methods In this experimental study, we initially isolated mesenchymal stem cells from amniotic membranes and analyzed them by flow cytometry. In addition, we synthesized superparamagnetic iron oxide nanoparticles (SPIONs) and characterized them by various methods. The SPIONs were incubated with hAMCs at concentrations of 25-800 μg/mL. The cytotoxicity of nanoparticles on hAMCs was measured by the MTT assay. Next, we evaluated the effectiveness of the magnetic nanoparticles as MRI contrast agents. Solutions of SPION were prepared in water at different iron concentrations for relaxivity measurements by a 1.5 Tesla clinical MRI instrument. Results The isolated cells showed an adherent spindle shaped morphology. Polyethylene glycol (PEG)-coated SPIONs exhibited a spherical morphology. The average particle size was 20 nm and magnetic saturation was 60 emu/g. Data analysis showed no significant reduction in the percentage of viable cells (97.86 ± 0.41%) after 72 hours at the 125 μg/ml concentration compared with the control. The relaxometry results of this SPION showed a transverse relaxivity of 6.966 (μg/ml.s)-1 Conclusion SPIONs coated with PEG used in this study at suitable concentrations had excellent labeling efficiency and biocompatibility for hAMCs. PMID:27602314

  5. Magnetic Resonance Imaging of Human-Derived Amniotic Membrane Stem Cells Using PEGylated Superparamagnetic Iron Oxide Nanoparticles

    PubMed Central

    Naseroleslami, Maryam; Parivar, Kazem; Khoei, Samideh; Aboutaleb, Nahid

    2016-01-01

    Objective The label and detection of cells injected into target tissues is an area of focus for researchers. Iron oxide nanoparticles can be used to label cells as they have special characteristics. The purpose of this study is to examine the effects of iron oxide nanoparticles on human-derived amniotic membrane stem cell (hAMCs) survival and to investigate the magnetic properties of these nanoparticles with increased contrast in magnetic resonance imaging (MRI). Materials and Methods In this experimental study, we initially isolated mesenchymal stem cells from amniotic membranes and analyzed them by flow cytometry. In addition, we synthesized superparamagnetic iron oxide nanoparticles (SPIONs) and characterized them by various methods. The SPIONs were incubated with hAMCs at concentrations of 25-800 μg/mL. The cytotoxicity of nanoparticles on hAMCs was measured by the MTT assay. Next, we evaluated the effectiveness of the magnetic nanoparticles as MRI contrast agents. Solutions of SPION were prepared in water at different iron concentrations for relaxivity measurements by a 1.5 Tesla clinical MRI instrument. Results The isolated cells showed an adherent spindle shaped morphology. Polyethylene glycol (PEG)-coated SPIONs exhibited a spherical morphology. The average particle size was 20 nm and magnetic saturation was 60 emu/g. Data analysis showed no significant reduction in the percentage of viable cells (97.86 ± 0.41%) after 72 hours at the 125 μg/ml concentration compared with the control. The relaxometry results of this SPION showed a transverse relaxivity of 6.966 (μg/ml.s)-1 Conclusion SPIONs coated with PEG used in this study at suitable concentrations had excellent labeling efficiency and biocompatibility for hAMCs.

  6. Direct Reprogramming of Human Amniotic Fluid Stem Cells by OCT4 and Application in Repairing of Cerebral Ischemia Damage

    PubMed Central

    Qin, Mingde; Chen, Ruihua; Li, Hong; Liang, Hansi; Xue, Qun; Li, Fang; Chen, Ying; Zhang, Xueguang

    2016-01-01

    Amniotic fluid stem cells (AFSCs) are a type of fetal stem cell whose stemness encompasses both embryonic and adult stem cells, suggesting that they may be easily and efficiently reprogrammed into induced pluripotent stem cells (iPSCs). To further simplify the reprogramming process, the creation of AFSC-derived iPSCs using a single factor is desirable. Here we report the generation of one-factor human AFSC-iPSCs (AiPSCs) from human AFSCs by ectopic expression of the transcription factor OCT4. Just like human embryonic stem cells, AiPSCs exhibited similar epigenetic status, global gene expression profiles, teratoma formation and in vitro & in vivo pluripotency. Our results indicate that the OCT4 is necessary and sufficient to directly reprogram human AFSCs into pluripotent AiPSCs. Moreover, reflecting the similar memory characteristics of AFSCs and neural stem cells, we show that AiPSC membrane-derived vesicles (MVs) repair cerebral ischemia damage. We anticipate that the successful generation of one-factor AiPSCs will facilitate the creation of patient-specific pluripotent stem cells without the need for transgenic expression of oncogenes. Moreover, MVs from tissue-specific AiPSCs have potential in tissue repair, representing a novel application of iPSCs. PMID:27019637

  7. Tuberin and PRAS40 are anti-apoptotic gatekeepers during early human amniotic fluid stem-cell differentiation.

    PubMed

    Fuchs, Christiane; Rosner, Margit; Dolznig, Helmut; Mikula, Mario; Kramer, Nina; Hengstschläger, Markus

    2012-03-01

    Embryoid bodies (EBs) are three-dimensional multicellular aggregates allowing the in vitro investigation of stem-cell differentiation processes mimicking early embryogenesis. Human amniotic fluid stem (AFS) cells harbor high proliferation potential, do not raise the ethical issues of embryonic stem cells, have a lower risk for tumor development, do not need exogenic induction of pluripotency and are chromosomal stable. Starting from a single human AFS cell, EBs can be formed accompanied by the differentiation into cells of all three embryonic germ layers. Here, we report that siRNA-mediated knockdown of the endogenous tuberous sclerosis complex-2 (TSC2) gene product tuberin or of proline-rich Akt substrate of 40 kDa (PRAS40), the two major negative regulators of mammalian target of rapamycin (mTOR), leads to massive apoptotic cell death during EB development of human AFS cells without affecting the endodermal, mesodermal and ectodermal cell differentiation spectrum. Co-knockdown of endogenous mTOR demonstrated these effects to be mTOR-dependent. Our findings prove this enzyme cascade to be an essential anti-apoptotic gatekeeper of stem-cell differentiation during EB formation. These data allow new insights into the regulation of early stem-cell maintenance and differentiation and identify a new role of the tumor suppressor tuberin and the oncogenic protein PRAS40 with the relevance for a more detailed understanding of the pathogenesis of diseases associated with altered activities of these gene products.

  8. Evaluation of Distinct Freezing Methods and Cryoprotectants for Human Amniotic Fluid Stem Cells Cryopreservation

    PubMed Central

    Janz, Felipe de Lara; Debes, Adriana de Aguiar; Cavaglieri, Rita de Cássia; Duarte, Sérgio Aloísio; Romão, Carolina Martinez; Morón, Antonio Fernandes; Zugaib, Marcelo; Bydlowski, Sérgio Paulo

    2012-01-01

    Amniotic fluid (AF) was described as a potential source of mesenchymal stem cells (MSCs) for biomedicine purposes. Therefore, evaluation of alternative cryoprotectants and freezing protocols capable to maintain the viability and stemness of these cells after cooling is still needed. AF stem cells (AFSCs) were tested for different freezing methods and cryoprotectants. Cell viability, gene expression, surface markers, and plasticity were evaluated after thawing. AFSCs expressed undifferentiated genes Oct4 and Nanog; presented typical markers (CD29, CD44, CD90, and CD105) and were able to differentiate into mesenchymal lineages. All tested cryoprotectants preserved the features of AFSCs however, variations in cell viability were observed. In this concern, dimethyl sulfoxide (Me2SO) showed the best results. The freezing protocols tested did not promote significant changes in the AFSCs viability. Time programmed and nonprogrammed freezing methods could be used for successful AFSCs cryopreservation for 6 months. Although tested cryoprotectants maintained undifferentiated gene expression, typical markers, and plasticity of AFSCs, only Me2SO and glycerol presented workable viability ratios. PMID:22665987

  9. Amniotic fluid (image)

    MedlinePlus

    Amniotic fluid surrounds the growing fetus in the womb and protects the fetus from injury and temperature changes. ... of fetal movement and permits musculoskeletal development. The amniotic fluid can be withdrawn in a procedure called amniocentsis ...

  10. [Differentiation of human amniotic mesenchymal stem cells into insulin-secreting cells induced by regenerating pancreatic extract].

    PubMed

    Zhang, Yanmei; Wang, Dianliang; Zeng, Hongyan; Wang, Lieming; Sun, Jinwei; Zhang, Zhen; Dong, Shasha

    2012-02-01

    In this study, the natural biological inducer, rat regenerating pancreatic extract (RPE), was used to induce human amniotic mesenchymal stem cells (hAMSCs) into insulin-secreting cells. We excised 60% of rat pancreas in order to stimulate pancreatic regeneration. RPE was extracted and used to induce hAMSCs at a final concentration of 20 microg/mL. The experiment methods used were as follows: morphological-identification, dithizone staining, immumofluorescence analysis, reverse transcription-PCR (RT-PCR) and insulin secretion stimulated by high glucose. The results show that the cell morphology of passge3 hAMSCs changed significantly after the induction of RPE, resulting in cluster shape after induction for 15 days. Dithizone staining showed that there were scarlet cell masses in RPE-treated culture. Immumofluorescence analysis indicated that induced cells were insulin-positive expression. RT-PCR showed the positive expression of human islet-related genes Pdx1 and insulin in the induced cells. The result of insulin secretion stimulated by high glucose indicated that insulin increasingly secreted and then kept stable with prolongation of high glucose stimulation. In conclusion, hAMSCs had the potential to differentiate into insulin-secreting cells induced by RPE in vitro. PMID:22667123

  11. Involvement of gene methylation changes in the differentiation of human amniotic epithelial cells into islet-like cell clusters.

    PubMed

    Peng, Lin; Wang, Jian; Lu, Guangxiu

    2014-09-01

    Insulin-dependent diabetes results from destruction of the insulin-producing β-cells of the pancreas. Islet cell transplantation is a promising cure for diabetes. Here, we induced human amniotic epithelial cells (hAECs) to differentiate into islet-like cell clusters by nicotinamide plus betacellulin in vitro, and further investigated the DNA methylation status by a Nimble MeDIP microarray before and after cell differentiation to shed light on the molecular mechanisms of this differentiation. In addition, 5-Aza-2'-deoxycytidine was used to investigate whether the differentiation of hAECs into islet-like cells occurred through demethylation. Purified hAECs (CK18(+)/E-cadherin(+)/CD29(+)/CD90(-)/CD34(-)/CD45(-)) were isolated from human amnia. After induction, hAECs were found to be insulin positive and sensitive to glucose, indicating successful induction to islet-like cells. The methylation status of cell cytoskeleton-related genes was down-regulated and that of negative regulation of cell adhesion-related genes was up-regulated. The methylation status of pancreas development-related genes such as HNF1α and DGAT1 was decreased in hAECs after induction. After brief demethylation, INS gene expression was up-regulated in islet-like cell clusters, suggesting that DNA methylation changes were associated with the differentiation of hAECs into islet-like cell clusters. PMID:24945458

  12. Involvement of gene methylation changes in the differentiation of human amniotic epithelial cells into islet-like cell clusters.

    PubMed

    Peng, Lin; Wang, Jian; Lu, Guangxiu

    2014-09-01

    Insulin-dependent diabetes results from destruction of the insulin-producing β-cells of the pancreas. Islet cell transplantation is a promising cure for diabetes. Here, we induced human amniotic epithelial cells (hAECs) to differentiate into islet-like cell clusters by nicotinamide plus betacellulin in vitro, and further investigated the DNA methylation status by a Nimble MeDIP microarray before and after cell differentiation to shed light on the molecular mechanisms of this differentiation. In addition, 5-Aza-2'-deoxycytidine was used to investigate whether the differentiation of hAECs into islet-like cells occurred through demethylation. Purified hAECs (CK18(+)/E-cadherin(+)/CD29(+)/CD90(-)/CD34(-)/CD45(-)) were isolated from human amnia. After induction, hAECs were found to be insulin positive and sensitive to glucose, indicating successful induction to islet-like cells. The methylation status of cell cytoskeleton-related genes was down-regulated and that of negative regulation of cell adhesion-related genes was up-regulated. The methylation status of pancreas development-related genes such as HNF1α and DGAT1 was decreased in hAECs after induction. After brief demethylation, INS gene expression was up-regulated in islet-like cell clusters, suggesting that DNA methylation changes were associated with the differentiation of hAECs into islet-like cell clusters.

  13. Human amniotic epithelial cells induce localized cell-mediated immune privilege in vitro: implications for pancreatic islet transplantation.

    PubMed

    Qureshi, Khalid M; Oliver, Robert J; Paget, Michelle B; Murray, Hilary E; Bailey, Clifford J; Downing, Richard

    2011-01-01

    Chronic systemic immunosuppression in cell replacement therapy restricts its clinical application. This study sought to explore the potential of cell-based immune modulation as an alternative to immunosuppressive drug therapy in the context of pancreatic islet transplantation. Human amniotic epithelial cells (AEC) possess innate anti-inflammatory and immunosuppressive properties that were utilized to create localized immune privilege in an in vitro islet cell culture system. Cellular constructs composed of human islets and AEC (islet/AEC) were bioengineered under defined rotational cell culture conditions. Insulin secretory capacity was validated by glucose challenge and immunomodulatory potential characterized using a peripheral blood lymphocyte (PBL) proliferation assay. Results were compared to control constructs composed of islets or AEC cultured alone. Studies employing AEC-conditioned medium examined the role of soluble factors, and fluorescence immunocytochemistry was used to identify putative mediators of the immunosuppressive response in isolated AEC monocultures. Sustained, physiologically appropriate insulin secretion was observed in both islets and islet/AEC constructs. Activation of resting PBL proliferation occurred on exposure to human islets alone but this response was significantly (p < 0.05) attenuated by the presence of AEC and AEC-conditioned medium. Mitogen (phytohaemagglutinin, 5 μg/ml)-induced PBL proliferation was sustained on contact with isolated islets but abrogated by AEC, conditioned medium, and the islet/AEC constructs. Immunocytochemical analysis of AEC monocultures identified a subpopulation of cells that expressed the proapoptosis protein Fas ligand. This study demonstrates that human islet/AEC constructs exhibit localized immunosuppressive properties with no impairment of β-cell function. The data suggest that transplanted islets may benefit from the immune privilege status conferred on them as a consequence of their close

  14. Human amniotic epithelial cells as novel feeder layers for promoting ex vivo expansion of limbal epithelial progenitor cells.

    PubMed

    Chen, Ying-Ting; Li, Wei; Hayashida, Yasutaka; He, Hua; Chen, Szu-Yu; Tseng, David Y; Kheirkhah, Ahmad; Tseng, Scheffer C G

    2007-08-01

    Human amniotic epithelial cells (HAECs) are a unique embryonic cell source that potentially can be used as feeder layers for expanding different types of stem cells. In vivo, HAECs uniformly expressed pan-cytokeratins (pan-CK) and heterogeneously expressed vimentin (Vim). The two phenotypes expressing either pan-CK(+)/Vim(+) or pan-CK(+)/Vim(-) were maintained in serum-free media with high calcium. In contrast, all HAECs became pan-CK(+)/Vim(+) in serum-containing media, which also promoted HAEC proliferation for at least eight passages, especially supplemented with epidermal growth factor and insulin. Mitomycin C-arrested HAEC feeder layers were more effective in promoting clonal growth of human limbal epithelial progenitors than conventional 3T3 murine feeder layers. Cells in HAEC-supported clones were uniformly smaller, sustained more proliferation, and expressed less CK12 and connexin 43 but higher levels of stem cell-associated markers such as p63, Musashi-1, and ATP-binding cassette subfamily G2 than those of 3T3-supported clones. Subculturing of clonally expanded limbal progenitors from HAEC feeder layers, but not from 3T3 feeder layers, gave rise to uniformly p63-positive epithelial progenitor cells as well as nestin-positive neuronal-like progenitors. Collectively, these results indicated that HAECs can be used as a human feeder layer equivalent for more effective ex vivo expansion of adult epithelial stem cells from the human limbus. Disclosure of potential conflicts of interest is found at the end of this article.

  15. Feasibility of Human Amniotic Fluid Derived Stem Cells in Alleviation of Neuropathic Pain in Chronic Constrictive Injury Nerve Model

    PubMed Central

    Chiang, Chien-Yi; Liu, Shih-An; Sheu, Meei-Ling; Chen, Fu-Chou; Chen, Chun-Jung; Su, Hong-Lin; Pan, Hung-Chuan

    2016-01-01

    Purpose The neurobehavior of neuropathic pain by chronic constriction injury (CCI) of sciatic nerve is very similar to that in humans, and it is accompanied by a profound local inflammation response. In this study, we assess the potentiality of human amniotic fluid derived mesenchymal stem cells (hAFMSCs) for alleviating the neuropathic pain in a chronic constriction nerve injury model. Methods and Methods This neuropathic pain animal model was conducted by four 3–0 chromic gut ligatures loosely ligated around the left sciatic nerve in Sprague—Dawley rats. The intravenous administration of hAFMSCs with 5x105 cells was conducted for three consecutive days. Results The expression IL-1β, TNF-α and synaptophysin in dorsal root ganglion cell culture was remarkably attenuated when co-cultured with hAFMSCs. The significant decrease of PGP 9.5 in the skin after CCI was restored by administration of hAFMSCs. Remarkably increased expression of CD 68 and TNF-α and decreased S-100 and neurofilament expression in injured nerve were rescued by hAFMSCs administration. Increases in synaptophysin and TNF-α over the dorsal root ganglion were attenuated by hAFMSCs. Significant expression of TNF-α and OX-42 over the dorsal spinal cord was substantially attenuated by hAFMSCs. The increased amplitude of sensory evoked potential as well as expression of synaptophysin and TNF-α expression was alleviated by hAFMSCs. Human AFMSCs significantly improved the threshold of mechanical allodynia and thermal hyperalgesia as well as various parameters of CatWalk XT gait analysis. Conclusion Human AFMSCs administration could alleviate the neuropathic pain demonstrated in histomorphological alteration and neurobehavior possibly through the modulation of the inflammatory response. PMID:27441756

  16. Analysis of nifedipine in human plasma and amniotic fluid by liquid chromatography-tandem mass spectrometry and its application to clinical pharmacokinetics in hypertensive pregnant women.

    PubMed

    Filgueira, Gabriela Campos de Oliveira; Filgueira, Osmany Alberto Silva; Carvalho, Daniela Miarelli; Marques, Maria Paula; Moisés, Elaine Christine Dantas; Duarte, Geraldo; Lanchote, Vera Lucia; Cavalli, Ricardo Carvalho

    2015-07-01

    Nifedipine is a dihydropyridine calcium channel blocker used for the treatment of hypertension in pregnant women. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for analysis of nifedipine in human plasma and amniotic fluid. Separation of nifedipine and nitrendipine (IS) was performed using a LiChroCART(®) RP-Select B column and a mixture of water:acetonitrile:glacial acetic acid (30:70:0.5 v/v) as the mobile phase. Aliquots of 500μL of biological samples were extracted at pH 13 using dichloromethane:n-pentane (3:7 v/v). The validated method was applied to a study of the pharmacokinetics of nifedipine in human plasma and amniotic fluid samples collected up to 12h after administration of the last slow-release nifedipine (20mg/12h) dose to 12 hypertensive pregnant women. The estimated pharmacokinetic parameters of nifedipine showed a mean AUC(0-12) of 250.2ngh/mL, ClT/F of 89.2L/h, Vd/F of 600.0L and t1/2 5.1h. The mean amniotic fluid/plasma concentration ratio was 0.05. The methods proved to be highly sensitive by showing a lower quantification limit of 0.1ng/mL for both matrices. And this study reports for the first time the complete development and validation of the method to quantify nifedipine in amniotic fluid using LC-MS-MS.

  17. Effects of different doses of gamma irradiation on oxygen and water vapour transmission rate of preserved human amniotic membrane.

    PubMed

    Zahari, Nor Kamalia; Sheikh Ab Hamid, Suzina; Yusof, Norimah

    2015-03-01

    Preserved human amniotic membrane either air dried or glycerol preserved has been used effectively to treat superficial and partial thickness wounds without leaving any obvious hypertrophic scar. The preserved amnion, sterilised by ionising radiation, is known as an effective barrier for heat, fluid and protein loss while adheres nicely on wound. Air drying slightly reduced the oxygen transmission rate (OTR) of the amnion and the value significantly dropped after 15 kGy (p < 0.05). Glycerol preservation significantly reduced (p < 0.05) the OTR indicating less oxygen transmitted through the well structured cells of the amnion. Increase in the OTR with the increasing radiation doses up to 35 kGy possibly due to direct effects of radiation that resulted in large intercellular gaps. Both preservation methods significantly increased (p < 0.05) the water vapour transmission rate (WVTR). However, the low WVTR in the air dried amnion at 15 and 25 kGy was postulated due to cross-linking of collagen. Changes in the biophysical properties can be linked to direct and indirect effects of radiation on collagen bundles. The radiation dose of 25 kGy caused no adverse effect on biophysical properties hence it is still acceptable to sterilize both the air dried and the glycerol preserved amnions.

  18. Evaluation of human amniotic membrane as a wound dressing for split-thickness skin-graft donor sites.

    PubMed

    Loeffelbein, Denys J; Rohleder, Nils H; Eddicks, Matthias; Baumann, Claudia M; Stoeckelhuber, Mechthild; Wolff, Klaus-D; Drecoll, Enken; Steinstraesser, Lars; Hennerbichler, Simone; Kesting, Marco R

    2014-01-01

    Human amniotic membrane (HAM) has been used as a biomaterial in various surgical procedures and exceeds some qualities of common materials. We evaluated HAM as wound dressing for split-thickness skin-graft (STSG) donor sites in a swine model (Part A) and a clinical trial (Part B). Part A: STSG donor sites in 4 piglets were treated with HAM or a clinically used conventional polyurethane (PU) foil (n = 8 each). Biopsies were taken on days 5, 7, 10, 20, 40, and 60 and investigated immunohistochemically for alpha-smooth muscle actin (αSMA: wound contraction marker), von Willebrand factor (vWF: angiogenesis), Ki-67 (cell proliferation), and laminin (basement membrane integrity). Part B: STSG donor sites in 45 adult patients (16 female/29 male) were treated with HAM covered by PU foam, solely by PU foam, or PU foil/paraffin gauze (n = 15 each). Part A revealed no difference in the rate of wound closure between groups. HAM showed improved esthetic results and inhibitory effects on cicatrization. Angioneogenesis was reduced, and basement membrane formation was accelerated in HAM group. Part B: no difference in re-epithelialization/infection rate was found. HAM caused less ichor exudation and less pruritus. HAM has no relevant advantage over conventional dressings but might be a cost-effective alternative.

  19. [Optimization of in vitro culture conditions for human amniotic epithelial cells and expression of stem cell markers].

    PubMed

    Chen, You-Yi; Lu, Yan; Wang, Ke; Wang, Yan; Wu, Dong-Ying; Liu, Bin; Yang, Ying; Lü, Shuang-Hong

    2011-04-01

    This study was purposed to optimize the culture conditions of the human amniotic epithelium cells (hAEC) in vitro, and detect the expression of hAEC pluripotent markers. Amnion tissues were separated from the underlying chorion through the spongy layer immediately after elective cesarean section of healthy pregnancy women at term. After the subsequent exposure to trypsin digestion, hAEC were cultured in DMEM with different supplements. The growth and proliferation potential of hAEC was evaluated, and the expression of cultured hAEC pluripotent markers was detected by using flow cytometry and immunohistochemistry methods. The results indicated that when being cultured in the mediums similar to that of embryonic stem cell culture supplemented with 10 ng/ml EGF, the hAEC grew better and the time for passage was shortened. In addition, compared to other culture conditions, under this condition, the cells could be passaged up to 5 times as much without obvious morphological changes, and the pluripotent marker SSEA-4 was detected in the cultured cells by flow cytometry. Meanwhile, the detection of immunofluorescence showed the expression of vimentin in cultured hAEC was strengthened as compared with primary cells. It is concluded that the culture condition similar to that for embryonic stem cells supplemented with EGF facilitates the proliferation and passage of hAEC in vitro.

  20. Electrospun Poly(l-lactide)/Poly(ethylene glycol) Scaffolds Seeded with Human Amniotic Mesenchymal Stem Cells for Urethral Epithelium Repair.

    PubMed

    Lv, Xiaokui; Guo, Qianping; Han, Fengxuan; Chen, Chunyang; Ling, Christopher; Chen, Weiguo; Li, Bin

    2016-01-01

    Tissue engineering-based urethral replacement holds potential for repairing large segmental urethral defects, which remains a great challenge at present. This study aims to explore the potential of combining biodegradable poly(l-lactide) (PLLA)/poly(ethylene glycol) (PEG) scaffolds and human amniotic mesenchymal cells (hAMSCs) for repairing urethral defects. PLLA/PEG fibrous scaffolds with various PEG fractions were fabricated via electrospinning. The scaffolds were then seeded with hAMSCs prior to implantation in New Zealand male rabbits that had 2.0 cm-long defects in the urethras. The rabbits were randomly divided into three groups. In group A, hAMSCs were grown on PLLA/PEG scaffolds for two days and then implanted to the urethral defects. In group B, only the PLLA/PEG scaffolds were used to rebuild the rabbit urethral defect. In group C, the urethral defect was reconstructed using a regular urethral reparation technique. The repair efficacy was compared among the three groups by examining the urethral morphology, tissue reconstruction, luminal patency, and complication incidence (including calculus formation, urinary fistula, and urethral stricture) using histological evaluation and urethral radiography methods. Findings from this study indicate that hAMSCs-loaded PLLA/PEG scaffolds resulted in the best urethral defect repair in rabbits, which predicts the promising application of a tissue engineering approach for urethral repair. PMID:27517902

  1. Human amniotic fluid stem cells labeled with up-conversion nanoparticles for imaging-monitored repairing of acute lung injury.

    PubMed

    Xu, Yunyun; Xiang, Jian; Zhao, He; Liang, Hansi; Huang, Jie; Li, Yan; Pan, Jian; Zhou, Huiting; Zhang, Xueguang; Wang, Jiang Huai; Liu, Zhuang; Wang, Jian

    2016-09-01

    Human amniotic fluid stem (hAFS) cells have generated a great deal of excitement in cell-based therapies and regenerative medicine. Here, we examined the effect of hAFS cells labeled with dual-polymer-coated UCNP-PEG-PEI nanoparticles in a murine model of acute lung injury (ALI). We observed hAFS cells migration to the lung using highly sensitive in vivo upconversion luminescence (UCL) imaging. We demonstrated that hAFS cells remained viable and retained their ability to differentiate even after UCNP-PEG-PEI labeling. More importantly, hAFS cells displayed remarkable positive effects on ALI-damaged lung tissue repair compared with mouse bone marrow mesenchymal stem cells (mBMSCs), which include recovery of the integrity of alveolar-capillary membrane, attenuation of transepithelial leukocyte and neutrophil migration, and down-regulation of proinflammatory cytokine and chemokine expression. Our work highlights a promising role for imaging-guided hAFS cell-based therapy in ALI. PMID:27244692

  2. Pathologic changes of wound tissue in rats with stage III pressure ulcers treated by transplantation of human amniotic epithelial cells.

    PubMed

    Zheng, Xilan; Jiang, Zhixia; Zhou, Aiting; Yu, Limei; Quan, Mingtao; Cheng, Huagang

    2015-01-01

    This study aims to determine the impact of orthotopic transplantation of human amniotic epithelial cells (hAECs) on the pathologic changes of wound tissues in a self-prepared rat stage III pressure ulcer model. Ninety-six SD rats were randomly divided into the model group (group M), hAEC transplantation group (group H), traditional treatment group (group T), and the control group (group C), with 24 rats in each group. The wound healing time was observed in 6 rats from each group, and 6 rats of each group were selected for post-modeling on day(s) (D) 1, 3, and 7 for HE staining to compare the pathological changes. The healing time of group H was significantly shorter than the other three groups. Moreover, pathological observations revealed that group H exhibited significant proliferation of fibrous tissues and vessels in the dermal layer, and the appearance time and degree of skin appendages were significantly greater than that observed in the other three groups. Pathological observations showed that hAEC transplantation could significantly speed up the healing of stage III pressure ulcer.

  3. Electrospun Poly(l-lactide)/Poly(ethylene glycol) Scaffolds Seeded with Human Amniotic Mesenchymal Stem Cells for Urethral Epithelium Repair.

    PubMed

    Lv, Xiaokui; Guo, Qianping; Han, Fengxuan; Chen, Chunyang; Ling, Christopher; Chen, Weiguo; Li, Bin

    2016-01-01

    Tissue engineering-based urethral replacement holds potential for repairing large segmental urethral defects, which remains a great challenge at present. This study aims to explore the potential of combining biodegradable poly(l-lactide) (PLLA)/poly(ethylene glycol) (PEG) scaffolds and human amniotic mesenchymal cells (hAMSCs) for repairing urethral defects. PLLA/PEG fibrous scaffolds with various PEG fractions were fabricated via electrospinning. The scaffolds were then seeded with hAMSCs prior to implantation in New Zealand male rabbits that had 2.0 cm-long defects in the urethras. The rabbits were randomly divided into three groups. In group A, hAMSCs were grown on PLLA/PEG scaffolds for two days and then implanted to the urethral defects. In group B, only the PLLA/PEG scaffolds were used to rebuild the rabbit urethral defect. In group C, the urethral defect was reconstructed using a regular urethral reparation technique. The repair efficacy was compared among the three groups by examining the urethral morphology, tissue reconstruction, luminal patency, and complication incidence (including calculus formation, urinary fistula, and urethral stricture) using histological evaluation and urethral radiography methods. Findings from this study indicate that hAMSCs-loaded PLLA/PEG scaffolds resulted in the best urethral defect repair in rabbits, which predicts the promising application of a tissue engineering approach for urethral repair.

  4. Electrospun Poly(l-lactide)/Poly(ethylene glycol) Scaffolds Seeded with Human Amniotic Mesenchymal Stem Cells for Urethral Epithelium Repair

    PubMed Central

    Lv, Xiaokui; Guo, Qianping; Han, Fengxuan; Chen, Chunyang; Ling, Christopher; Chen, Weiguo; Li, Bin

    2016-01-01

    Tissue engineering-based urethral replacement holds potential for repairing large segmental urethral defects, which remains a great challenge at present. This study aims to explore the potential of combining biodegradable poly(l-lactide) (PLLA)/poly(ethylene glycol) (PEG) scaffolds and human amniotic mesenchymal cells (hAMSCs) for repairing urethral defects. PLLA/PEG fibrous scaffolds with various PEG fractions were fabricated via electrospinning. The scaffolds were then seeded with hAMSCs prior to implantation in New Zealand male rabbits that had 2.0 cm-long defects in the urethras. The rabbits were randomly divided into three groups. In group A, hAMSCs were grown on PLLA/PEG scaffolds for two days and then implanted to the urethral defects. In group B, only the PLLA/PEG scaffolds were used to rebuild the rabbit urethral defect. In group C, the urethral defect was reconstructed using a regular urethral reparation technique. The repair efficacy was compared among the three groups by examining the urethral morphology, tissue reconstruction, luminal patency, and complication incidence (including calculus formation, urinary fistula, and urethral stricture) using histological evaluation and urethral radiography methods. Findings from this study indicate that hAMSCs-loaded PLLA/PEG scaffolds resulted in the best urethral defect repair in rabbits, which predicts the promising application of a tissue engineering approach for urethral repair. PMID:27517902

  5. Complication prevalence following use of tutoplast-derived human acellular dermal matrix in prosthetic breast reconstruction: a retrospective review of 203 patients.

    PubMed

    Rundell, V L M; Beck, R T; Wang, C E; Gutowski, K A; Sisco, M; Fenner, G; Howard, M A

    2014-10-01

    Use of human acellular dermal matrix (ADM) during prosthetic breast reconstruction has increased. Several ADM products are available produced by differing manufacturing techniques. It is not known if outcomes vary with different products. This study reports the complication prevalence following use of a tutoplast-derived ADM (T-ADM) in prosthetic breast reconstruction. We performed a retrospective chart review of 203 patients (mean follow-up times 12.2 months) who underwent mastectomy and immediate prosthetic breast reconstruction utilizing T-ADM, recording demographic data, surgical indications and complication (infection, seroma, hematoma, wound healing exceeding three weeks and reconstruction failure). During a four-year period, 348 breast reconstructions were performed Complications occurred in 16.4% of reconstructed breasts. Infection occurred in 6.6% of breast reconstructions (3.7% - major infection, requiring intravenous antibiotics and 2.9% minor infection, requiring oral antibiotics only). Seromas occurred in 3.4% and reconstruction failure occurred in 0.6% of breast reconstructions. Analysis suggested that complication prevalence was significantly higher in patients with a BMI >30 (p = 0.03). The complication profile following T-ADM use is this series is comparable to that reported for with other ADM products. T-ADM appears to be a safe and acceptable option for use in ADM-assisted breast reconstruction. PMID:24917371

  6. Effects of hydroxyapatite nanostructure on channel surface of porcine acellular dermal matrix scaffold on cell viability and osteogenic differentiation of human periodontal ligament stem cells.

    PubMed

    Ge, Shaohua; Zhao, Ning; Wang, Lu; Liu, Hong; Yang, Pishan

    2013-01-01

    A new nanostructured hydroxyapatite-coated porcine acellular dermal matrix (HAp-PADM) was fabricated by a biomimetic mineralization method. Human periodontal ligament stem cells were seeded on HAp-PADM and the effects of this scaffold on cell shape, cytoskeleton organization, cell viability, and osteogenic differentiation were examined. Periodontal ligament stem cells cultured on HAp-PADM exhibited different cell shape when compared with those on pure PADM. Moreover, HAp-PADM promoted cell viability and alkaline phosphatase activity significantly. Based on quantitative real-time polymerase chain reaction, the expression of bone-related markers runt-related transcription factor 2 (Runx2), osteopontin (OPN), and osteocalcin (OCN) upregulated in the HAp-PADM scaffold. The enhancement of osteogenic differentiation of periodontal ligament stem cells on the HAp-PADM scaffold was proposed based on the research results. The results of this study highlight the micro-nano, two-level, three-dimensional HAp-PADM composite as a promising scaffold for periodontal tissue engineering.

  7. Human amniotic fluid stem cells support undifferentiated propagation and pluripotency of human embryonic stem cell without b-FGF in a density dependent manner.

    PubMed

    Ma, Xiaorong; Li, Huanqi; Xin, Shujia; Ma, Yueting; Ouyang, Tianxiang

    2014-01-01

    Human embryonic stem cells (hESCs) are pluripotent cells which can give rise to almost all adult cell lineages. Culture system of hESCs is complex, requiring exogenous b-FGF and feeder cell layer. Human mesenchymal stem cells (MSCs) not only synthesize soluble cytokines or factors such as b-FGF, but also provide other mechanism which might play positive role on sustaining hESCs propagation and pluripotency. Human amniotic fluid stem (AFS) cells, which share characteristics of both embryonic and adult stem cells, have been regarded as promising cells for regenerative medicine. Taking advantage by AFS cells, we studied the ability of AFS cells in supporting undifferentiated propagation and pluripotency of Chinese population derived X-01 hESCs. Human AF-type amniotic fluid stem cells (hAF-AFSCs) transcribed genes including Activin A, TGF-β1, Noggin and b-FGF, which involved in maintaining pluripotency and self-renewal of hESCs. Compared to mouse embryonic fibroblasts (MEFs), hAF-AFSCs secreted higher concentration of b-FGF which was important in hESCs culture (P < 0.05). The hESCs were propagated more than 30 passages on hAF-AFSCs layer with exogenous b-FGF supplementation, keeping undifferentiated status. While exogenous b-FGF was obviated, propagation of hESCs with undifferentiated status was dependent on density of hAF-AFSC feeder layer. Lower density of hAF-AFSCs resulted in rapid decline in undifferentiated clone number, while higher ones hindered the growth of colonies. The most appropriate hAF-AFSCs feeder density to maintain the X-01 hESC line without exogenous b-FGF was 15-20×10(4)/well. To the best of our knowledge, this is the first study demonstrating that hAF-AFSCs could support undifferentiated propagation and pluripotency of Chinese population derived hESCs without exogenous b-FGF supplementation.

  8. Physics of amniote formation.

    PubMed

    Fleury, Vincent; Murukutla, Ameya Vaishnavi; Chevalier, Nicolas R; Gallois, Benjamin; Capellazzi-Resta, Marina; Picquet, Pierre; Peaucelle, Alexis

    2016-08-01

    We present a detailed study of the formation of the amniotic sac in the avian embryo, and a comparison with the crocodile amniotic sac. We show that the amniotic sac forms at a circular line of stiffness contrast, separating rings of cell domains. Cells align at this boundary, and this in turn orients and concentrates the tension forces. The tissue fold which forms the amniotic sac is locked exactly along this line due to the colocalization of the stiffness contrast and of the tensile force. In addition, the tensile force plays a regenerative role when the amniotic sac is cut. The fold forming the ventral side of the embryo displays the same characteristics. This work shows that amniote embryogenesis consists of a cascade of buckling events taking place at the boundaries between regions of differing mechanical properties. Hence, amniote embryogenesis relies on a simple and robust biomechanical scheme used repeatedly, and selected ancestrally.

  9. Physics of amniote formation.

    PubMed

    Fleury, Vincent; Murukutla, Ameya Vaishnavi; Chevalier, Nicolas R; Gallois, Benjamin; Capellazzi-Resta, Marina; Picquet, Pierre; Peaucelle, Alexis

    2016-08-01

    We present a detailed study of the formation of the amniotic sac in the avian embryo, and a comparison with the crocodile amniotic sac. We show that the amniotic sac forms at a circular line of stiffness contrast, separating rings of cell domains. Cells align at this boundary, and this in turn orients and concentrates the tension forces. The tissue fold which forms the amniotic sac is locked exactly along this line due to the colocalization of the stiffness contrast and of the tensile force. In addition, the tensile force plays a regenerative role when the amniotic sac is cut. The fold forming the ventral side of the embryo displays the same characteristics. This work shows that amniote embryogenesis consists of a cascade of buckling events taking place at the boundaries between regions of differing mechanical properties. Hence, amniote embryogenesis relies on a simple and robust biomechanical scheme used repeatedly, and selected ancestrally. PMID:27627351

  10. Physics of amniote formation

    NASA Astrophysics Data System (ADS)

    Fleury, Vincent; Murukutla, Ameya Vaishnavi; Chevalier, Nicolas R.; Gallois, Benjamin; Capellazzi-Resta, Marina; Picquet, Pierre; Peaucelle, Alexis

    2016-08-01

    We present a detailed study of the formation of the amniotic sac in the avian embryo, and a comparison with the crocodile amniotic sac. We show that the amniotic sac forms at a circular line of stiffness contrast, separating rings of cell domains. Cells align at this boundary, and this in turn orients and concentrates the tension forces. The tissue fold which forms the amniotic sac is locked exactly along this line due to the colocalization of the stiffness contrast and of the tensile force. In addition, the tensile force plays a regenerative role when the amniotic sac is cut. The fold forming the ventral side of the embryo displays the same characteristics. This work shows that amniote embryogenesis consists of a cascade of buckling events taking place at the boundaries between regions of differing mechanical properties. Hence, amniote embryogenesis relies on a simple and robust biomechanical scheme used repeatedly, and selected ancestrally.

  11. Transplantation of human umbilical cord blood or amniotic epithelial stem cells alleviates mechanical allodynia after spinal cord injury in rats.

    PubMed

    Roh, Dae-Hyun; Seo, Min-Soo; Choi, Hoon-Seong; Park, Sang-Bum; Han, Ho-Jae; Beitz, Alvin J; Kang, Kyung-Sun; Lee, Jang-Hern

    2013-01-01

    Stem cell therapy is a potential treatment for spinal cord injury (SCI), and a variety of different stem cell types have been grafted into humans suffering from spinal cord trauma or into animal models of spinal injury. Although several studies have reported functional motor improvement after transplantation of stem cells into injured spinal cord, the benefit of these cells for treating SCI-induced neuropathic pain is not clear. In this study, we investigated the therapeutic effect of transplanting human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) or amniotic epithelial stem cells (hAESCs) on SCI-induced mechanical allodynia (MA) and thermal hyperalgesia (TH) in T13 spinal cord hemisected rats. Two weeks after SCI, hUCB-MSCs or hAESCs were transplanted around the spinal cord lesion site, and behavioral tests were performed to evaluate changes in SCI-induced MA and TH. Immunohistochemical and Western blot analyses were also performed to evaluate possible therapeutic effects on SCI-induced inflammation and the nociceptive-related phosphorylation of the NMDA NR1 receptor subunit. While transplantation of hUCB-MSCs showed a tendency to reduce MA, transplantation of hAESCs significantly reduced MA. Neither hUCB-MSC nor hAESC transplantation had any effect on SCI-induced TH. Transplantation of hAESCs also significantly reduced the SCI-induced increase in NMDA receptor NR1 subunit phosphorylation (pNR1) expression in the spinal cord. Both hUCB-MSCs and hAESCs reduced the SCI-induced increase in spinal cord expression of the microglial marker, F4/80, but not the increased expression of GFAP or iNOS. Taken together, these findings demonstrate that the transplantation of hAESCs into the injured spinal cord can suppress mechanical allodynia, and this effect seems to be closely associated with the modulation of spinal cord microglia activity and NR1 phosphorylation.

  12. Comparison of toxicities of acellular pertussis vaccine with whole cell pertussis vaccine in experimental animals.

    PubMed

    Sato, Y; Sato, H

    1991-01-01

    There is no suitable animal model for pertussis encephalopathy in humans. In this study, we have compared the toxicity of acellular pertussis vaccine with whole cell pertussis vaccine in mice or guinea pigs. Two lots of acellular and two lots of whole cell vaccine produced in different countries were assayed in the test. 1. There was no statistical difference in mouse protective potency between these acellular or whole cell pertussis vaccines. 2. There were no differences in chemical ingredients between acellular and whole cell pertussis vaccines except for protein nitrogen content. The protein nitrogen content of whole cell vaccine was at least three times higher than that of the acellular product. 3. Anti-PT antibody productivity of the acellular vaccine was higher than that of the whole cell vaccine. 4. Anti-agglutinogen antibody productivity of the whole cell vaccine was higher than that of the acellular vaccine. 5. There was no pyrogenic activity with the acellular vaccine, but high pyrogenicity was seen with whole cell vaccine. 6. There was high body-weight decreasing toxicity in mice and guinea pigs by the whole cell vaccine. 7. The mice died when they received whole cell pertussis vaccine iv, but no deaths occurred in the mice which received acellular pertussis vaccine. PMID:1778317

  13. Investigation of Efficacy of Mitomycin-C, Sodium Hyaluronate and Human Amniotic Fluid in Preventing Epidural Fibrosis and Adhesion Using a Rat Laminectomy Model

    PubMed Central

    Bolat, Elif; Kocamaz, Erdoğan; Kulahcilar, Zeki; Yilmaz, Ali; Topcu, Abdullah; Coskun, Mehmet Erdal

    2013-01-01

    Study Design A retrospective study. Purpose The aim of this study was to evalute the effects of mitomycin-C, sodium hyaluronate and human amniotic fluid on preventing spinal epidural fibrosis. Overview of Literature The role of scar tissue in pain formation is not exactly known, but it is reported that scar tissue causes adhesions between anatomic structures. Intensive fibrotic tissue compresses on anatomic structures and increases the sensitivity of the nerve root for recurrent herniation and lateral spinal stenosis via limiting movements of the root. Also, neuronal atrophy and axonal degeneration occur under scar tissue. Methods The study design included 4 groups of rats: group 1 was the control group, groups 2, 3, and 4 receieved antifibrotic agents, mitomycin-C (group 2), sodium hyaluronate (group 3), and human amniotic fluid (group 4). Midline incision for all animals were done on L5 for total laminectomy. Four weeks after the surgery, the rats were sacrificed and specimens were stained with hematoxylin-eosin and photos of the slides were taken for quantitive assesment of the scar tissue. Results There was no significant scar tissue in the experimental animals of groups 2, 3, and 4. It was found that there was no significant difference between drug groups, but there was a statistically significant difference between the drug groups and the control group. Conclusions This experimental study shows that implantation of mitomycin-C, sodium hyaluronate and human amniotic fluid reduces epidural fibrosis and adhesions after spinal laminectomy in rat models. Further studies in humans are needed to determine the complications of the agents researched. PMID:24353840

  14. BMP15 gene is activated during human amniotic fluid stem cell differentiation into oocyte-like cells.

    PubMed

    Cheng, Xiang; Chen, Shuai; Yu, Xiaoli; Zheng, Pengsheng; Wang, Huayan

    2012-07-01

    The generation of oocyte-like cells (OLCs) from stem cell differentiation in vitro provides an optimal approach for studying the mechanism of oocyte development and maturation. The aim of this study was to investigate the activation of bone morphogenetic protein 15 gene (BMP15) during the differentiation of human amniotic fluid stem cells (hAFSCs) into OLCs. After 15 days of differentiation, OLCs with a diameter of 50-60 μm and zona pellucida (ZP)-like morphology were observed. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed the BMP15 was activated from approximately day 10 of differentiating hAFSCs and thereafter. The reporter construct pBMP15-enhanced green fluorescent protein (EGFP) was transiently transfected into the differentiated hAFSCs and the EGFP expression driven by the BMP15 promoter was positive in the OLCs. Moreover, RT-PCR analysis showed that the oocyte-specific markers including ZP1, ZP2, ZP3, and c-kit were expressed in the differentiated hAFSCs, and the immunofluorescence assay confirmed that the ZP2 was detected in the OLCs. Quantitative RT-PCR revealed that ZP2 and ZP3 were significantly elevated in the differentiated hAFSCs. Further, in the OLCs derived from hAFSCs, the BMP15 promoter directing the EGFP reporter was colocalized with ZP2. Together, these results illustrated that the BMP15 could be used as an oogenesis marker to track hAFSCs differentiation into the OLCs.

  15. Human amniotic mesenchymal stem cell-derived induced pluripotent stem cells may generate a universal source of cardiac cells.

    PubMed

    Ge, Xiaohu; Wang, I-Ning E; Toma, Ildiko; Sebastiano, Vittorio; Liu, Jianwei; Butte, Manish J; Reijo Pera, Renee A; Yang, Phillip C

    2012-10-10

    Human amniotic mesenchymal stem cells (hAMSCs) demonstrated partially pluripotent characteristics with a strong expression of Oct4 and Nanog genes and immunomodulatory properties characterized by the absence of HLA-DR and the presence of HLA-G and CD59. The hAMSCs were reprogrammed into induced pluripotent stem cells (iPSCs) that generate a promising source of universal cardiac cells. The hAMSC-derived iPSCs (MiPSCs) successfully underwent robust cardiac differentiation to generate cardiomyocytes. This study investigated 3 key properties of the hAMSCs and MiPSCs: (1) the reprogramming efficiency of the partially pluripotent hAMSCs to generate MiPSCs; (2) immunomodulatory properties of the hAMSCs and MiPSCs; and (3) the cardiac differentiation potential of the MiPSCs. The characteristic iPSC colony formation was observed within 10 days after the transduction of the hAMSCs with a single integration polycistronic vector containing 4 Yamanaka factors. Immunohistology and reverse transcription-polymerase chain reaction assays revealed that the MiPSCs expressed stem cell surface markers and pluripotency-specific genes. Furthermore, the hAMSCs and MiPSCs demonstrated immunomodulatory properties enabling successful engraftment in the SVJ mice. Finally, the cardiac differentiation of MiPSCs exhibited robust spontaneous contractility, characteristic calcium transience across the membrane, a high expression of cardiac genes and mature cardiac phenotypes, and a contractile force comparable to cardiomyocytes. Our results demonstrated that the hAMSCs are reprogrammed with a high efficiency into MiPSCs, which possess pluripotent, immunomodulatory, and precardiac properties. The MiPSC-derived cardiac cells express a c-kit cell surface marker, which may be employed to purify the cardiac cell population and enable allogeneic cardiac stem cell therapy.

  16. Improved Neurological Outcome by Intramuscular Injection of Human Amniotic Fluid Derived Stem Cells in a Muscle Denervation Model

    PubMed Central

    Su, Hong-Lin; Sheu, Meei-Ling; Lu, Zong-Han; Chiang, Chien-Yi; Yang, Dar-Yu; Sheehan, Jason; Pan, Hung-Chuan

    2015-01-01

    Purpose The skeletal muscle develops various degrees of atrophy and metabolic dysfunction following nerve injury. Neurotrophic factors are essential for muscle regeneration. Human amniotic fluid derived stem cells (AFS) have the potential to secrete various neurotrophic factors necessary for nerve regeneration. In the present study, we assess the outcome of neurological function by intramuscular injection of AFS in a muscle denervation and nerve anastomosis model. Materials and Methods Seventy two Sprague-Dawley rats weighing 200–250 gm were enrolled in this study. Muscle denervation model was conducted by transverse resection of a sciatic nerve with the proximal end sutured into the gluteal muscle. The nerve anastomosis model was performed by transverse resection of the sciatic nerve followed by four stitches reconnection. These animals were allocated to three groups: control, electrical muscle stimulation, and AFS groups. Results NT-3 (Neurotrophin 3), BDNF (Brain derived neurotrophic factor), CNTF (Ciliary neurotrophic factor), and GDNF (Glia cell line derived neurotrophic factor) were highly expressed in AFS cells and supernatant of culture medium. Intra-muscular injection of AFS exerted significant expression of several neurotrophic factors over the distal end of nerve and denervated muscle. AFS caused high expression of Bcl-2 in denervated muscle with a reciprocal decrease of Bad and Bax. AFS preserved the muscle morphology with high expression of desmin and acetylcholine receptors. Up to two months, AFS produced significant improvement in electrophysiological study and neurological functions such as SFI (sciatic nerve function index) and Catwalk gait analysis. There was also significant preservation of the number of anterior horn cells and increased nerve myelination as well as muscle morphology. Conclusion Intramuscular injection of AFS can protect muscle apoptosis and likely does so through the secretion of various neurotrophic factors. This protection

  17. Porcine acellular lung matrix for wound healing and abdominal wall reconstruction: A pilot study.

    PubMed

    Fernandez-Moure, Joseph S; Van Eps, Jeffrey L; Rhudy, Jessica R; Cabrera, Fernando J; Acharya, Ghanashyam S; Tasciotti, Ennio; Sakamoto, Jason; Nichols, Joan E

    2016-01-01

    Surgical wound healing applications require bioprosthetics that promote cellular infiltration and vessel formation, metrics associated with increased mechanical strength and resistance to infection. Porcine acellular lung matrix is a novel tissue scaffold known to promote cell adherence while minimizing inflammatory reactions. In this study, we evaluate the capacity of porcine acellular lung matrix to sustain cellularization and neovascularization in a rat model of subcutaneous implantation and chronic hernia repair. We hypothesize that, compared to human acellular dermal matrix, porcine acellular lung matrix would promote greater cell infiltration and vessel formation. Following pneumonectomy, porcine lungs were processed and characterized histologically and by scanning electron microscopy to demonstrate efficacy of the decellularization. Using a rat model of subcutaneou implantation, porcine acellular lung matrices (n = 8) and human acellular dermal matrices (n = 8) were incubated in vivo for 6 weeks. To evaluate performance under mechanically stressed conditions, porcine acellular lung matrices (n = 7) and human acellular dermal matrices (n = 7) were implanted in a rat model of chronic ventral incisional hernia repair for 6 weeks. After 6 weeks, tissues were evaluated using hematoxylin and eosin and Masson's trichrome staining to quantify cell infiltration and vessel formation. Porcine acellular lung matrices were shown to be successfully decellularized. Following subcutaneous implantation, macroscopic vessel formation was evident. Porcine acellular lung matrices demonstrated sufficient incorporation and showed no evidence of mechanical failure after ventral hernia repair. Porcine acellular lung matrices demonstrated significantly greater cellular density and vessel formation when compared to human acellular dermal matrix. Vessel sizes were similar across all groups. Cell infiltration and vessel formation are well-characterized metrics of incorporation

  18. Porcine acellular lung matrix for wound healing and abdominal wall reconstruction: A pilot study

    PubMed Central

    Fernandez-Moure, Joseph S; Van Eps, Jeffrey L; Rhudy, Jessica R; Cabrera, Fernando J; Acharya, Ghanashyam S; Tasciotti, Ennio; Sakamoto, Jason; Nichols, Joan E

    2016-01-01

    Surgical wound healing applications require bioprosthetics that promote cellular infiltration and vessel formation, metrics associated with increased mechanical strength and resistance to infection. Porcine acellular lung matrix is a novel tissue scaffold known to promote cell adherence while minimizing inflammatory reactions. In this study, we evaluate the capacity of porcine acellular lung matrix to sustain cellularization and neovascularization in a rat model of subcutaneous implantation and chronic hernia repair. We hypothesize that, compared to human acellular dermal matrix, porcine acellular lung matrix would promote greater cell infiltration and vessel formation. Following pneumonectomy, porcine lungs were processed and characterized histologically and by scanning electron microscopy to demonstrate efficacy of the decellularization. Using a rat model of subcutaneou implantation, porcine acellular lung matrices (n = 8) and human acellular dermal matrices (n = 8) were incubated in vivo for 6 weeks. To evaluate performance under mechanically stressed conditions, porcine acellular lung matrices (n = 7) and human acellular dermal matrices (n = 7) were implanted in a rat model of chronic ventral incisional hernia repair for 6 weeks. After 6 weeks, tissues were evaluated using hematoxylin and eosin and Masson’s trichrome staining to quantify cell infiltration and vessel formation. Porcine acellular lung matrices were shown to be successfully decellularized. Following subcutaneous implantation, macroscopic vessel formation was evident. Porcine acellular lung matrices demonstrated sufficient incorporation and showed no evidence of mechanical failure after ventral hernia repair. Porcine acellular lung matrices demonstrated significantly greater cellular density and vessel formation when compared to human acellular dermal matrix. Vessel sizes were similar across all groups. Cell infiltration and vessel formation are well-characterized metrics of incorporation

  19. Human gut colonisation may be initiated in utero by distinct microbial communities in the placenta and amniotic fluid.

    PubMed

    Collado, Maria Carmen; Rautava, Samuli; Aakko, Juhani; Isolauri, Erika; Salminen, Seppo

    2016-03-22

    Interaction with intestinal microbes in infancy has a profound impact on health and disease in later life through programming of immune and metabolic pathways. We collected maternal faeces, placenta, amniotic fluid, colostrum, meconium and infant faeces samples from 15 mother-infant pairs in an effort to rigorously investigate prenatal and neonatal microbial transfer and gut colonisation. To ensure sterile sampling, only deliveries at full term by elective caesarean section were studied. Microbiota composition and activity assessment by conventional bacterial culture, 16S rRNA gene pyrosequencing, quantitative PCR, and denaturing gradient gel electrophoresis revealed that the placenta and amniotic fluid harbour a distinct microbiota characterised by low richness, low diversity and the predominance of Proteobacteria. Shared features between the microbiota detected in the placenta and amniotic fluid and in infant meconium suggest microbial transfer at the foeto-maternal interface. At the age of 3-4 days, the infant gut microbiota composition begins to resemble that detected in colostrum. Based on these data, we propose that the stepwise microbial gut colonisation process may be initiated already prenatally by a distinct microbiota in the placenta and amniotic fluid. The link between the mother and the offspring is continued after birth by microbes present in breast milk.

  20. Human gut colonisation may be initiated in utero by distinct microbial communities in the placenta and amniotic fluid.

    PubMed

    Collado, Maria Carmen; Rautava, Samuli; Aakko, Juhani; Isolauri, Erika; Salminen, Seppo

    2016-01-01

    Interaction with intestinal microbes in infancy has a profound impact on health and disease in later life through programming of immune and metabolic pathways. We collected maternal faeces, placenta, amniotic fluid, colostrum, meconium and infant faeces samples from 15 mother-infant pairs in an effort to rigorously investigate prenatal and neonatal microbial transfer and gut colonisation. To ensure sterile sampling, only deliveries at full term by elective caesarean section were studied. Microbiota composition and activity assessment by conventional bacterial culture, 16S rRNA gene pyrosequencing, quantitative PCR, and denaturing gradient gel electrophoresis revealed that the placenta and amniotic fluid harbour a distinct microbiota characterised by low richness, low diversity and the predominance of Proteobacteria. Shared features between the microbiota detected in the placenta and amniotic fluid and in infant meconium suggest microbial transfer at the foeto-maternal interface. At the age of 3-4 days, the infant gut microbiota composition begins to resemble that detected in colostrum. Based on these data, we propose that the stepwise microbial gut colonisation process may be initiated already prenatally by a distinct microbiota in the placenta and amniotic fluid. The link between the mother and the offspring is continued after birth by microbes present in breast milk. PMID:27001291

  1. Human gut colonisation may be initiated in utero by distinct microbial communities in the placenta and amniotic fluid

    PubMed Central

    Collado, Maria Carmen; Rautava, Samuli; Aakko, Juhani; Isolauri, Erika; Salminen, Seppo

    2016-01-01

    Interaction with intestinal microbes in infancy has a profound impact on health and disease in later life through programming of immune and metabolic pathways. We collected maternal faeces, placenta, amniotic fluid, colostrum, meconium and infant faeces samples from 15 mother-infant pairs in an effort to rigorously investigate prenatal and neonatal microbial transfer and gut colonisation. To ensure sterile sampling, only deliveries at full term by elective caesarean section were studied. Microbiota composition and activity assessment by conventional bacterial culture, 16S rRNA gene pyrosequencing, quantitative PCR, and denaturing gradient gel electrophoresis revealed that the placenta and amniotic fluid harbour a distinct microbiota characterised by low richness, low diversity and the predominance of Proteobacteria. Shared features between the microbiota detected in the placenta and amniotic fluid and in infant meconium suggest microbial transfer at the foeto-maternal interface. At the age of 3–4 days, the infant gut microbiota composition begins to resemble that detected in colostrum. Based on these data, we propose that the stepwise microbial gut colonisation process may be initiated already prenatally by a distinct microbiota in the placenta and amniotic fluid. The link between the mother and the offspring is continued after birth by microbes present in breast milk. PMID:27001291

  2. Therapeutic potential of amniotic fluid stem cells.

    PubMed

    Abdulrazzak, Hassan; De Coppi, Paolo; Guillot, Pascale V

    2013-03-01

    Human amniotic fluid cells have been used traditionally as a diagnostic tool for genetic anomalies. More recently it has been recognized that amniotic fluid contains populations of stem cells. Mesenchymal stem cells (AFMSC) were first to be described. These cells are able to differentiate towards mesodermal lineages. More recently cells with broader potential, defined as amniotic fluid stem cells (AFSC), were also isolated. They have intermediate characteristics between embryonic and adult stem cells and are able to differentiate into lineages representative of all three germ layers but unlike ES cells they do not form tumours in vivo. Furthermore, AFSC have been reverted to functional pluripotency in a transgene-free approach using an epigenetics modifier. These characteristics, together with absence of ethical issues concerning their employment, have made stem cells from amniotic fluid a promising candidate for cell therapy and tissue engineering.

  3. Editorial Commentary: Reflections From a Mature Arthroscopic Shoulder Surgeon on the History and Current Benefits of Augmentation for the Revision of a Massive Rotator Cuff Tear Using Acellular Human Dermal Matrix Allograft.

    PubMed

    Snyder, Stephen J

    2016-09-01

    Acellular human dermal matrix allografts are now being used to augment and sometimes replace severely damaged rotator cuff tissue. I have been interested in this important aspect of orthopaedics for 15 years and am pleased to have the opportunity to share my personal reflections of some of the highlights in science and the literature that helped get to the point now where we can expect greater than 80% healing even in these difficult cases of revision after massive failed cuff repair. The field of tissue engineering will certainly be a critical part of our rotator cuff surgical future.

  4. Editorial Commentary: Reflections From a Mature Arthroscopic Shoulder Surgeon on the History and Current Benefits of Augmentation for the Revision of a Massive Rotator Cuff Tear Using Acellular Human Dermal Matrix Allograft.

    PubMed

    Snyder, Stephen J

    2016-09-01

    Acellular human dermal matrix allografts are now being used to augment and sometimes replace severely damaged rotator cuff tissue. I have been interested in this important aspect of orthopaedics for 15 years and am pleased to have the opportunity to share my personal reflections of some of the highlights in science and the literature that helped get to the point now where we can expect greater than 80% healing even in these difficult cases of revision after massive failed cuff repair. The field of tissue engineering will certainly be a critical part of our rotator cuff surgical future. PMID:27594327

  5. In situ vascularization of injectable fibrin/poly(ethylene glycol) hydrogels by human amniotic fluid-derived stem cells.

    PubMed

    Benavides, Omar M; Brooks, Abigail R; Cho, Sung Kyung; Petsche Connell, Jennifer; Ruano, Rodrigo; Jacot, Jeffrey G

    2015-08-01

    One of the greatest challenges in regenerative medicine is generating clinically relevant engineered tissues with functional blood vessels. Vascularization is a key hurdle faced in designing tissue constructs larger than the in vivo limit of oxygen diffusion. In this study, we utilized fibrin-based hydrogels to serve as a foundation for vascular formation, poly(ethylene glycol) (PEG) to modify fibrinogen and increase scaffold longevity, and human amniotic fluid-derived stem cells (AFSC) as a source of vascular cell types (AFSC-EC). AFSC hold great potential for use in regenerative medicine strategies, especially those involving autologous congenital applications, and we have shown previously that AFSC-seeded fibrin-PEG hydrogels have the potential to form three-dimensional vascular-like networks in vitro. We hypothesized that subcutaneously injecting these hydrogels in immunodeficient mice would both induce a fibrin-driven angiogenic host response and promote in situ AFSC-derived neovascularization. Two weeks postinjection, hydrogels were sectioned, and the following was demonstrated: the average maximum invasion distance of host murine cells into the subcutaneous fibrin/PEG scaffold was 147 ± 90 µm after 1 week and 395 ± 138 µm after 2 weeks; the average number of cell-lined lumen per square millimeter was significantly higher in hydrogels seeded with stem cells or cocultures containing stem cells (MSC, 36.5 ± 11.4; AFSC, 47.0 ± 18.9; AFSC/AFSC-EC, 32.8 ± 11.6; and MSC/HUVEC, 43.1 ± 25.1) versus endothelial cell types alone (AFSC-EC, 9.7 ± 6.1; HUVEC, 14.2 ± 8.8); and a subset of these lumen were characterized by the presence of red blood cells. Select areas of cell-seeded hydrogels contained CD31(+) lumen surrounded by α-smooth muscle cell support cells, whereas control hydrogels with no cells only showed infiltration of α-smooth muscle cell-positive host cells.

  6. Capillary-Like Network Formation by Human Amniotic Fluid-Derived Stem Cells Within Fibrin/Poly(Ethylene Glycol) Hydrogels

    PubMed Central

    Benavides, Omar M.; Quinn, Joseph P.; Pok, Seokwon; Petsche Connell, Jennifer; Ruano, Rodrigo

    2015-01-01

    A major limitation in tissue engineering strategies for congenital birth defects is the inability to provide a significant source of oxygen, nutrient, and waste transport in an avascular scaffold. Successful vascularization requires a reliable method to generate vascular cells and a scaffold capable of supporting vessel formation. The broad potential for differentiation, high proliferation rates, and autologous availability for neonatal surgeries make amniotic fluid-derived stem cells (AFSC) well suited for regenerative medicine strategies. AFSC-derived endothelial cells (AFSC-EC) express key proteins and functional phenotypes associated with endothelial cells. Fibrin-based hydrogels were shown to stimulate AFSC-derived network formation in vitro but were limited by rapid degradation. Incorporation of poly(ethylene glycol) (PEG) provided mechanical stability (65%±9% weight retention vs. 0% for fibrin-only at day 14) while retaining key benefits of fibrin-based scaffolds—quick formation (10±3 s), biocompatibility (88%±5% viability), and vasculogenic stimulation. To determine the feasibility of AFSC-derived microvasculature, we compared AFSC-EC as a vascular cell source and AFSC as a perivascular cell source to established sources of these cell types—human umbilical vein endothelial cells (HUVEC) and mesenchymal stem cells (MSC), respectively. Cocultures were seeded at a 4:1 endothelial-to-perivascular cell ratio, and gels were incubated at 37°C for 2 weeks. Mechanical testing was performed using a stress-controlled rheometer (G′=95±10 Pa), and cell-seeded hydrogels were assessed based on morphology. Network formation was analyzed based on key parameters such as vessel thickness, length, and area, as well as the degree of branching. There was no statistical difference between individual cultures of AFSC-EC and HUVEC in regard to these parameters, suggesting the vasculogenic potential of AFSC-EC; however, the development of robust vessels required the

  7. Challenges in validating the sterilisation dose for processed human amniotic membranes

    NASA Astrophysics Data System (ADS)

    Yusof, Norimah; Hassan, Asnah; Firdaus Abd Rahman, M. N.; Hamid, Suzina A.

    2007-11-01

    Most of the tissue banks in the Asia Pacific region have been using ionising radiation at 25 kGy to sterilise human tissues for save clinical usage. Under tissue banking quality system, any dose employed for sterilisation has to be validated and the validation exercise has to be a part of quality document. Tissue grafts, unlike medical items, are not produced in large number per each processing batch and tissues relatively have a different microbial population. A Code of Practice established by the International Atomic Energy Agency (IAEA) in 2004 offers several validation methods using smaller number of samples compared to ISO 11137 (1995), which is meant for medical products. The methods emphasise on bioburden determination, followed by sterility test on samples after they were exposed to verification dose for attaining of sterility assurance level (SAL) of 10 -1. This paper describes our experience in using the IAEA Code of Practice in conducting the validation exercise for substantiating 25 kGy as sterilisation dose for both air-dried amnion and those preserved in 99% glycerol.

  8. Clone-derived human AF-amniotic fluid stem cells are capable of skeletal myogenic differentiation in vitro and in vivo.

    PubMed

    Ma, Xiaorong; Zhang, Shengli; Zhou, Junmei; Chen, Baisong; Shang, Yafeng; Gao, Tongbing; Wang, Xue; Xie, Hua; Chen, Fang

    2012-08-01

    Stem cell-based therapy may be the most promising method to cure skeletal muscle degenerative diseases such as Duchenne muscular dystrophy (DMD) and trauma in the future. Human amniotic fluid is enriched with early-stage stem cells from developing fetuses and these cells have cardiomyogenic potential both in vitro and in vivo. In the present study, we investigated the characteristics of human amniotic fluid-derived AF-type stem (HAF-AFS) cells by flow cytometry, immunofluorescence staining, reverse-transcription polymerase chain reaction, and osteogenic and adipogenic differentiation analysis. After confirming the stemness of HAF-AFS cells, we tested whether HAF-AFS cells could differentiate into skeletal myogenic cells in vitro and incorporate into regenerating skeletal muscle in vivo. By temporary exposure to the DNA demethylation agent 5-aza-2'-deoxycytidine (5-Aza dC) or co-cultured with C2C12 myoblasts, HAF-AFS cells differentiated into skeletal myogenic cells, expressing skeletal myogenic cell-specific markers such as Desmin, Troponin I (Tn I) and α-Actinin. Four weeks after transplantation into cardiotoxin-injured and X-ray-irradiated tibialis anterior (TA) muscles of NOD/SCID mice, HAF-AFS cells survived, differentiated into myogenic precursor cells and fused with host myofibres. The findings that HAF-AFS cells differentiate into myogenic cells in vitro and incorporate in skeletal muscle regeneration in vivo hold the promise of HAF-AFS cell-based therapy for skeletal muscle degenerative diseases.

  9. Cut, copy, move, delete: The study of human interferon genes reveal multiple mechanisms underlying their evolution in amniotes.

    PubMed

    Krause, Christopher D; Pestka, Sidney

    2015-12-01

    Interferons (IFNs) are rapidly evolving cytokines released when viral infections are detected in cells. Previous research suggests that genes encoding IFNs and their receptors duplicated extensively throughout vertebrate evolution. We present molecular genetic evidence that supports the use of nonallelic homologous recombination (NAHR) to expand select IFN genes during amniote evolution. The duplication of long regions of genome (encompassing at least one functional IFN gene) followed by the insertion of this genome fragment near its parent's location, is commonly observed in many amniote genomes. Duplicates inserted away from duplication hotspots are not as frequently perturbed with new duplicates, and tend to survive long periods of evolution, sometimes becoming new IFN subtypes. Although most duplicates are inserted parallel to and near the original sequence, the insertion of the Kelch-like 9 gene within the Type I IFN locus of placental mammals promoted antiparallel insertion of gene duplicates between the Kelch-like 9 and IFN-ε loci. Genetic exchange between highly similar Type I gene duplicates as well as between Type III IFN gene duplicates homogenized their diversification. Oddly, Type III IFN genes migrated long distances throughout the genome more frequently than did Type I IFN genes. The inter-chromosomal movement of Type I IFN genes in amniotes correlated with complete intron loss in their gene structure, and repeatedly occurred with occasional Type III IFN genes.

  10. Human amniotic epithelial cell feeder layers maintain human iPS cell pluripotency via inhibited endogenous microRNA-145 and increased Sox2 expression

    SciTech Connect

    Liu, Te; Cheng, Weiwei; Huang, Yongyi; Huang, Qin; Jiang, Lizhen; Guo, Lihe

    2012-02-15

    Currently, human induced pluripotent stem (iPS) cells were generated from patient or disease-specific sources and share the same key properties as embryonic stem cells. This makes them attractive for personalized medicine, drug screens or cellular therapy. Long-term cultivation and maintenance of normal iPS cells in an undifferentiated self-renewing state are a major challenge. Our previous studies have shown that human amniotic epithelial cells (HuAECs) could provide a good source of feeder cells for mouse and human embryonic stem cells, or spermatogonial stem cells, but the mechanism for this is unknown. Here, we examined the effect of endogenous microRNA-145 regulation on Sox2 expression in human iPS cells by HuAECs feeder cells regulation, and in turn on human iPS cells pluripotency. We found that human IPS cells transfected with a microRNA-145 mutant expressed Sox2 at high levels, allowing iPS to maintain a high level of AP activity in long-term culture and form teratomas in SCID mice. Expression of stem cell markers was increased in iPS transfected with the microRNA-145 mutant, compared with iPS was transfected with microRNA-145. Besides, the expression of Drosha proteins of the microRNA-processor complex, required for the generation of precursor pre-miRNA, was significantly increased in human iPS cells cultured on MEF but not on HuAECs. Taken together, these results suggest that endogenous Sox2 expression may be regulated by microRNA-145 in human iPS cells with HuAECs feeder cells, and Sox2 is a crucial component required for maintenance of them in an undifferentiated, proliferative state capable of self-renewal. Highlights: Black-Right-Pointing-Pointer microRNA-145 inhibits Sox2 expression in human iPS cells. Black-Right-Pointing-Pointer microRNA-145 suppresses the self-renewal and pluripotency of human iPS cells. Black-Right-Pointing-Pointer HuAECs regulate expression of microRNA-145 and Sox2 in human iPS cells. Black-Right-Pointing-Pointer HuAECs feeder

  11. Conditioned medium from human amniotic epithelial cells may induce the differentiation of human umbilical cord blood mesenchymal stem cells into dopaminergic neuron-like cells.

    PubMed

    Yang, Shu; Sun, Hai-Mei; Yan, Ji-Hong; Xue, Hong; Wu, Bo; Dong, Fang; Li, Wen-Shuai; Ji, Feng-Qing; Zhou, De-Shan

    2013-07-01

    Dopaminergic (DA) neuron therapy has been established as a new clinical tool for treating Parkinson's disease (PD). Prior to cell transplantation, there are two primary issues that must be resolved: one is the appropriate seed cell origin, and the other is the efficient inducing technique. In the present study, human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) were used as the available seed cells, and conditioned medium from human amniotic epithelial cells (ACM) was used as the inducing reagent. Results showed that the proportion of DA neuron-like cells from hUCB-MSCs was significantly increased after cultured in ACM, suggested by the upregulation of DAT, TH, Nurr1, and Pitx3. To identify the process by which ACM induces DA neuron differentiation, we pretreated hUCB-MSCs with k252a, the Trk receptor inhibitor of brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF), and found that the proportion of DA neuron-like cells was significantly decreased compared with ACM-treated hUCB-MSCs, suggesting that NGF and BDNF in ACM were involved in the differentiation process. However, we could not rule out the involvement of other unidentified factors in the ACM, because ACM + k252a treatment does not fully block DA neuron-like cell differentiation compared with control. The transplantation of ACM-induced hUCB-MSCs could ameliorate behavioral deficits in PD rats, which may be associated with the survival of engrafted DA neuron-like cells. In conclusion, we propose that hUCB-MSCs are a good source of DA neuron-like cells and that ACM is a potential inducer to obtain DA neuron-like cells from hUCB-MSCs in vitro for an ethical and legal cell therapy for PD.

  12. Comparative study of effects of magnesium and taurine on electrical parameters of natural and artificial membranes. VIII. Effect on the ultrastructure of human amniotic epithelial cells.

    PubMed

    Guiet-Bara, A; Bara, M; Durlach, J

    1991-03-01

    The ultrastructure of human amniotic epithelial cells from normal pregnancies, at term, was studied using transmission electron microscopy. The results were analysed by a stereological method which indicates the ratio between the volume of the intercellular space (R1, the microvilli (R2), and the podocytes (R3) versus the cell volume. At low concentration (2 mM), MgCl2 decreased R1 and R3 and had no significant effect on R2. In contrast, taurine (2 mM) increased R1 and had no significant effect on R2 and R3. There is no vicarious action between Mg and taurine. These data are in contrast to the results obtained after electrophysiological studies, which indicates that the structural targets for Mg and taurine are different from the targets responsible for ionic transfer.

  13. Acellular organ scaffolds for tumor tissue engineering

    NASA Astrophysics Data System (ADS)

    Guller, Anna; Trusova, Inna; Petersen, Elena; Shekhter, Anatoly; Kurkov, Alexander; Qian, Yi; Zvyagin, Andrei

    2015-12-01

    Rationale: Tissue engineering (TE) is an emerging alternative approach to create models of human malignant tumors for experimental oncology, personalized medicine and drug discovery studies. Being the bottom-up strategy, TE provides an opportunity to control and explore the role of every component of the model system, including cellular populations, supportive scaffolds and signalling molecules. Objectives: As an initial step to create a new ex vivo TE model of cancer, we optimized protocols to obtain organ-specific acellular matrices and evaluated their potential as TE scaffolds for culture of normal and tumor cells. Methods and results: Effective decellularization of animals' kidneys, ureter, lungs, heart, and liver has been achieved by detergent-based processing. The obtained scaffolds demonstrated biocompatibility and growthsupporting potential in combination with normal (Vero, MDCK) and tumor cell lines (C26, B16). Acellular scaffolds and TE constructs have been characterized and compared with morphological methods. Conclusions: The proposed methodology allows creation of sustainable 3D tumor TE constructs to explore the role of organ-specific cell-matrix interaction in tumorigenesis.

  14. Comparative studies on amniotic fluid and plasma fibronectins.

    PubMed Central

    Ruoslahti, E; Engvall, E; Hayman, E G; Spiro, R G

    1981-01-01

    Human fibronectin was isolated from second-trimester amniotic fluid, from amniotic fluid obtained at term and from adult plasma. The amniotic-fluid fibronectins had a slightly higher apparent molecular weight on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis than the plasma fibronectin. Early- and late-amniotic-fluid fibronectin had 9.5 and 9.6% carbohydrate respectively, whereas plasma fibronectin had 5.8%. The amniotic-fluid fibronectins had similar mannose and sialic acid contents to plasma fibronectin, but greater amounts of glucosamine, galactosamine, galactose and fucose. There were no detectable differences in the amino-acid composition of amniotic-fluid and plasma fibronectins, and the patterns of peptides obtained after tryptic digestion of fibronectin from the two sources showed extensive similarities. Fibronectins from plasma and amniotic fluid were equally active in promoting cell attachment and were immunologically indistinguishable. These results show that fibronectin from amniotic fluid is more heavily glycosylated than plasma fibronectin or previously analysed fibronectins from cultured fibroblasts. The observed differences in glycosylation may be related to cell type and/or stage of development. Images Fig. 2. PMID:7305927

  15. Amniotic fluid embolism.

    PubMed

    Kaur, Kiranpreet; Bhardwaj, Mamta; Kumar, Prashant; Singhal, Suresh; Singh, Tarandeep; Hooda, Sarla

    2016-01-01

    Amniotic fluid embolism (AFE) is one of the catastrophic complications of pregnancy in which amniotic fluid, fetal cells, hair, or other debris enters into the maternal pulmonary circulation, causing cardiovascular collapse. Etiology largely remains unknown, but may occur in healthy women during labour, during cesarean section, after abnormal vaginal delivery, or during the second trimester of pregnancy. It may also occur up to 48 hours post-delivery. It can also occur during abortion, after abdominal trauma, and during amnio-infusion. The pathophysiology of AFE is not completely understood. Possible historical cause is that any breach of the barrier between maternal blood and amniotic fluid forces the entry of amniotic fluid into the systemic circulation and results in a physical obstruction of the pulmonary circulation. The presenting signs and symptoms of AFE involve many organ systems. Clinical signs and symptoms are acute dyspnea, cough, hypotension, cyanosis, fetal bradycardia, encephalopathy, acute pulmonary hypertension, coagulopathy etc. Besides basic investigations lung scan, serum tryptase levels, serum levels of C3 and C4 complements, zinc coproporphyrin, serum sialyl Tn etc are helpful in establishing the diagnosis. Treatment is mainly supportive, but exchange transfusion, extracorporeal membrane oxygenation, and uterine artery embolization have been tried from time to time. The maternal prognosis after amniotic fluid embolism is very poor though infant survival rate is around 70%.

  16. Amniotic fluid embolism

    PubMed Central

    Kaur, Kiranpreet; Bhardwaj, Mamta; Kumar, Prashant; Singhal, Suresh; Singh, Tarandeep; Hooda, Sarla

    2016-01-01

    Amniotic fluid embolism (AFE) is one of the catastrophic complications of pregnancy in which amniotic fluid, fetal cells, hair, or other debris enters into the maternal pulmonary circulation, causing cardiovascular collapse. Etiology largely remains unknown, but may occur in healthy women during labour, during cesarean section, after abnormal vaginal delivery, or during the second trimester of pregnancy. It may also occur up to 48 hours post-delivery. It can also occur during abortion, after abdominal trauma, and during amnio-infusion. The pathophysiology of AFE is not completely understood. Possible historical cause is that any breach of the barrier between maternal blood and amniotic fluid forces the entry of amniotic fluid into the systemic circulation and results in a physical obstruction of the pulmonary circulation. The presenting signs and symptoms of AFE involve many organ systems. Clinical signs and symptoms are acute dyspnea, cough, hypotension, cyanosis, fetal bradycardia, encephalopathy, acute pulmonary hypertension, coagulopathy etc. Besides basic investigations lung scan, serum tryptase levels, serum levels of C3 and C4 complements, zinc coproporphyrin, serum sialyl Tn etc are helpful in establishing the diagnosis. Treatment is mainly supportive, but exchange transfusion, extracorporeal membrane oxygenation, and uterine artery embolization have been tried from time to time. The maternal prognosis after amniotic fluid embolism is very poor though infant survival rate is around 70%. PMID:27275041

  17. Amniotic fluid embolism.

    PubMed

    Kaur, Kiranpreet; Bhardwaj, Mamta; Kumar, Prashant; Singhal, Suresh; Singh, Tarandeep; Hooda, Sarla

    2016-01-01

    Amniotic fluid embolism (AFE) is one of the catastrophic complications of pregnancy in which amniotic fluid, fetal cells, hair, or other debris enters into the maternal pulmonary circulation, causing cardiovascular collapse. Etiology largely remains unknown, but may occur in healthy women during labour, during cesarean section, after abnormal vaginal delivery, or during the second trimester of pregnancy. It may also occur up to 48 hours post-delivery. It can also occur during abortion, after abdominal trauma, and during amnio-infusion. The pathophysiology of AFE is not completely understood. Possible historical cause is that any breach of the barrier between maternal blood and amniotic fluid forces the entry of amniotic fluid into the systemic circulation and results in a physical obstruction of the pulmonary circulation. The presenting signs and symptoms of AFE involve many organ systems. Clinical signs and symptoms are acute dyspnea, cough, hypotension, cyanosis, fetal bradycardia, encephalopathy, acute pulmonary hypertension, coagulopathy etc. Besides basic investigations lung scan, serum tryptase levels, serum levels of C3 and C4 complements, zinc coproporphyrin, serum sialyl Tn etc are helpful in establishing the diagnosis. Treatment is mainly supportive, but exchange transfusion, extracorporeal membrane oxygenation, and uterine artery embolization have been tried from time to time. The maternal prognosis after amniotic fluid embolism is very poor though infant survival rate is around 70%. PMID:27275041

  18. Effect of the Mode of Application of Cryopreserved Human Amniotic Membrane on Adhesion Formation after Abdomino-Pelvic Surgery in a Mouse Model

    PubMed Central

    Nassif, Joseph; Abbasi, Sehrish A.; Kechli, Mohamad Karim; Boutary, Suzan S.; Ghulmiyyah, Labib; Khalifeh, Ibrahim; Abou Ghaddara, Hussein; Nassar, Anwar H.

    2016-01-01

    Adhesions after abdomino-pelvic surgery are a cause of morbidity and reoperations. The use of human amniotic membrane (HAM) for adhesion prevention has given controversial results. The mode of administration of the amniotic membrane has not been well studied. This study assessed the efficacy of two modes of application of cryopreserved HAM, patch or fragmented in Lactated Ringer (LR) solution, for the prevention of pelvic adhesion formation postabdomino-pelvic surgery in a mice model. After a midline laparotomy incision, a small cautery lesion was done on each side of the abdominal wall peritoneum in mice. In Group A (control; n = 42), the abdomen was closed directly, Group B (n = 42) received 2.5 ml of LR prior to closure. In Groups C (n = 42) and D (n = 42), a 2 cm × 2 cm patch of HAM and another one fragmented and dispersed in 2.5 ml of LR were applied prior to closure, respectively. Two weeks later, a laparotomy was performed, and gross and pathological evaluation of adhesions, fibrosis, angiogenesis, and inflammation were conducted. Group D exhibited a significantly lower rate of gross adhesion formation. Fibrosis was significantly lowest in Group C as compared to the control. Group B had the lowest vascular formation in the adhesions. The use of HAM fragmented in LR solution is associated with a significantly lower incidence of postoperative adhesions in mice when compared to LR alone, HAM patch, or control. The mechanism of action of this reduction needs to be elucidated by future studies. PMID:27066485

  19. Nitric oxide-mediated immunosuppressive effect of human amniotic membrane-derived mesenchymal stem cells on the viability and migration of microglia.

    PubMed

    Yan, Ke; Zhang, Run; Chen, Lei; Chen, Fanfan; Liu, Yi; Peng, Lingmei; Sun, Haitao; Huang, Weiyi; Sun, Chengmei; Lv, Bingke; Li, Feng; Cai, Yingqian; Tang, Yanping; Zou, Yuxi; Du, Mouxuan; Qin, Lingsha; Zhang, Hengzhu; Jiang, Xiaodan

    2014-11-24

    Human amniotic membrane-derived mesenchymal stem cells (AMSCs) are considered a novel and promising source of stem cells for cell replacement-based therapy. Current research is mostly limited to investigating the cellular differentiation potential of AMSCs, while few have focused on their immunosuppressive properties. This study is aimed at exploring and evaluating the immunosuppressive effect of human AMSCs on the viability and migratory properties of microglia. We found, from results of cell viability assays, that AMSCs can reduce the activity of inflammatory cells by secreting nitric oxide (NO). Also, based on results from wound healing and transwell migration assays, we show that AMSCs can inhibit the migration of human microglia as well as the mouse microglial cell line BV2, suggesting that they have the ability to inhibit the recruitment of certain immune cells to injury sites. Furthermore, we found that NO contributes significantly to this inhibitory effect. Our study provides evidence that human AMSCs can have detrimental effects on the viability and migration of microglia, through secretion of NO. This mechanism may contribute to anti-inflammatory processes in the central nervous system.

  20. Pdx1 and controlled culture conditions induced differentiation of human amniotic fluid-derived stem cells to insulin-producing clusters.

    PubMed

    Chun, So Young; Mack, David L; Moorefield, Emily; Oh, Se Heang; Kwon, Tae Gyun; Pettenati, Mark J; Yoo, James J; Coppi, Paolo De; Atala, Anthony; Soker, Shay

    2015-05-01

    This study investigated the differentiation of human amniotic fluid-derived stem cells (hAFSCs) into insulin-producing clusters in vitro. Adenovirally-delivered mouse Pdx1 (Ad-Pdx1) induced human Pdx1 expression in hAFSCs and enhanced the coordinated expression of downstream β-cell markers. When Ad-Pdx1-transduced hAFSCs were sequentially treated with activin A, bFGF and nicotinamide and the culture plate surface coated with poly-l-ornithine, the expression of islet-associated human mRNAs for Pdx1, Pax6, Ngn3 and insulin was increased. C-peptide ELISA confirmed that Ad-Pdx1-transduced hAFSCs processed and secreted insulin in a manner consistent with that pathway in pancreatic β-cells. To sustain the β-cell-like phenotype and investigate the effect of three-dimensional (3D) conformation on the differentiation of hAFSCs, Pdx1-transduced cells were encapsulated in alginate and cultured long-term under serum-free conditions. Over 2 weeks, partially differentiated hAFSC clusters increased in size and increased insulin secretion. Taken together, these data demonstrate that ectopic Pdx1 expression initiates pancreatic differentiation in hAFSCs and that a β-cell-like phenotype can be augmented by culture conditions that mimic the stromal components and 3D geometry associated with pancreatic islets.

  1. Reconstruction of limbal stem cell deficient corneal surface with induced human bone marrow mesenchymal stem cells on amniotic membrane.

    PubMed

    Rohaina, Che Man; Then, Kong Yong; Ng, Angela Min Hwei; Wan Abdul Halim, Wan Haslina; Zahidin, Aida Zairani Mohd; Saim, Aminuddin; Idrus, Ruszymah B H

    2014-03-01

    The cornea can be damaged by a variety of clinical disorders or chemical, mechanical, and thermal injuries. The objectives of this study were to induce bone marrow mesenchymal stem cells (BMSCs) to corneal lineage, to form a tissue engineered corneal substitute (TEC) using BMSCs, and to treat corneal surface defects in a limbal stem cell deficiency model. BMSCs were induced to corneal lineage using limbal medium for 10 days. Induced BMSCs demonstrated upregulation of corneal stem cell markers; β1-integrin, C/EBPδ, ABCG2, and p63, increased protein expression of CK3 and p63 significantly compared with the uninduced ones. For TEC formation, passage 1 BMSCs were trypsinized and seeded on amniotic membrane in a transwell co-culture system and were grown in limbal medium. Limbal stem cell deficiency models were induced by alkaline injury, and the TEC was implanted for 8 weeks. Serial slit lamp evaluation revealed remarkable improvement in corneal regeneration in terms of corneal clarity and reduced vascularization. Histologic and optical coherence tomography analyses demonstrated comparable corneal thickness and achieved stratified epithelium with a compact stromal layer resembling that of normal cornea. CK3 and p63 were expressed in the newly regenerated cornea. In conclusion, BMSCs can be induced into corneal epithelial lineage, and these cells are viable for the formation of TEC, to be used for the reconstruction of the corneal surface in the limbal stem cell deficient model.

  2. The effect of amniotic membrane preparation method on its ability to serve as a substrate for the ex-vivo expansion of limbal epithelial cells.

    PubMed

    Shortt, Alex J; Secker, Genevieve A; Lomas, Richard J; Wilshaw, Stacy P; Kearney, John N; Tuft, Stephen J; Daniels, Julie T

    2009-02-01

    Human amniotic membrane (HAM) is employed as a substrate for the ex-vivo expansion of limbal epithelial cells (LECs) used to treat corneal epithelial stem cell deficiency in humans. The optimal method of HAM preparation for this purpose is unknown. This study evaluated the ability of different preparations of stored HAM to serve as substrates for LEC expansion ex-vivo. The effect of removing the amniotic epithelial cells (decellularisation) from HAM prior to seeding of LECs, the effect of glycerol cryopreservation and the effect of peracetic acid (PAA) sterilization and antibiotic disinfection were evaluated using different HAM test groups. Human LECs were cultured on each preparation and the following outcomes were assessed: confluence of growth, cell density, cell morphology and expression of the putative LESC markers deltaN-p63alpha and ABCG2. Removing amniotic epithelial cells prior to seeding of LECs resulted in a higher percentage of confluence but a lower cell density than intact HAM suggesting that decellularisation does not increase proliferation, but rather that it facilitates migration of LECs resulting in larger cells. Decellularisation did not affect the percentage of cells expressing the putative LESC markers deltaN-p63alpha (< or =4% in both intact and acellular groups) and ABCG2 (< or =3% in both intact and acellular groups). Glycerol cryopreservation of HAM resulted in poor morphology and a low proportion of cells expressing deltaN-p63alpha (< or =6%) and ABCG2 (< or =8%). HAM frozen at -80 degrees C in Hank's Balanced Salt Solution (HBSS) was superior, demonstrating excellent morphology of cultured LECs and high levels of deltaN-p63alpha (< or =68%) and ABCG2 (< or =62%) expression (p<0.001). The use of PAA or antibiotics to decontaminate HAM does not appear to affect this function. The variables affecting the ability of HAM to serve as a substrate for LEC expansion ex-vivo are poorly understood. The use of glycerol as a cryoprotectant impairs

  3. Effect of targeted ovarian cancer therapy using amniotic fluid mesenchymal stem cells transfected with enhanced green fluorescent protein-human interleukin-2 in vivo

    PubMed Central

    YOU, QI; YAO, YUAN; ZHANG, YUANLONG; FU, SONGBIN; DU, MEI; ZHANG, GUANGMEI

    2015-01-01

    The aim of the present study was to investigate the effect of using amniotic fluid mesenchymal stem cells (AF-MSCs) in targeted ovarian cancer therapy in vivo. AF-MSCs were isolated from human second trimester AF and a plasmid, enhanced green fluorescent protein-human interleukin-2 (pEGFP-hIL-2) was formed. The plasmid was stably transfected into the AF-MSCs and the cells were intravenously injected into ovarian cancer nude mice models. Following stable transfection of the vector, tumor formation, and the expression and activity of hIL-2 were investigated, and microscopic pathological examinations of the tumor were performed. It was found that AF-MSCs exhibited high motility during migration in vivo, and the vector, pEGFP-hIL-2 can be stably transfected into AF-MSCs. Following stable transfection, this type of stem cell is able to successfully transport the therapeutic gene, IL-2, migrate to the ovarian cancer tumor site to secrete the functional IL-2 and treat the tumor. Thus, AF-MSCs may serve as transporters for therapeutic genes targeting ovarian tumor sites and, therefore, be involved in the treatment of tumors. PMID:26179662

  4. Pleiotrophin is involved in the amniotic epithelial cell-induced differentiation of human umbilical cord blood-derived mesenchymal stem cells into dopaminergic neuron-like cells.

    PubMed

    Yang, Shu; Xue, Dan-Dan; Wu, Bo; Sun, Hai-Mei; Li, Xiao-Shuang; Dong, Fang; Li, Wen-Shuai; Ji, Feng-Qing; Zhou, De-Shan

    2013-02-28

    We have reported that human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) are capable of differentiating into dopaminergic (DA) neuron-like cells upon being induced by amniotic epithelial cells (AECs). However, what factor(s) is involved in the differentiation process has not been explored out thoroughly. Because pleiotrophin (PTN) is known to exert important trophic effects on DA neurons, in the present study, we investigated whether PTN is released by AECs and whether it is involved in the differentiation of hUCB-MSCs into DA neuron-like cells. The expression and secretion of PTN by AECs were detected by immunofluorescence, RT-PCR and ELISA. The hUCB-MSCs were isolated and treated with AEC-conditioned medium (ACM) or recombinant human PTN. Compared to the controls, a higher proportion of treated cells differentiated into DA neuron-like cells, indicated by the increased expression of TH and DAT and the increased dopamine content. These results indicate that PTN released by AECs acts as a synergetic factor with other neurotrophic factors and is involved in the differentiation of hUCB-MSCs into DA neuron-like cells. We suggest that ACM, which contains PTN and other neurotrophic factors, could potentially be used as an agent to promote the differentiation of DA neuron-like cells from hUCB-MSCs for cell therapy of Parkinson's disease without creating legal or ethical issues.

  5. Effect of targeted ovarian cancer therapy using amniotic fluid mesenchymal stem cells transfected with enhanced green fluorescent protein-human interleukin-2 in vivo.

    PubMed

    You, Qi; Yao, Yuan; Zhang, Yuanlong; Fu, Songbin; Du, Mei; Zhang, Guangmei

    2015-10-01

    The aim of the present study was to investigate the effect of using amniotic fluid mesenchymal stem cells (AF-MSCs) in targeted ovarian cancer therapy in vivo. AF-MSCs were isolated from human second trimester AF and a plasmid, enhanced green fluorescent protein‑human interleukin‑2 (pEGFP‑hIL‑2) was formed. The plasmid was stably transfected into the AF‑MSCs and the cells were intravenously injected into ovarian cancer nude mice models. Following stable transfection of the vector, tumor formation, and the expression and activity of hIL‑2 were investigated, and microscopic pathological examinations of the tumor were performed. It was found that AF‑MSCs exhibited high motility during migration in vivo, and the vector, pEGFP‑hIL‑2 can be stably transfected into AF‑MSCs. Following stable transfection, this type of stem cell is able to successfully transport the therapeutic gene, IL-2, migrate to the ovarian cancer tumor site to secrete the functional IL-2 and treat the tumor. Thus, AF-MSCs may serve as transporters for therapeutic genes targeting ovarian tumor sites and, therefore, be involved in the treatment of tumors.

  6. [Astragalus induces human amniotic epithelial cells (WISH) to differentiate toward neurons, inhibits the expression of Notch1 and promotes cell survival].

    PubMed

    Chen, Xu-Dong; Wang, Jian-Guo

    2012-12-25

    The aim of the study was to investigate the effect of astragalus on differentiation of human amniotic epithelial cell line WISH into neurons, the expression of Notch1 gene and cell viability. WISH were randomly divided into astragalus group (4 subgroups), alltransretinoic acid (RA) group and control group. Astragalus group and RA group were induced to differentiate into neurocytes by using chemical inducer RA and astragalus, respectively. The expression of neuron-specific enolase (NSE), microtubule associated protein 2 (MAP-2), Nestin and GFAP of induced cells in three groups were detected using immunocytochemical method. RT- PCR was further used to detect the expression of Oct4, Notch1, Hes1, Nestin and NSE. The cell viability was measured by methyl thiazolyl tetrazolium methods. Under the convert microscope it was observed that WISH cells started to change their shape, and there were several axon or dendrite-like processes out from the cell body induced by astragalus for 24 h or RA for 12 h. The positive cell rates of NSE and MAP-2 in 100 μL/mL astragalus-induced group were less than those in RA-induced group at 48 h (P < 0.05), but higher than those in control group. Cell viability in astragalus group was higher than that of RA group (P < 0.05). While the positive cell rates of Nestin and GFAP in 100 μL/mL astragalus-induced group were higher than those in RA-induced group at 48 h (P < 0.05). The positive cell rates of Nestin in the two induced groups were lower than those in control group. RT-PCR showed that the expressions of Oct4, Notch1 and Hes1 in RA and astragalus (100 μL/mL) groups were less than those in control group, but the expression of NSE was higher than that in control group. These results suggest that astragalus (especially at 100 μL/mL, 48 h) and RA can both induce human amniotic epithelial cell line WISH cells into neuron-like cells, but astragalus induction has a higher cell survival rate than RA induction, and the expression of Notch1

  7. Acellular pertussis vaccines in China.

    PubMed

    Wang, Lichan; Lei, Dianliang; Zhang, Shumin

    2012-11-26

    In China, whole-cell pertussis (Pw) vaccines were produced in the early 1960s and acellular pertussis (Pa) vaccines were introduced in 1995. Pa vaccines have now almost completely replaced Pw vaccines in the national immunization program. To strengthen the regulation of vaccines used in China, a vaccine lot release system was established in 2001 and Pa vaccines have been included in the system since 2006. This paper mainly described the current status of production and the quality control measures in place for Pa vaccines; and analyses quality control test data accumulated between 2006 and 2010.

  8. Xenogeneic acellular conjunctiva matrix as a scaffold of tissue-engineered corneal epithelium.

    PubMed

    Zhao, Haifeng; Qu, Mingli; Wang, Yao; Wang, Zhenyu; Shi, Weiyun

    2014-01-01

    Amniotic membrane-based tissue-engineered corneal epithelium has been widely used in the reconstruction of the ocular surface. However, it often degrades too early to ensure the success of the transplanted corneal epithelium when treating patients with severe ocular surface disorders. In the present study, we investigated the preparation of xenogeneic acellular conjunctiva matrix (aCM) and evaluated its efficacy and safety as a scaffold of tissue-engineered corneal epithelium. Native porcine conjunctiva was decellularized with 0.1% sodium dodecyl sulfate (SDS) for 12 h at 37°C and sterilized via γ-irradiation. Compared with native conjunctiva, more than 92% of the DNA was removed, and more than 90% of the extracellular matrix components (glycosaminoglycan and collagen) remained after the decellularization treatment. Compared with denuded amniotic membrane (dAM), the aCM possessed favorable optical transmittance, tensile strength, stability and biocompatibility as well as stronger resistance to degradation both in vitro and in vivo. The corneal epithelial cells seeded on aCM formed a multilayered epithelial structure and endured longer than did those on dAM. The aCM-based tissue-engineered corneal epithelium was more effective in the reconstruction of the ocular surface in rabbits with limbal stem cell deficiency. These findings support the application of xenogeneic acellular conjunctiva matrix as a scaffold for reconstructing the ocular surface.

  9. Investigating the effect of hypoxic culture on the endothelial differentiation of human amniotic fluid-derived stem cells.

    PubMed

    Lloyd-Griffith, Cai; Duffy, Garry P; O'Brien, Fergal J

    2015-12-01

    Amniotic fluid-derived stem cells (AFSCs) are a unique stem cell source that may have great potential for use in tissue engineering (TE) due to their pluripotentiality. AFSCs have previously shown angiogenic potential and may present an alternative cell source for endothelial-like cells that could be used in range of applications, including the pre-vascularisation of TE constructs and the treatment of ischaemic diseases. This study investigated the ability of these cells to differentiate down an endothelial lineage with the aim of producing an endothelial-like cell suitable for use in pre-vascularisation. As hypoxia and the associated HIF-1 pathway have been implicated in the induction of angiogenesis in a number of biological processes, it was hypothesised that culture in hypoxic conditions could enhance the endothelial differentiation of AFSCs. The cells were cultured in endothelial cell media supplemented with 50 ng mL(-1) of VEGF, maintained in normoxia, intermittent hypoxia or continuous hypoxia and assessed for markers of endothelial differentiation at day 7 and 14. The results demonstrated that AFSCs subjected to these culture conditions display an endothelial gene expression profile and adopted functional endothelial cell characteristics indicative of early endothelial differentiation. Culture in continuous hypoxia enhanced endothelial gene expression but did not enhance functional endothelial cell characteristics. Overall, AFSCs subjected to endothelial stimuli demonstrated a less mature endothelial gene expression profile and phenotype when compared with HUVECs, the endothelial cell control. However, this study is the first time that the positive effect of an extended period of continuous hypoxic culture on endothelial differentiation in AFSCs has been demonstrated.

  10. Genetic reprogramming of human amniotic cells with episomal vectors: neural rosettes as sentinels in candidate selection for validation assays

    PubMed Central

    Payne, Tiffany

    2014-01-01

    The promise of genetic reprogramming has prompted initiatives to develop banks of induced pluripotent stem cells (iPSCs) from diverse sources. Sentinel assays for pluripotency could maximize available resources for generating iPSCs. Neural rosettes represent a primitive neural tissue that is unique to differentiating PSCs and commonly used to identify derivative neural/stem progenitors. Here, neural rosettes were used as a sentinel assay for pluripotency in selection of candidates to advance to validation assays. Candidate iPSCs were generated from independent populations of amniotic cells with episomal vectors. Phase imaging of living back up cultures showed neural rosettes in 2 of the 5 candidate populations. Rosettes were immunopositive for the Sox1, Sox2, Pax6 and Pax7 transcription factors that govern neural development in the earliest stage of development and for the Isl1/2 and Otx2 transcription factors that are expressed in the dorsal and ventral domains, respectively, of the neural tube in vivo. Dissociation of rosettes produced cultures of differentiation competent neural/stem progenitors that generated immature neurons that were immunopositive for βIII-tubulin and glia that were immunopositive for GFAP. Subsequent validation assays of selected candidates showed induced expression of endogenous pluripotency genes, epigenetic modification of chromatin and formation of teratomas in immunodeficient mice that contained derivatives of the 3 embryonic germ layers. Validated lines were vector-free and maintained a normal karyotype for more than 60 passages. The credibility of rosette assembly as a sentinel assay for PSCs is supported by coordinate loss of nuclear-localized pluripotency factors Oct4 and Nanog in neural rosettes that emerge spontaneously in cultures of self-renewing validated lines. Taken together, these findings demonstrate value in neural rosettes as sentinels for pluripotency and selection of promising candidates for advance to validation

  11. Genetic reprogramming of human amniotic cells with episomal vectors: neural rosettes as sentinels in candidate selection for validation assays.

    PubMed

    Wilson, Patricia G; Payne, Tiffany

    2014-01-01

    The promise of genetic reprogramming has prompted initiatives to develop banks of induced pluripotent stem cells (iPSCs) from diverse sources. Sentinel assays for pluripotency could maximize available resources for generating iPSCs. Neural rosettes represent a primitive neural tissue that is unique to differentiating PSCs and commonly used to identify derivative neural/stem progenitors. Here, neural rosettes were used as a sentinel assay for pluripotency in selection of candidates to advance to validation assays. Candidate iPSCs were generated from independent populations of amniotic cells with episomal vectors. Phase imaging of living back up cultures showed neural rosettes in 2 of the 5 candidate populations. Rosettes were immunopositive for the Sox1, Sox2, Pax6 and Pax7 transcription factors that govern neural development in the earliest stage of development and for the Isl1/2 and Otx2 transcription factors that are expressed in the dorsal and ventral domains, respectively, of the neural tube in vivo. Dissociation of rosettes produced cultures of differentiation competent neural/stem progenitors that generated immature neurons that were immunopositive for βIII-tubulin and glia that were immunopositive for GFAP. Subsequent validation assays of selected candidates showed induced expression of endogenous pluripotency genes, epigenetic modification of chromatin and formation of teratomas in immunodeficient mice that contained derivatives of the 3 embryonic germ layers. Validated lines were vector-free and maintained a normal karyotype for more than 60 passages. The credibility of rosette assembly as a sentinel assay for PSCs is supported by coordinate loss of nuclear-localized pluripotency factors Oct4 and Nanog in neural rosettes that emerge spontaneously in cultures of self-renewing validated lines. Taken together, these findings demonstrate value in neural rosettes as sentinels for pluripotency and selection of promising candidates for advance to validation

  12. Ex vivo expanded autologous limbal epithelial cells on amniotic membrane using a culture medium with human serum as single supplement.

    PubMed

    Shahdadfar, Aboulghassem; Haug, Kristiane; Pathak, Meeta; Drolsum, Liv; Olstad, Ole Kristoffer; Johnsen, Erik O; Petrovski, Goran; Moe, Morten C; Nicolaissen, Bjørn

    2012-04-01

    In patients with limbal stem cell deficiency (LSCD), transplantation of ex vivo expanded human limbal epithelial cells (HLECs) can restore the structural and functional integrity of the corneal surface. However, the protocol for cultivation and transplantation of HLECs differ significantly, and in most protocols growth additives such as cholera toxins, exogenous growth factors, hormones and fetal calf serum are used. In the present article, we compare for the first time human limbal epithelial cells (HLECs) cultivated on human amniotic membrane (HAM) in a complex medium (COM) including fetal bovine serum to a medium with human serum as single growth supplement (HSM), and report on our first examinations of HLECs expanded in autologous HSM and used for transplant procedures in patients with LSCD. Expanded HLECs were examined by genome-wide microarray, RT-PCR, Western blotting, and for cell viability, morphology, expression of immunohistochemical markers and colony forming efficiency. Cultivation of HLECs in HSM produced a multilayered epithelium where cells with markers associated with LESCs were detected in the basal layers. There were few transcriptional differences and comparable cell viability between cells cultivated in HSM and COM. The p63 gene associated with LESCs were expressed 3.5 fold more in HSM compared to COM, and Western blotting confirmed a stronger p63α band in HSM cultures. The cornea-specific keratin CK12 was equally found in both culture conditions, while there were significantly more CK3 positive cells in HSM. Cells in epithelial sheets on HAM remaining after transplant surgery of patients with LSCD expressed central epithelial characteristics, and dissociated cells cultured at low density on growth-arrested fibroblasts produced clones containing 21 ± 12% cells positive for p63α (n = 3). In conclusion, a culture medium without growth additives derived from animals or from animal cell cultures and with human serum as single growth supplement

  13. Enhanced Ex Vivo Expansion of Human Hematopoietic Progenitors on Native and Spin Coated Acellular Matrices Prepared from Bone Marrow Stromal Cells

    PubMed Central

    Wasnik, Samiksha; Kantipudi, Suma; Kirkland, Mark A.; Pande, Gopal

    2016-01-01

    The extracellular microenvironment in bone marrow (BM) is known to regulate the growth and differentiation of hematopoietic stem and progenitor cells (HSPC). We have developed cell-free matrices from a BM stromal cell line (HS-5), which can be used as substrates either in native form or as tissue engineered coatings, for the enhanced ex vivo expansion of umbilical cord blood (UCB) derived HSPC. The physicochemical properties (surface roughness, thickness, and uniformity) of native and spin coated acellular matrices (ACM) were studied using scanning and atomic force microscopy (SEM and AFM). Lineage-specific expansion of HSPC, grown on these substrates, was evaluated by immunophenotypic (flow cytometry) and functional (colony forming) assays. Our results show that the most efficient expansion of lineage-specific HSPC occurred on spin coated ACM. Our method provides an improved protocol for ex vivo HSPC expansion and it offers a system to study the in vivo roles of specific molecules in the hematopoietic niche that influence HSPC expansion. PMID:26981135

  14. Elemental analysis of human amniotic fluid and placenta by total-reflection X-ray fluorescence and energy-dispersive X-ray fluorescence: child weight and maternal age dependence

    NASA Astrophysics Data System (ADS)

    Carvalho, M. L.; Custódio, P. J.; Reus, U.; Prange, A.

    2001-11-01

    This work is an attempt to evaluate the possible influence of the mother's age in trace element concentrations in human amniotic fluid and placenta and whether these concentrations are correlated to the weight of the newborn infants. Total-reflection X-ray fluorescence (TXRF) was used to analyze 16 amniotic fluid samples, and the placenta samples were analyzed by energy dispersive X-ray fluorescence (EDXRF). The whole samples were collected during delivery from healthy mothers and healthy infants and full-term pregnancies. According to the age of the mother, three different groups were considered: 20-25, 25-30 and 30-40 years old. Only two mothers were aged more than 35 years. The weight of the infants ranged from 2.56 to 4.05 kg and three groups were also considered: 2.5-3, 3-3.5 and 3.5-4 kg. The organic matrix of the amniotic fluid samples was removed by treatment with HNO 3 followed by oxygen plasma ashing. Yttrium was used as the internal standard for TXRF analysis. Placenta samples were lyophilized and analyzed by EDXRF without any chemical treatment. Very low levels of Ni and Sr were found in the amniotic fluid samples, and were independent of the age of the mother and weight of the child. Cr, Mn, Se and Pb were at the level of the detection limit. Zn, considered one of the key elements in neonatal health, was not significantly different in the samples analyzed; however, it was weakly related to birth weigh. The concentrations obtained ranged from 0.11 to 0.92 mg/l and 30 to 65 μg/g in amniotic fluid and placenta, respectively. The only two elements which seemed to be significantly correlated with mother's age and newborn weight were Ca and Fe for both types of sample: Ca levels were increased in heavier children and older mothers; however, Fe increased with increasing maternal age, but decreased for heavier babies. The same conclusions were obtained for placenta and amniotic fluid samples. Cu is closely associated with Fe in its function in the organism

  15. The safety of photochemical tissue bonding for treating damaged corneal epithelium using limbal stem cells pre-cultured on human amniotic membrane.

    PubMed

    Gu, Chuan; Yang, Jun; Yuan, Ying; Yao, Min; Zhang, Xiong

    2015-07-01

    We previously demonstrated the feasibility of treating limbal stem cell deficiency (LSCD) with limbal stem cells (LSCs) pre-cultured on human amniotic membrane (HAM), using a suture-free technique called photochemical tissue bonding (PTB). However, important issues regarding the safety and the influence of PTB on LSCs have not been elucidated. In this study, LSCs, isolated from rabbit eyes and identified by cell markers, were labeled with BrdU prior to cultivation on de-epithelialized HAM to fabricate grafts. Rabbit LSCD models were created and randomly divided into groups for transplantation of fabricated grafts using sutures or PTB (n=10). Possible phototoxicity of PTB to LSCs was analyzed in vitro and in vivo. Restoration of corneal epithelium was evaluated at 28 days after grafting. Our results showed that phototoxicity did not occur in the LSCs cultured on HAM after PTB in vitro. Transplantation of grafts with PTB restored the damaged cornea epithelium effectively and no significant influences on LSC characteristics were found in both sutured and PTB groups. BrdU positive cells were tracked at 28 days post grafting suggesting that the restored epithelium was derived from the in vitro fabricated HAM/LSC graft. These data suggest that PTB is a safe and potential strategy for securing LSC/HAM grafts that produces with better outcomes than sutured attachment.

  16. Escalated regeneration in sciatic nerve crush injury by the combined therapy of human amniotic fluid mesenchymal stem cells and fermented soybean extracts, Natto

    PubMed Central

    Pan, Hung-Chuan; Yang, Dar-Yu; Ho, Shu-Peng; Sheu, Meei-Ling; Chen, Chung-Jung; Hwang, Shiaw-Min; Chang, Ming-Hong; Cheng, Fu-Chou

    2009-01-01

    Attenuation of inflammatory cell deposits and associated cytokines prevented the apoptosis of transplanted stem cells in a sciatic nerve crush injury model. Suppression of inflammatory cytokines by fermented soybean extracts (Natto) was also beneficial to nerve regeneration. In this study, the effect of Natto on transplanted human amniotic fluid mesenchymal stem cells (AFS) was evaluated. Peripheral nerve injury was induced in SD rats by crushing a sciatic nerve using a vessel clamp. Animals were categorized into four groups: Group I: no treatment; Group II: fed with Natto (16 mg/day for 7 consecutive days); Group III: AFS embedded in fibrin glue; Group IV: Combination of group II and III therapy. Transplanted AFS and Schwann cell apoptosis, inflammatory cell deposits and associated cytokines, motor function, and nerve regeneration were evaluated 7 or 28 days after injury. The deterioration of neurological function was attenuated by AFS, Natto, or the combined therapy. The combined therapy caused the most significantly beneficial effects. Administration of Natto suppressed the inflammatory responses and correlated with decreased AFS and Schwann cell apoptosis. The decreased AFS apoptosis was in line with neurological improvement such as expression of early regeneration marker of neurofilament and late markers of S-100 and decreased vacuole formation. Administration of either AFS, or Natto, or combined therapy augmented the nerve regeneration. In conclusion, administration of Natto may rescue the AFS and Schwann cells from apoptosis by suppressing the macrophage deposits, associated inflammatory cytokines, and fibrin deposits. PMID:19698158

  17. The effects of different preservation processes on the total protein and growth factor content in a new biological product developed from human amniotic membrane.

    PubMed

    Russo, Alessandra; Bonci, Paola; Bonci, Paolo

    2012-06-01

    The aim of this work is to quantify the total protein and growth factors content in a tissue-suspension obtained from processed human amniotic membrane (hAM). hAM was collected, frozen, freeze dried, powdered and sterilized by γ-irradiation. At each step of the process, samples were characterized for the total protein amounts by a Bradford protein assay and for the growth factor concentrations by ELISA test of the tissue suspensions. Frozen-hAM samples show higher release of total proteins and specific growth factors in the tissue suspension in comparison with freeze-dried hAM. We observed that even if the protein extraction is hindered once the tissue is dried, the powdering process allows a greater release in the tissue suspension of total proteins and growth factors after tissue re-solubilization in comparison with only the freeze-drying process (+91 ± 13% for EGF, +16 ± 4% for HGF, +11 ± 5% for FGF, +16 ± 9% for TGF-β1), and a greater release of EGF (85 ± 10%) in comparison with only the freezing process, because proteins become much readily solubilized in the solution. According with these results, we describe a protocol to obtain a new sterile biological product from hAM tissue, with well-known effects of thermal, mechanical and physical processes on the total protein and grow factors contents.

  18. Human amniotic membrane-derived epithelial stem cells display anticancer activity in BALB/c female nude mice bearing disseminated breast cancer xenografts.

    PubMed

    Kang, Nam-Hee; Yi, Bo-Rim; Lim, So Yoon; Hwang, Kyung-A; Baek, Young Seok; Kang, Kyung-Sun; Choi, Kyung-Chul

    2012-06-01

    Breast cancer is one of the most common malignant tumors and the leading cause of mortality among women. In this study, we propose a human stem cell transplantation strategy, an important method for treating various cancers, as a potential breast cancer therapy. To this end, we used human amniotic membrane-derived epithelial stem cells (hAECs) as a cell source for performing human stem cell transplantation. hAECs have multipotent differentiation abilities and possess high proliferative potential. We transplanted hAECs into female BALB/c nude mice bearing tumors originating from MDA-MB-231 breast cancer cells. Co-culturred hAECs and MDA-MB-231 cells at a ratio of 1:4 or 1:8 (tumor cells to stem cells) inhibited breast cancer cell growth by 67.29 and 67.33%, respectively. In the xenograft mouse model, tumor volumes were significantly decreased by 5-flurouracil (5-FU) treatment and two different ratios of hAECs (1:4 and 1:8) by 84.33, 73.88 and 56.89%, respectively. Treatment of nude mice with hAECs (1:4) produced remarkable antitumor effects without any side-effects (e.g., weight loss, death and bruising) compared to the mice that received only 5-FU treatment. Tumor progression was significantly reduced by hAEC treatment compared to the xenograft model. On the other hand, breast tissues (e.g., the epidermis, dermis and reticular layer) appeared to be well-maintained following treatment with hAECs. Taken together, these results provide strong evidence that hAECs can be used as a safe and effective cancer-targeting cytotherapy for treating breast cancer.

  19. A Prospective Study Assessing Complication Rates and Patient-Reported Outcomes in Breast Reconstructions Using a Novel, Deep Dermal Human Acellular Dermal Matrix

    PubMed Central

    Vu, Michael M.; De Oliveira, Gildasio S.; Mayer, Kristen E.; Blough, Jordan T.

    2015-01-01

    Abstract Background: The value proposition of an acellular dermal matrix (ADM) taken from the deep dermis is that the allograft may be more porous, allowing for enhanced integration and revascularization. In turn, this characteristic may attenuate complications related to foreign body reactions, seromas, and infection. However, this is juxtaposed against the potential loss of allograft structural integrity, with subsequent risk of malposition and extrusion. Despite the active use of novel, deep dermal ADMs, the clinical outcomes of this new technology has not been well studied. Methods: This is a prospective study to evaluate surgical and patient-reported outcomes using a deep dermal ADM, FlexHD Pliable. Surgical outcomes and BREAST-Q patient-reported outcomes were evaluated postoperatively at 2- and 6-month time points. Results: Seventy-two breasts (41 patients) underwent reconstruction. Complication rate was 12.5%, including 2 hematomas and 7 flap necroses. One case of flap necrosis led to reconstructive failure. Notably, there were no cases of infection, seroma, or implant extrusion or malposition. Average BREAST-Q scores were satisfaction with outcome (70.13 ± 23.87), satisfaction with breasts (58.53 ± 20.00), psychosocial well being (67.97 ± 20.93), sexual well being (54.11 ± 27.72), and physical well being (70.45 ± 15.44). Two-month postoperative BREAST-Q scores decreased compared with baseline and returned to baseline by 6 months. Postoperative radiation therapy had a negative effect on satisfaction with breasts (P = 0.004) and sexual well being (P = 0.006). Conclusions: Deep dermal ADM is a novel modification of traditional allograft technology. Use of the deep dermal ADM yielded acceptably low complication rates and satisfactory patient-reported outcomes. PMID:26894010

  20. The targeted inhibitory effects of human amniotic fluid stem cells carrying CXCR4 promoter and DAL-1 on non-small cell lung carcinoma growth.

    PubMed

    Li, L; Li, S; Cai, T; Wang, H; Xie, X; Liu, Z; Zhang, Y

    2016-02-01

    The differentially expressed in adenocarcinoma of the lung-1 (DAL-1) protein has been demonstrated to be suppressive to various types of tumors including lung cancer. This study aimed to determine the targeted effects of human amniotic fluid stem cells (hAFS cells) carrying CXCR4 promoter driven conditionally replicable adenovirus vector overexpressing DAL-1 (Ad-CXCR4-DAL-1) on non-small cell lung carcinoma (NSCLC) growth. The apoptotic effects of virus vectors were assessed using flow cytometry, and the cytotoxicity analyzed by CCK-8 assay. In vivo imaging system was used to determine the homing capability of hAFS cells. A549 cell xenograft mouse model was created to assess the in vivo effect of DAL-1 overexpression on NSCLC growth. We found that infection of Ad-CXCR4-DAL-1 increased the apoptosis of A549 NSCLC cells but not 16HBE normal human bronchial epithelial cells. Ad-CXCR4-DAL-1 administered via intratumoral injection led to significant reduced growth and greater necrosis of A549 xenograft tumors comparing to null vector treated animals. When infused via tail vein, hAFS cells carrying Ad-CXCR4-DAL-1 homed to lung cancer xenografts, caused virus replication and DAL-1 overexpression, and led to significant lower growth and greater necrosis of A549 cell xenografts comparing to non-treatment control. In conclusion, hAFS cells are capable of carrying Ad-CXCR4-DAL-1 vectors, specifically targeting to lung cancer, and causing oncolytic effects when administered in vivo.

  1. Human amniotic epithelial cells can differentiate into granulosa cells and restore folliculogenesis in a mouse model of chemotherapy-induced premature ovarian failure

    PubMed Central

    2013-01-01

    Introduction Ovarian dysfunction frequently occurs in female cancer patients after chemotherapy, but human amniotic epithelial cells (hAECs) that can differentiate into cell types that arise from all three germ layers may offer promise for restoration of such dysfunction. Previous studies confirmed that hAECs could differentiate into cells that express germ cell-specific markers, but at this time hAECs have not been shown to restore ovarian function. Methods To model premature ovarian failure, hAECs infected with lenti-virus carrying green fluorescent protein were injected into the tail vein of mice sterilized with cyclophosphamide and busulphan. hAECs migrated to the mouse ovaries and overall ovarian function was measured using immunohistochemical techniques. Results Seven days to two months after hAECs transplantation, ovarian cells were morphologically restored in sterilized mice. Hemotoxylin and eosin staining revealed that restored ovarian cells developed follicles at all stages. No follicles were observed in control mice at the same time period. Immunostaining with anti-human antigen antibodies and pre-transplantation labeling with green fluorescent protein (GFP) revealed that the grafted hAECs survived and migrated to mouse ovary, differentiating into granulosa cells. Furthermore, the ovarian function marker, anti-Müllerian hormone, was evident in treated mouse ovaries after hAEC transplantation. Conclusions Intravenously injected hAECs reached the ovaries of chemotherapy-treated mice and restored folliculogenesis, data which suggest promise for hAECs for promoting reproductive health and improving the quality of life for female cancer survivors. PMID:24406076

  2. Amniotic fluid fibronectin. Characterization and synthesis by cells in culture

    PubMed Central

    1978-01-01

    A glycoprotein immunologically related to plasma cold-insoluble globulin (CIG) and fetal skin fibroblast fibronectin has been purified from second-trimester human amniotic fluid. This protein (amniotic fluid fibronectin) migrated more slowly than CIG on sodium dodecyl sulfate gel electrophoresis and showed greater polydispersity which could result, at least in part, from heterogeneity in glycosylation. Cloned human amniotic fluid epithelioid and fibroblastic cells synthesized and secreted a protein with similar properties into the culture medium. Fibronectin was shown to be associated with the pericellular and extracellular matrix of cultured amniotic fluid cells by immunofluorescence, lactoperoxidase-catalyzed iodination, and labeling with ferritin-conjugated antibodies. The kinetics of secretion of the protein were consistent with its role as a matrix protein. We anticipate that amniotic fluid fibronectin will prove to be the same protein which elsewhere in the body is incorporated into connective tissues and basement membranes. Amniotic fluid could, therefore, serve as a convenient source of in vivo synthesized fibronectin for biological and structural studies. PMID:701356

  3. Investigation of the regenerative capacity of an acellular porcine medial meniscus for tissue engineering applications.

    PubMed

    Stapleton, Thomas W; Ingram, Joanne; Fisher, John; Ingham, Eileen

    2011-01-01

    Previously, we have described the development of an acellular porcine meniscal scaffold. The aims of this study were to determine the immunocompatibility of the scaffold and capacity for cellular attachment and infiltration to gain insight into its potential for meniscal repair and replacement. Porcine menisci were decellularized by exposing the tissue to freeze-thaw cycles, incubation in hypotonic tris buffer, 0.1% (w/v) sodium dodecyl sulfate in hypotonic buffer plus protease inhibitors, nucleases, hypertonic buffer followed by disinfection using 0.1% (v/v) peracetic, and final washing in phosphate-buffered saline. In vivo immunocompatibility was assessed after implantation of the acellular meniscal scaffold subcutaneously into galactosyltransferase knockout mice for 3 months in comparison to fresh and acellular tissue treated with α-galactosidase (negative control). The cellular infiltrates in the explants were assessed by histology and characterized using monoclonal antibodies against: CD3, CD4, CD34, F4/80, and C3c. Static culture was used to assess the potential of acellular porcine meniscal scaffold to support the attachment and infiltration of primary human dermal fibroblasts and primary porcine meniscal cells in vitro. The explants were surrounded by capsules that were more pronounced for the fresh meniscal tissue compared to the acellular tissues. Cellular infiltrates compromised mononuclear phagocytes, CD34-positive cells, and nonlabeled fibroblastic cells. T-lymphocytes were sparse in all explanted tissue types and there was no evidence of C3c deposition. The analysis revealed an absence of a specific immune response to all of the implanted tissues. Acellular porcine meniscus was shown to be capable of supporting the attachment and infiltration of primary human fibroblasts and primary porcine meniscal cells. In conclusion, acellular porcine meniscal tissue exhibits excellent immunocompatibility and potential for cellular regeneration in the longer term.

  4. Recent advances in acellular regenerative tissue scaffolds.

    PubMed

    Protzman, Nicole M; Brigido, Stephen A

    2015-01-01

    The management of chronic wounds is a considerable challenge for foot and ankle surgeons. The well-established tenets of adequate vascular supply, debridement with eradication of infection, and offloading must be employed in the management of all extremity wounds. Regenerative scaffolds are a viable means of reestablishing a favorable wound environment. The matrix facilitates cell migration, chemoattraction, angiogenesis, wound bed granulation, and expedited wound closure. Although studies have demonstrated success with acellular matrices, a multimodal approach should always be employed to improve healing success. Negative pressure wound therapy, compression, offloading, and antibiotics are advocated to improve outcomes. Acellular graft selection requires a multifactorial analysis, taking into consideration the specific patient and wound characteristics as well as the differences between acellular matrices. Patient age, comorbidities, activity level, and ability to comply with protocol as well as wound etiology, duration, depth, surface area, exudate, bacterial burden, location, vascular status, ischemic status, and presentation are all critical components. To effectively choose a matrix, the clinician must have a comprehensive understanding of the products available and the data validating their use. The mechanisms by which the acellular matrix accelerates wound healing and increases the likelihood of wound healing continue to be investigated. However, it is clear that these acellular biologic tissue scaffolds are incorporating into the host tissue, with resultant revascularization and cellular repopulation. Moving forward, additional investigations examining the effectiveness of acellular biologic tissue scaffolds to improve healing in complex, nondiabetic wounds are warranted.

  5. Induction of E-cadherin+ human amniotic fluid cell differentiation into oocyte-like cells via culture in medium supplemented with follicular fluid.

    PubMed

    Liu, Te; Huang, Yongyi; Bu, Yanzhen; Zhao, Yanhui; Zou, Gang; Liu, Zhixue

    2014-07-01

    Pluripotent human amniotic fluid cells (HuAFCs) can differentiate into various types of somatic cell in vitro. However, their differentiation into oocyte-like cells has never been described to the best of our knowledge. In the present study, differentiation of E-cadherin+ and E-cadherin- HuAFC sub-populations into oocyte-like cells was induced via culture in medium containing bovine follicular fluid and β-mercaptoethanol. The E-cadherin+ HuAFCs expressed DAZL highly. Post-induction, cells with an oocyte-like phenotype were found among the E-cadherin+ HuAFCs, expressing markers specific to germ cells and oocytes (VASA, ZP3 and GDF9) and meiosis (DMC1 and SCP3). When specific small interfering RNA (siRNA) was used to suppress E-cadherin in the E-cadherin+ HuAFCs, the levels of DAZL expression were reduced. Post-induction, the morphology of the siRNA‑E‑cadherin HuAFCs was poorer and the expression levels of germ cell-specific markers were lower compared with those of the siRNA-mock HuAFCs. Therefore, E-cadherin+ HuAFCs could be more easily induced to differentiate into oocyte-like cells by bovine follicular fluid and β-mercaptoethanol. In addition, the E-cadherin+ HuAFCs exhibited potential characteristics of DAZL protein expression, and thus it was conjectured that bovine follicular fluid acts on DAZL protein and promotes E-cadherin+ HuAFC differentiation into oocyte-like cells.

  6. Induction of E-cadherin+ human amniotic fluid cell differentiation into oocyte-like cells via culture in medium supplemented with follicular fluid.

    PubMed

    Liu, Te; Huang, Yongyi; Bu, Yanzhen; Zhao, Yanhui; Zou, Gang; Liu, Zhixue

    2014-07-01

    Pluripotent human amniotic fluid cells (HuAFCs) can differentiate into various types of somatic cell in vitro. However, their differentiation into oocyte-like cells has never been described to the best of our knowledge. In the present study, differentiation of E-cadherin+ and E-cadherin- HuAFC sub-populations into oocyte-like cells was induced via culture in medium containing bovine follicular fluid and β-mercaptoethanol. The E-cadherin+ HuAFCs expressed DAZL highly. Post-induction, cells with an oocyte-like phenotype were found among the E-cadherin+ HuAFCs, expressing markers specific to germ cells and oocytes (VASA, ZP3 and GDF9) and meiosis (DMC1 and SCP3). When specific small interfering RNA (siRNA) was used to suppress E-cadherin in the E-cadherin+ HuAFCs, the levels of DAZL expression were reduced. Post-induction, the morphology of the siRNA‑E‑cadherin HuAFCs was poorer and the expression levels of germ cell-specific markers were lower compared with those of the siRNA-mock HuAFCs. Therefore, E-cadherin+ HuAFCs could be more easily induced to differentiate into oocyte-like cells by bovine follicular fluid and β-mercaptoethanol. In addition, the E-cadherin+ HuAFCs exhibited potential characteristics of DAZL protein expression, and thus it was conjectured that bovine follicular fluid acts on DAZL protein and promotes E-cadherin+ HuAFC differentiation into oocyte-like cells. PMID:24788191

  7. Non-integrating episomal plasmid-based reprogramming of human amniotic fluid stem cells into induced pluripotent stem cells in chemically defined conditions.

    PubMed

    Slamecka, Jaroslav; Salimova, Lilia; McClellan, Steven; van Kelle, Mathieu; Kehl, Debora; Laurini, Javier; Cinelli, Paolo; Owen, Laurie; Hoerstrup, Simon P; Weber, Benedikt

    2016-01-01

    Amniotic fluid stem cells (AFSC) represent an attractive potential cell source for fetal and pediatric cell-based therapies. However, upgrading them to pluripotency confers refractoriness toward senescence, higher proliferation rate and unlimited differentiation potential. AFSC were observed to rapidly and efficiently reacquire pluripotency which together with their easy recovery makes them an attractive cell source for reprogramming. The reprogramming process as well as the resulting iPSC epigenome could potentially benefit from the unspecialized nature of AFSC. iPSC derived from AFSC also have potential in disease modeling, such as Down syndrome or β-thalassemia. Previous experiments involving AFSC reprogramming have largely relied on integrative vector transgene delivery and undefined serum-containing, feeder-dependent culture. Here, we describe non-integrative oriP/EBNA-1 episomal plasmid-based reprogramming of AFSC into iPSC and culture in fully chemically defined xeno-free conditions represented by vitronectin coating and E8 medium, a system that we found uniquely suited for this purpose. The derived AF-iPSC lines uniformly expressed a set of pluripotency markers Oct3/4, Nanog, Sox2, SSEA-1, SSEA-4, TRA-1-60, TRA-1-81 in a pattern typical for human primed PSC. Additionally, the cells formed teratomas, and were deemed pluripotent by PluriTest, a global expression microarray-based in-silico pluripotency assay. However, we found that the PluriTest scores were borderline, indicating a unique pluripotent signature in the defined condition. In the light of potential future clinical translation of iPSC technology, non-integrating reprogramming and chemically defined culture are more acceptable. PMID:26654216

  8. Hepatocyte-like cells derived from human amniotic epithelial cells can be encapsulated without loss of viability or function in vitro.

    PubMed

    Vaghjiani, Vijesh; Vaithilingam, Vijayaganapathy; Saraswati, Indah; Sali, Adnan; Murthi, Padma; Kalionis, Bill; Tuch, Bernard E; Manuelpillai, Ursula

    2014-04-15

    Placenta derived human amniotic epithelial cells (hAEC) are an attractive source of stem cells for the generation of hepatocyte-like cells (HLC) for therapeutic applications to treat liver diseases. During hAEC differentiation into HLC, they become increasingly immunogenic, which may result in immune cell-mediated rejection upon transplantation into allogeneic recipients. Placing cells within devices such as alginate microcapsules can prevent immune cell-mediated rejection. The aim of this study was to investigate the characteristics of HLC generated from hAEC and to examine the effects of encapsulation on HLC viability, gene expression, and function. hAEC were differentiated for 4 weeks and evaluated for hepatocyte-specific gene expression and function. Differentiated cells were encapsulated in barium alginate microcapsules and cultured for 7 days and the effect of encapsulation on cell viability, function, and hepatocyte related gene expression was determined. Differentiated cells performed key functions of hepatocytes including urea synthesis, drug-metabolizing cytochrome P450 (CYP)3A4 activity, indocyanine green (ICG) uptake, low-density lipoprotein (LDL) uptake, and exhibited glutathione antioxidant capacity. A number of hepatocyte-related genes involved in fat, cholesterol, bile acid synthesis, and xenobiotic metabolism were also expressed showing that the hAEC had differentiated into HLC. Upon encapsulation, the HLC remained viable for at least 7 days in culture, continued to express genes involved in fat, cholesterol, bile acid, and xenobiotic metabolism and had glutathione antioxidant capacity. CYP3A4 activity and urea synthesis by the encapsulated HLC were higher than that of monolayer HLC cultures. Functional HLC can be derived from hAEC, and HLC can be encapsulated within alginate microcapsules without losing viability or function in vitro. PMID:24295364

  9. Non-integrating episomal plasmid-based reprogramming of human amniotic fluid stem cells into induced pluripotent stem cells in chemically defined conditions.

    PubMed

    Slamecka, Jaroslav; Salimova, Lilia; McClellan, Steven; van Kelle, Mathieu; Kehl, Debora; Laurini, Javier; Cinelli, Paolo; Owen, Laurie; Hoerstrup, Simon P; Weber, Benedikt

    2016-01-01

    Amniotic fluid stem cells (AFSC) represent an attractive potential cell source for fetal and pediatric cell-based therapies. However, upgrading them to pluripotency confers refractoriness toward senescence, higher proliferation rate and unlimited differentiation potential. AFSC were observed to rapidly and efficiently reacquire pluripotency which together with their easy recovery makes them an attractive cell source for reprogramming. The reprogramming process as well as the resulting iPSC epigenome could potentially benefit from the unspecialized nature of AFSC. iPSC derived from AFSC also have potential in disease modeling, such as Down syndrome or β-thalassemia. Previous experiments involving AFSC reprogramming have largely relied on integrative vector transgene delivery and undefined serum-containing, feeder-dependent culture. Here, we describe non-integrative oriP/EBNA-1 episomal plasmid-based reprogramming of AFSC into iPSC and culture in fully chemically defined xeno-free conditions represented by vitronectin coating and E8 medium, a system that we found uniquely suited for this purpose. The derived AF-iPSC lines uniformly expressed a set of pluripotency markers Oct3/4, Nanog, Sox2, SSEA-1, SSEA-4, TRA-1-60, TRA-1-81 in a pattern typical for human primed PSC. Additionally, the cells formed teratomas, and were deemed pluripotent by PluriTest, a global expression microarray-based in-silico pluripotency assay. However, we found that the PluriTest scores were borderline, indicating a unique pluripotent signature in the defined condition. In the light of potential future clinical translation of iPSC technology, non-integrating reprogramming and chemically defined culture are more acceptable.

  10. Hepatocyte-Like Cells Derived from Human Amniotic Epithelial Cells Can Be Encapsulated Without Loss of Viability or Function In Vitro

    PubMed Central

    Vaghjiani, Vijesh; Vaithilingam, Vijayaganapathy; Saraswati, Indah; Sali, Adnan; Murthi, Padma; Kalionis, Bill; Tuch, Bernard E.

    2014-01-01

    Placenta derived human amniotic epithelial cells (hAEC) are an attractive source of stem cells for the generation of hepatocyte-like cells (HLC) for therapeutic applications to treat liver diseases. During hAEC differentiation into HLC, they become increasingly immunogenic, which may result in immune cell-mediated rejection upon transplantation into allogeneic recipients. Placing cells within devices such as alginate microcapsules can prevent immune cell-mediated rejection. The aim of this study was to investigate the characteristics of HLC generated from hAEC and to examine the effects of encapsulation on HLC viability, gene expression, and function. hAEC were differentiated for 4 weeks and evaluated for hepatocyte-specific gene expression and function. Differentiated cells were encapsulated in barium alginate microcapsules and cultured for 7 days and the effect of encapsulation on cell viability, function, and hepatocyte related gene expression was determined. Differentiated cells performed key functions of hepatocytes including urea synthesis, drug-metabolizing cytochrome P450 (CYP)3A4 activity, indocyanine green (ICG) uptake, low-density lipoprotein (LDL) uptake, and exhibited glutathione antioxidant capacity. A number of hepatocyte-related genes involved in fat, cholesterol, bile acid synthesis, and xenobiotic metabolism were also expressed showing that the hAEC had differentiated into HLC. Upon encapsulation, the HLC remained viable for at least 7 days in culture, continued to express genes involved in fat, cholesterol, bile acid, and xenobiotic metabolism and had glutathione antioxidant capacity. CYP3A4 activity and urea synthesis by the encapsulated HLC were higher than that of monolayer HLC cultures. Functional HLC can be derived from hAEC, and HLC can be encapsulated within alginate microcapsules without losing viability or function in vitro. PMID:24295364

  11. The immunomodulatory activity of human amniotic fluid can be correlated with transforming growth factor-beta 1 (TGF-beta 1) and beta 2 activity.

    PubMed Central

    Lang, A K; Searle, R F

    1994-01-01

    The role of alphafetoprotein (AFP) in the immunomodulatory activity of amniotic fluids (AF) from normally progressing human pregnancy (weeks 14-16) was investigated. A panel of 42 AF (25% v/v) reduced significantly phytohaemagglutinin (PHA)-induced peripheral blood mononuclear cell (PBMC) proliferation in serum-free cultures with a mean per cent inhibition of 68.4 +/- 5.5%. In contrast, AFP preparations, with one exception (U.AFP), failed to display inhibitory activity. Pretreatment of AF with anti-TGF-beta 1 and beta 2 antibodies used alone resulted in the mean per cent loss of inhibition of 33.1 +/- 3.9% and 52.3 +/- 7.5%, respectively. A summative loss of AF-mediated inhibition was detected when anti-TGF-beta 1 and beta 2 antibodies were used in combination, but immunomodulation was rarely abolished 100% by this treatment. Anti-TGF-beta 2 antibody treatment, unlike anti-TGF-beta 1 antibody treatment, reversed the inhibitory activity of U.AFP. The amount of TGF-beta 1 and beta 2 contained in human AF was studied by growth inhibition of Mv1 Lu cells. The mean levels of TGF-beta 1 and beta 2 in AF were 11 +/- 0.9 U/ml and 2.3 +/- 0.4 U/ml, respectively, which corresponds with a mean per cent inhibition of 49 +/- 4.7%. U.AFP also significantly inhibited Mv1 Lu cell growth. To investigate the mechanism of AF-mediated inhibition, the effect of AF and AFP on IL-2 production by concanavalin A (Con A)-stimulated PBMC blasts was determined by the CTLL-2 cell bioassay. IL-2 production was reduced 55.5% in AF-treated blasts and 61% in U.AFP-treated blasts compared with controls. Our findings indicate that the immunomodulatory activity of human AF can be correlated with TGF-beta 1 and beta 2 and not with AFP, the inhibitory activity of U.AFP preparation reflecting copurifying TGF-beta 2 activity. PMID:7518368

  12. In vivo Quantification of the Effects of Radiation and Presence of Hair Follicle Pores on the Proliferation of Fibroblasts in an Acellular Human Dermis in a Dorsal Skinfold Chamber: Relevance for Tissue Reconstruction following Neoadjuvant Therapy

    PubMed Central

    Maier, Patrick; Hohenberger, Peter; Roessner, Eric Dominic

    2015-01-01

    Introduction In neoadjuvant therapy, irradiation has a deleterious effect on neoangiogenesis. The aim of this study was to examine the post-implantation effects of neoadjuvant irradiation on the survival and proliferation of autologous cells seeded onto an acellular human dermis (hAD; Epiflex). Additionally, we examined the influence of dermal hair follicle pores on viability and proliferation. We used dorsal skinfold chambers implanted in rats and in-situ microscopy to quantify cell numbers over 9 days. Methods 24 rats received a skinfold chamber and were divided into 2 main groups; irradiated and unirradiated. In the irradiated groups 20Gy were applied epicutaneously at the dorsum. Epiflex pieces were cut to size 5x5mm such that each piece had either one or more visible hair follicle pores, or no such visible pores. Fibroblasts were transduced lentiviral with a fluorescent protein for cell tracking. Matrices were seeded statically with 2.5x104 fluorescent fibroblasts and implanted into the chambers. In each of the two main groups, half of the rats received Epiflex with hair follicle pores and half received Epiflex without pores. Scaffolds were examined in-situ at 0, 3, 6 and 9 days after transplantation. Visible cells on the surface were quantified using ImageJ. Results In all groups cell numbers were decreased on day 3. A treatment-dependent increase in cell numbers was observed at subsequent time points. Irradiation had an adverse effect on cell survival and proliferation. The number of cells detected in both irradiated and non-irradiated subjects was increased in those subjects that received transplants with hair follicle pores. Discussion This in-vivo study confirms that radiation negatively affects the survival and proliferation of fibroblasts seeded onto a human dermis transplant. The presence of hair follicle pores in the dermis transplants is shown to have a positive effect on cell survival and proliferation even in irradiated subjects. PMID:25955842

  13. CD117+ amniotic fluid stem cells

    PubMed Central

    Cananzi, Mara; De Coppi, Paolo

    2012-01-01

    Broadly multipotent stem cells can be isolated from amniotic fluid by selection for the expression of the membrane stem cell factor receptor c-Kit, a common marker for multipotential stem cells. They have clonogenic capability and can be directed into a wide range of cell types representing the three primary embryonic lineages. Amniotic fluid stem cells maintained for over 250 population doublings retained long telomeres and a normal karyotype. Clonal human lines verified by retroviral marking were induced to differentiate into cell types representing each embryonic germ layer, including cells of adipogenic, osteogenic, myogenic, endothelial, neuronal and hepatic lineages. AFS cells could be differentiate toward cardiomyogenic lineages, when co-cultured with neonatal cardiomyocytes, and have the potential to generate myogenic and hematopoietic lineages both in vitro and in vivo. Very recently first trimester AFS cells could be reprogrammed without any genetic manipulation opening new possibilities in the field of fetal/neonatal therapy and disease modeling. In this review we are aiming to summarize the knowledge on amniotic fluid stem cells and highlight the most promising results. PMID:23037870

  14. In vitro angiogenesis on the human amniotic membrane: requirement for basic fibroblast growth factor-induced proteinases

    PubMed Central

    1989-01-01

    The role of basic fibroblast growth factor-(bFGF) induced proteinases in basement membrane (BM) invasion by bovine capillary endothelial (BCE) cells was studied using a quantitative in vitro assay previously described (Mignatti et al., 1986). 125I-iododeoxyuridine-labeled BCE cells were grown for 72 h on the human amnion BM, and cell invasion was determined by measuring the radioactivity associated with the tissue after removal of the noninvasive cell layer. BCE cells were noninvasive under normal conditions. Addition of human bFGF to either the BM or to the stromal aspect of the amnion induced BCE cell invasion with a dose- dependent response. This effect was maximal in the presence of 70 ng/ml bFGF, and was inhibited by anti-FGF antibody. Transforming growth factor beta, as well as plasmin inhibitors and anti-tissue type plasminogen activator antibody inhibited BCE cell invasion. The tissue inhibitor of metalloproteinases, 1-10 phenanthroline, anti-type IV and anti-interstitial collagenase antibodies had the same effect. On the contrary, anti-stromelysin antibody and Eglin, an inhibitor of elastase, were ineffective. The results obtained show that both the plasminogen activator-plasmin system and specific collagenases are involved in the invasive process occurring during angiogenesis. PMID:2465298

  15. Ethmocephaly with amniotic band syndrome.

    PubMed

    Das, Gobinda; Gayen, Sibnath; Bandyopadhyay, Sabyasachi; Das, Debabrata

    2012-10-01

    Ethmocephaly is the rarest form of holoprosencephaly, which occurs due to an incomplete cleavage of the forebrain. Clinically, the disease presents with a proboscis, hypotelorism, microphthalmos and malformed ears. Amniotic band syndrome is another rare congenital malformation with ring-like constriction bands in the limbs, head, face or trunk. We present a case of ethmocephaly with amniotic band syndrome, which is likely the first of its kind, published in the literature. PMID:23248551

  16. High aminopeptidase N/CD13 levels characterize human amniotic mesenchymal stem cells and drive their increased adipogenic potential in obese women.

    PubMed

    Iaffaldano, Laura; Nardelli, Carmela; Raia, Maddalena; Mariotti, Elisabetta; Ferrigno, Maddalena; Quaglia, Filomena; Labruna, Giuseppe; Capobianco, Valentina; Capone, Angela; Maruotti, Giuseppe Maria; Pastore, Lucio; Di Noto, Rosa; Martinelli, Pasquale; Sacchetti, Lucia; Del Vecchio, Luigi

    2013-08-15

    Maternal obesity is associated to increased fetal risk of obesity and other metabolic diseases. Human amniotic mesenchymal stem cells (hA-MSCs) have not been characterized in obese women. The aim of this study was to isolate and compare hA-MSC immunophenotypes from obese (Ob-) and normal weight control (Co-) women, to identify alterations possibly predisposing the fetus to obesity. We enrolled 16 Ob- and 7 Co-women at delivery (mean/SEM prepregnancy body mass index: 40.3/1.8 and 22.4/1.0 kg/m2, respectively), and 32 not pregnant women. hA-MSCs were phenotyped by flow cytometry; several maternal and newborn clinical and biochemical parameters were also measured. The expression of membrane antigen CD13 was higher on Ob-hA-MSCs than on Co-hA-MSCs (P = 0.005). Also, serum levels of CD13 at delivery were higher in Ob- versus Co-pregnant women and correlated with CD13 antigen expression on Ob-hA-MSCs (r2 = 0.84, P < 0.0001). Adipogenesis induction experiments revealed that Ob-hA-MSCs had a higher adipogenic potential than Co-hA-MSCs as witnessed by higher peroxisome proliferator-activated receptor gamma and aP2 mRNA levels (P = 0.05 and P = 0.05, respectively), at postinduction day 14 associated with increased CD13 mRNA levels from baseline to day 4 postinduction (P < 0.05). Adipogenesis was similar in the two sets of hA-MSCs after CD13 silencing, whereas it was increased in Co-hA-MSCs after CD13 overexpression. CD13 expression was high also in Ob-h-MSCs from umbilical cords or visceral adipose tissue of not pregnant women. In conclusion, antigen CD13, by influencing the adipogenic potential of hA-MSCs, could be an in utero risk factor for obesity. Our data strengthen the hypothesis that high levels of serum and MSC CD13 are obesity markers.

  17. High Aminopeptidase N/CD13 Levels Characterize Human Amniotic Mesenchymal Stem Cells and Drive Their Increased Adipogenic Potential in Obese Women

    PubMed Central

    Iaffaldano, Laura; Nardelli, Carmela; Raia, Maddalena; Mariotti, Elisabetta; Ferrigno, Maddalena; Quaglia, Filomena; Labruna, Giuseppe; Capobianco, Valentina; Capone, Angela; Maruotti, Giuseppe Maria; Pastore, Lucio; Di Noto, Rosa; Martinelli, Pasquale; Del Vecchio, Luigi

    2013-01-01

    Maternal obesity is associated to increased fetal risk of obesity and other metabolic diseases. Human amniotic mesenchymal stem cells (hA-MSCs) have not been characterized in obese women. The aim of this study was to isolate and compare hA-MSC immunophenotypes from obese (Ob-) and normal weight control (Co-) women, to identify alterations possibly predisposing the fetus to obesity. We enrolled 16 Ob- and 7 Co-women at delivery (mean/SEM prepregnancy body mass index: 40.3/1.8 and 22.4/1.0 kg/m2, respectively), and 32 not pregnant women. hA-MSCs were phenotyped by flow cytometry; several maternal and newborn clinical and biochemical parameters were also measured. The expression of membrane antigen CD13 was higher on Ob-hA-MSCs than on Co-hA-MSCs (P=0.005). Also, serum levels of CD13 at delivery were higher in Ob- versus Co-pregnant women and correlated with CD13 antigen expression on Ob-hA-MSCs (r2=0.84, P<0.0001). Adipogenesis induction experiments revealed that Ob-hA-MSCs had a higher adipogenic potential than Co-hA-MSCs as witnessed by higher peroxisome proliferator-activated receptor gamma and aP2 mRNA levels (P=0.05 and P=0.05, respectively), at postinduction day 14 associated with increased CD13 mRNA levels from baseline to day 4 postinduction (P<0.05). Adipogenesis was similar in the two sets of hA-MSCs after CD13 silencing, whereas it was increased in Co-hA-MSCs after CD13 overexpression. CD13 expression was high also in Ob-h-MSCs from umbilical cords or visceral adipose tissue of not pregnant women. In conclusion, antigen CD13, by influencing the adipogenic potential of hA-MSCs, could be an in utero risk factor for obesity. Our data strengthen the hypothesis that high levels of serum and MSC CD13 are obesity markers. PMID:23488598

  18. Amniotic fluid index: correlation with amniotic fluid volume.

    PubMed

    Hoskins, I A; McGovern, P G; Ordorica, S A; Frieden, F J; Young, B K

    1992-01-01

    We calculated the amniotic fluid indexes (AFIs) of 310 women on 459 occasions. Normative data were analyzed and compared with data in several high-risk groups. In the normal gestations there was a progressive increase in AFI with advancing gestation until 32 weeks, after which there was a decline. The mean AFIs in abnormal gestations varied with the clinical diagnoses. These values were compared to those obtained by assessing amniotic fluid volume (AFV), that is a pocket more than 2 cm. There were 51 patients with abnormal AFVs. Forty-two had decreased fluid, six also had decreased AFIs; nine had increased AFVs and five (all with diabetes) also had increased AFIs. Thus, AFIs in normal pregnancies showed an orderly pattern of change with gestational age, and there was no accurate correlation between AFI and AFV. Thus, using AFV alone may lead to false interpretations of amniotic fluid status. PMID:1418123

  19. Acellular dermal matrices in breast reconstructions - a literature review.

    PubMed

    Skovsted Yde, Simon; Brunbjerg, Mette Eline; Damsgaard, Tine Engberg

    2016-08-01

    During the last two decades, acellular dermal matrices (ADM) have been more widely used in reconstructive procedures i.e. breast reconstructions. Several, both synthetic and biologic products derived from human, porcine and bovine tissue, have been introduced. Until this point postoperative complications for the acellular dermal matrices, as a group, have been the main focus. The purpose of this literature review is to summarize the current knowledge on the each biologic product used in breast reconstructions, including product specific complication frequencies. A systematic search of the literature was performed in the PubMed and EMBASE databases, identifying 55 relevant articles, mainly evidence level III. AlloDerm seems to be associated with severe complicating matters in the reconstructive process compared to other products. This could be due to the higher number of investigating studies relative to the others. The surgical area faces certain challenges comparing results, due to surgical variance, the data collection and follow-up. More well-defined guidelines and more high-evidence randomized studies could increase the overall level of evidence in this area. PMID:26881927

  20. Cartilage oligomeric matrix protein enhances the vascularization of acellular nerves

    PubMed Central

    Cui, Wei-ling; Qiu, Long-hai; Lian, Jia-yan; Li, Jia-chun; Hu, Jun; Liu, Xiao-lin

    2016-01-01

    Vascularization of acellular nerves has been shown to contribute to nerve bridging. In this study, we used a 10-mm sciatic nerve defect model in rats to determine whether cartilage oligomeric matrix protein enhances the vascularization of injured acellular nerves. The rat nerve defects were treated with acellular nerve grafting (control group) alone or acellular nerve grafting combined with intraperitoneal injection of cartilage oligomeric matrix protein (experimental group). As shown through two-dimensional imaging, the vessels began to invade into the acellular nerve graft from both anastomotic ends at day 7 post-operation, and gradually covered the entire graft at day 21. The vascular density, vascular area, and the velocity of revascularization in the experimental group were all higher than those in the control group. These results indicate that cartilage oligomeric matrix protein enhances the vascularization of acellular nerves. PMID:27127495

  1. Early mesozoic coexistence of amniotes and hepadnaviridae.

    PubMed

    Suh, Alexander; Weber, Claudia C; Kehlmaier, Christian; Braun, Edward L; Green, Richard E; Fritz, Uwe; Ray, David A; Ellegren, Hans

    2014-12-01

    Hepadnaviridae are double-stranded DNA viruses that infect some species of birds and mammals. This includes humans, where hepatitis B viruses (HBVs) are prevalent pathogens in considerable parts of the global population. Recently, endogenized sequences of HBVs (eHBVs) have been discovered in bird genomes where they constitute direct evidence for the coexistence of these viruses and their hosts from the late Mesozoic until present. Nevertheless, virtually nothing is known about the ancient host range of this virus family in other animals. Here we report the first eHBVs from crocodilian, snake, and turtle genomes, including a turtle eHBV that endogenized >207 million years ago. This genomic "fossil" is >125 million years older than the oldest avian eHBV and provides the first direct evidence that Hepadnaviridae already existed during the Early Mesozoic. This implies that the Mesozoic fossil record of HBV infection spans three of the five major groups of land vertebrates, namely birds, crocodilians, and turtles. We show that the deep phylogenetic relationships of HBVs are largely congruent with the deep phylogeny of their amniote hosts, which suggests an ancient amniote-HBV coexistence and codivergence, at least since the Early Mesozoic. Notably, the organization of overlapping genes as well as the structure of elements involved in viral replication has remained highly conserved among HBVs along that time span, except for the presence of the X gene. We provide multiple lines of evidence that the tumor-promoting X protein of mammalian HBVs lacks a homolog in all other hepadnaviruses and propose a novel scenario for the emergence of X via segmental duplication and overprinting of pre-existing reading frames in the ancestor of mammalian HBVs. Our study reveals an unforeseen host range of prehistoric HBVs and provides novel insights into the genome evolution of hepadnaviruses throughout their long-lasting association with amniote hosts. PMID:25501991

  2. [Antibacterial and antimycotic activity of the amniotic fluid against selected germs (author's transl)].

    PubMed

    Jankowski, R P; Rauskolb, R; Gupta, K G

    1977-03-01

    60 samples of amniotic fluid from 60 patients were obtained between 14 and 42 weeks gestation by dates and tested for their antibacterial activity against staphlococcus aureus strains and brucella abortus and their antimycotic activity against Candida albicans. The antibacterial and antimycotic activity of the amniotic fluid was confirmed. This activity increases steadily from the 15th week and reaches its maximum around term between 36 and 42 weeks. The amniotic samples obtained at 20 weeks gestation showed a weak antibacterial activity especially against staphlococcus aureus of human origin. The above results are important because of the increasing frequency of early and late diagnostic amniocentesis in pregnancy.

  3. Neurogenic differentiation of amniotic fluid stem cells.

    PubMed

    Rosner, M; Mikula, M; Preitschopf, A; Feichtinger, M; Schipany, K; Hengstschläger, M

    2012-05-01

    In 2003, human amniotic fluid has been shown to contain stem cells expressing Oct-4, a marker for pluripotency. This finding initiated a rapidly growing and very promising new stem cell research field. Since then, amniotic fluid stem (AFS) cells have been demonstrated to harbour the potential to differentiate into any of the three germ layers and to form three-dimensional aggregates, so-called embryoid bodies, known as the principal step in the differentiation of pluripotent stem cells. Marker selection and minimal dilution approaches allow the establishment of monoclonal AFS cell lineages with high proliferation potential. AFS cells have a lower risk for tumour development and do not raise the ethical issues of embryonic stem cells. Compared to induced pluripotent stem cells, AFS cells do not need exogenic treatment to induce pluripotency, are chromosomal stable and do not harbour the epigenetic memory and accumulated somatic mutations of specific differentiated source cells. Compared to adult stem cells, AFS can be grown in larger quantities and show higher differentiation potential. Accordingly, in the recent past, AFS became increasingly accepted as an optimal tool for basic research and probably also for specific cell-based therapies. Here, we review the current knowledge on the neurogenic differentiation potential of AFS cells.

  4. Acellular ostrich corneal stroma used as scaffold for construction of tissue-engineered cornea

    PubMed Central

    Liu, Xian-Ning; Zhu, Xiu-Ping; Wu, Jie; Wu, Zheng-Jie; Yin, Yong; Xiao, Xiang-Hua; Su, Xin; Kong, Bin; Pan, Shi-Yin; Yang, Hua; Cheng, Yan; An, Na; Mi, Sheng-Li

    2016-01-01

    AIM To assess acellular ostrich corneal matrix used as a scaffold to reconstruct a damaged cornea. METHODS A hypertonic saline solution combined with a digestion method was used to decellularize the ostrich cornea. The microstructure of the acellular corneal matrix was observed by transmission electron microscopy (TEM) and hematoxylin and eosin (H&E) staining. The mechanical properties were detected by a rheometer and a tension machine. The acellular corneal matrix was also transplanted into a rabbit cornea and cytokeratin 3 was used to check the immune phenotype. RESULTS The microstructure and mechanical properties of the ostrich cornea were well preserved after the decellularization process. In vitro, the methyl thiazolyl tetrazolium results revealed that extracts of the acellular ostrich corneas (AOCs) had no inhibitory effects on the proliferation of the corneal epithelial or endothelial cells or on the keratocytes. The rabbit lamellar keratoplasty showed that the transplanted AOCs were transparent and completely incorporated into the host cornea while corneal turbidity and graft dissolution occurred in the acellular porcine cornea (APC) transplantation. The phenotype of the reconstructed cornea was similar to a normal rabbit cornea with a high expression of cytokeratin 3 in the superficial epithelial cell layer. CONCLUSION We first used AOCs as scaffolds to reconstruct damaged corneas. Compared with porcine corneas, the anatomical structures of ostrich corneas are closer to those of human corneas. In accordance with the principle that structure determines function, a xenograft lamellar keratoplasty also confirmed that the AOC transplantation generated a superior outcome compared to that of the APC graft. PMID:27158598

  5. Acellular dermal matrix in abdominal wall reconstruction.

    PubMed

    Silverman, Ronald P

    2011-09-01

    Abdominal wall reconstruction is a complex and challenging surgical undertaking. While permanent prosthetic mesh is considered the gold standard for minimizing hernia recurrence, placement of synthetic mesh is sometimes imprudent due to contamination or risk of infection. Acellular dermal matrices (ADM) offer an exciting biologic alternative. This article provides a historical perspective on the evolution of complex ventral hernia repair leading up to and including the placement of ADM, an explanation of the biology of ADM as it relates to ventral hernia repair, and a description of the current indications, techniques, benefits, and shortcomings of its use in the abdominal wall.

  6. [Amniotic fluid embolism: an update].

    PubMed

    Legrand, M; Rossignol, M; Muller, F; Payen, D

    2013-03-01

    Amniotic fluid embolism (AFE) results from the passage of fœtal and amniotic fragments into the maternal circulation, occurring mostly within minutes before or after delivery. Although maternal and fœtal mortality of AFE remains high (about 40%), AFE should no longer be considered as having an ineluctable fatal course. Diagnosis is often made upon clinical presentation but histological confirmation is difficult owing favorable outcome and because an autopsy has not been performed. Identification of squamous cells in the maternal circulation could not confirm the diagnosis because of their possible maternal origin. High plasma level of insulin-like growth factor-binding protein-1 (IGFBP-1) has recently been identified as a biomarker of amniotic fluid passage into the maternal circulation and might therefore be used to confirm the diagnosis when lung tissue histology is not available. Treatment of AFE remains supportive with a special focus on correction of the coagulopathy and search for acute core pulmonale. In this later case, physicians should consider initiating an extracorporeal life support when facing a patient with refractory shock. Finally, caution is needed with the use of recombinant factor VIIa in this context. PMID:23422343

  7. Amniotic fluid: the use of high-dimensional biology to understand fetal well-being.

    PubMed

    Kamath-Rayne, Beena D; Smith, Heather C; Muglia, Louis J; Morrow, Ardythe L

    2014-01-01

    Our aim was to review the use of high-dimensional biology techniques, specifically transcriptomics, proteomics, and metabolomics, in amniotic fluid to elucidate the mechanisms behind preterm birth or assessment of fetal development. We performed a comprehensive MEDLINE literature search on the use of transcriptomic, proteomic, and metabolomic technologies for amniotic fluid analysis. All abstracts were reviewed for pertinence to preterm birth or fetal maturation in human subjects. Nineteen articles qualified for inclusion. Most articles described the discovery of biomarker candidates, but few larger, multicenter replication or validation studies have been done. We conclude that the use of high-dimensional systems biology techniques to analyze amniotic fluid has significant potential to elucidate the mechanisms of preterm birth and fetal maturation. However, further multicenter collaborative efforts are needed to replicate and validate candidate biomarkers before they can become useful tools for clinical practice. Ideally, amniotic fluid biomarkers should be translated to a noninvasive test performed in maternal serum or urine.

  8. Isolation and characterization of canine amniotic membrane-derived multipotent stem cells.

    PubMed

    Park, Sang-Bum; Seo, Min-Soo; Kim, Hyung-Sik; Kang, Kyung-Sun

    2012-01-01

    Recent studies have shown that amniotic membrane tissue is a rich source of stem cells in humans. In clinical applications, the amniotic membrane tissue had therapeutic effects on wound healing and corneal surface reconstruction. Here, we successfully isolated and identified multipotent stem cells (MSCs) from canine amniotic membrane tissue. We cultured the canine amniotic membrane-derived multipotent stem cells (cAM-MSCs) in low glucose DMEM medium. cAM-MSCs have a fibroblast-like shape and adhere to tissue culture plastic. We characterized the immunophenotype of cAM-MSCs by flow cytometry and measured cell proliferation by the cumulative population doubling level (CPDL). We performed differentiation studies for the detection of trilineage multipotent ability, under the appropriate culture conditions. Taken together, our results show that cAM-MSCs could be a rich source of stem cells in dogs. Furthermore, cAM-MSCs may be useful as a cell therapy application for veterinary regenerative medicine.

  9. Prenatal diagnosis of amniotic band syndrome

    PubMed Central

    Padmanabhan, Laxmi Devi; Hamza, Zareena V; Thampi, Madhavan Venugopalan; Nampoothiri, Sheela

    2016-01-01

    Amniotic band can cause a broad spectrum of anomalies ranging from simple band constrictions to major craniofacial and visceral defects. It can cause significant neonatal morbidity. Accurate diagnosis will help in the management of the present pregnancy and in counseling with regard to future pregnancies. Here we report three cases of amniotic band syndrome detected in the prenatal period. PMID:27081225

  10. Prenatal diagnosis of amniotic band syndrome.

    PubMed

    Padmanabhan, Laxmi Devi; Hamza, Zareena V; Thampi, Madhavan Venugopalan; Nampoothiri, Sheela

    2016-01-01

    Amniotic band can cause a broad spectrum of anomalies ranging from simple band constrictions to major craniofacial and visceral defects. It can cause significant neonatal morbidity. Accurate diagnosis will help in the management of the present pregnancy and in counseling with regard to future pregnancies. Here we report three cases of amniotic band syndrome detected in the prenatal period.

  11. Early Mesozoic Coexistence of Amniotes and Hepadnaviridae

    PubMed Central

    Suh, Alexander; Weber, Claudia C.; Kehlmaier, Christian; Braun, Edward L.; Green, Richard E.; Fritz, Uwe; Ray, David A.; Ellegren, Hans

    2014-01-01

    Hepadnaviridae are double-stranded DNA viruses that infect some species of birds and mammals. This includes humans, where hepatitis B viruses (HBVs) are prevalent pathogens in considerable parts of the global population. Recently, endogenized sequences of HBVs (eHBVs) have been discovered in bird genomes where they constitute direct evidence for the coexistence of these viruses and their hosts from the late Mesozoic until present. Nevertheless, virtually nothing is known about the ancient host range of this virus family in other animals. Here we report the first eHBVs from crocodilian, snake, and turtle genomes, including a turtle eHBV that endogenized >207 million years ago. This genomic “fossil” is >125 million years older than the oldest avian eHBV and provides the first direct evidence that Hepadnaviridae already existed during the Early Mesozoic. This implies that the Mesozoic fossil record of HBV infection spans three of the five major groups of land vertebrates, namely birds, crocodilians, and turtles. We show that the deep phylogenetic relationships of HBVs are largely congruent with the deep phylogeny of their amniote hosts, which suggests an ancient amniote–HBV coexistence and codivergence, at least since the Early Mesozoic. Notably, the organization of overlapping genes as well as the structure of elements involved in viral replication has remained highly conserved among HBVs along that time span, except for the presence of the X gene. We provide multiple lines of evidence that the tumor-promoting X protein of mammalian HBVs lacks a homolog in all other hepadnaviruses and propose a novel scenario for the emergence of X via segmental duplication and overprinting of pre-existing reading frames in the ancestor of mammalian HBVs. Our study reveals an unforeseen host range of prehistoric HBVs and provides novel insights into the genome evolution of hepadnaviruses throughout their long-lasting association with amniote hosts. PMID:25501991

  12. Spectral luminescence analysis of amniotic fluid

    NASA Astrophysics Data System (ADS)

    Slobozhanina, Ekaterina I.; Kozlova, Nataly M.; Kasko, Leonid P.; Mamontova, Marina V.; Chernitsky, Eugene A.

    1997-12-01

    It is shown that the amniotic fluid has intensive ultra-violet luminescence caused by proteins. Along with it amniotic fluid radiated in the field of 380 - 650 nm with maxima at 430 - 450 nm and 520 - 560 nm. The first peak of luminescence ((lambda) exc equals 350 nm; (lambda) em equals 430 - 440 nm) is caused (most probably) by the presence in amniotic fluid of some hormones, NADH2 and NADPH2. A more long-wave component ((lambda) exc equals 460 nm; (lambda) em equals 520 - 560 nm) is most likely connected with the presence in amniotic fluid pigments (bilirubin connected with protein and other). It is shown that intensity and maximum of ultra-violet luminescence spectra of amniotic fluid in normality and at pathology are identical. However both emission spectra and excitation spectra of long-wave ((lambda) greater than 450 nm) luminescence of amniotic fluid from pregnant women with such prenatal abnormal developments of a fetus as anencephaly and spina bifida are too long-wave region in comparison with the norm. Results of research testify that spectral luminescent analysis of amniotic fluid can be used for screening of malformations of the neural tube. It is very difficult for a practical obstetrician to reveal pregnant women with a high risk of congenital malformations of the fetus. Apart from ultrasonic examination, cytogenetic examination of amniotic fluid and defumination of concentrations of alpha-fetoprotein and acetylcholin-esterases in the amniotic fluid and blood plasma are the most widely used diagnostic approaches. However, biochemical and cytogenetic diagnostic methods are time-consuming. In the present work spectral luminescence properties of the amniotic fluid are investigated to determine spectral parameters that can be used to reveal pregnant women with a high risk of congenital malformations of their offsprings.

  13. 21 CFR 884.1550 - Amniotic fluid sampler (amniocentesis tray).

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Amniotic fluid sampler (amniocentesis tray). 884... Diagnostic Devices § 884.1550 Amniotic fluid sampler (amniocentesis tray). (a) Identification. The amniotic fluid sampler (amniocentesis tray) is a collection of devices used to aspirate amniotic fluid from...

  14. 21 CFR 884.1550 - Amniotic fluid sampler (amniocentesis tray).

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Amniotic fluid sampler (amniocentesis tray). 884... Diagnostic Devices § 884.1550 Amniotic fluid sampler (amniocentesis tray). (a) Identification. The amniotic fluid sampler (amniocentesis tray) is a collection of devices used to aspirate amniotic fluid from...

  15. 21 CFR 884.1550 - Amniotic fluid sampler (amniocentesis tray).

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Amniotic fluid sampler (amniocentesis tray). 884... Diagnostic Devices § 884.1550 Amniotic fluid sampler (amniocentesis tray). (a) Identification. The amniotic fluid sampler (amniocentesis tray) is a collection of devices used to aspirate amniotic fluid from...

  16. 21 CFR 884.1550 - Amniotic fluid sampler (amniocentesis tray).

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Amniotic fluid sampler (amniocentesis tray). 884... Diagnostic Devices § 884.1550 Amniotic fluid sampler (amniocentesis tray). (a) Identification. The amniotic fluid sampler (amniocentesis tray) is a collection of devices used to aspirate amniotic fluid from...

  17. 21 CFR 884.1550 - Amniotic fluid sampler (amniocentesis tray).

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Amniotic fluid sampler (amniocentesis tray). 884... Diagnostic Devices § 884.1550 Amniotic fluid sampler (amniocentesis tray). (a) Identification. The amniotic fluid sampler (amniocentesis tray) is a collection of devices used to aspirate amniotic fluid from...

  18. Proposed diagnostic criteria for the case definition of amniotic fluid embolism in research studies.

    PubMed

    Clark, Steven L; Romero, Roberto; Dildy, Gary A; Callaghan, William M; Smiley, Richard M; Bracey, Arthur W; Hankins, Gary D; D'Alton, Mary E; Foley, Mike; Pacheco, Luis D; Vadhera, Rakesh B; Herlihy, J Patrick; Berkowitz, Richard L; Belfort, Michael A

    2016-10-01

    Amniotic fluid embolism is a leading cause of maternal mortality in developed countries. Our understanding of risk factors, diagnosis, treatment, and prognosis is hampered by a lack of uniform clinical case definition; neither histologic nor laboratory findings have been identified unique to this condition. Amniotic fluid embolism is often overdiagnosed in critically ill peripartum women, particularly when an element of coagulopathy is involved. Previously proposed case definitions for amniotic fluid embolism are nonspecific, and when viewed through the eyes of individuals with experience in critical care obstetrics, would include women with a number of medical conditions much more common than amniotic fluid embolism. We convened a working group under the auspices of a committee of the Society for Maternal-Fetal Medicine and the Amniotic Fluid Embolism Foundation whose task was to develop uniform diagnostic criteria for the research reporting of amniotic fluid embolism. These criteria rely on the presence of the classic triad of hemodynamic and respiratory compromise accompanied by strictly defined disseminated intravascular coagulopathy. It is anticipated that limiting research reports involving amniotic fluid embolism to women who meet these criteria will enhance the validity of published data and assist in the identification of risk factors, effective treatments, and possibly useful biomarkers for this condition. A registry has been established in conjunction with the Perinatal Research Branch of the Eunice Kennedy Shriver National Institute of Child Health and Human Development to collect both clinical information and laboratory specimens of women with suspected amniotic fluid embolism in the hopes of identifying unique biomarkers of this condition.

  19. On the origin of amniotic stem cells: of mice and men.

    PubMed

    Dobreva, Mariya P; Pereira, Paulo N G; Deprest, Jan; Zwijsen, An

    2010-01-01

    A common characteristic of mammals is the development of extraembryonic supporting tissues and organs that are required for embryonic implantation, survival and development in utero. The amnion is the innermost extraembryonic membrane that eventually surrounds the fetus of amniotes, and contains the amniotic fluid. Next to its function in in utero development, the amnion has been shown to have an important potential for clinical applications. It is mainly used as a dressing to stimulate healing in skin and ocular wounds. Moreover, cells derived from the amniotic membrane and amniotic fluid have been reported to possess stem cell features, like pluripotent differentiation ability. Little is known about the early development of this membrane in humans. The mouse is a powerful genetic model organism that can be used to address the dynamics and the developmental origin of amnion and amnion-derived stem cells. Here, we discuss some fundamental differences in amnion development in the disc-shaped primate embryo and in the cup-shaped mouse embryo. We emphasize the consequences that this may have on the derivation of amniotic "stem" cells. After revision of the different isolation procedures of amniotic (fluid) derived "stem" cells from rodents, we reveal striking differences in the sources used to derive these cells across studies. The profound differences in the development of the extraembryonic membranes and cavities between primates and rodents may result in comparing cell types of different developmental origins, eventually leading to missinterpretations.

  20. Rat full term amniotic fluid harbors highly potent stem cells.

    PubMed

    Mun-Fun, Hoo; Ferdaos, Nurfarhana; Hamzah, Siti Nurusaadah; Ridzuan, Noridzzaida; Hisham, Nurul Afiqah; Abdullah, Syahril; Ramasamy, Rajesh; Cheah, Pike See; Thilakavathy, Karrupiah; Yazid, Mohd Nazri; Nordin, Norshariza

    2015-10-01

    Amniotic fluid stem cells (AFSCs) are commonly isolated from mid-term amniotic fluid (AF) of animals and human collected via an invasive technique, amniocentesis. Alternatively, AFSCs could be collected at full-term. However, it is unclear whether AFSCs are present in the AF at full term. Here, we aimed to isolate and characterize stem cells isolated from AF of full term pregnant rats. Three stem cell lines have been established following immuno-selection against the stem cell marker, c-kit. Two of the new lines expressed multiple markers of pluripotency until more than passage 90. Further, they spontaneously differentiated into derivatives of the three primary germ layers through the formation of good quality embryoid bodies (EBs), and can be directly differentiated into neural lineage. Their strong stemness and potent neurogenic properties highlight the presence of highly potent stem cells in AF of full-term pregnancies, which could serve as a potential source of stem cells for regenerative medicine.

  1. Tooth histology in the cretaceous ichthyosaur Platypterygius australis, and its significance for the conservation and divergence of mineralized tooth tissues in amniotes.

    PubMed

    Maxwell, Erin E; Caldwell, Michael W; Lamoureux, Denis O

    2011-02-01

    Ichthyosaurs are an extinct group of secondarily aquatic reptiles that show ligamentous tooth attachment to the jaw in some derived forms. Here, we provide a modern description of tooth histology in ichthyosaurs, using Platypterygius australis, a large ichthyosaur from the Cretaceous of Australia. Our study supports evolutionary conservation of the principal mineralized tooth tissue types in amniotes with ligamentous tooth attachment: enamel, dentine, cellular, and acellular cementum. This is the first time that the latter tissue has been located in ichthyosaurs. Vascularized cementum (osteocementum) is reduced or absent in amniotes in which the teeth are ankylosed to the jaw bone, such as basal ichthyosaurs, and raises questions regarding the function of this tissue and the potential developmental or selective conditions leading to its convergent evolution. PMID:21210486

  2. Human second trimester amniotic fluid cells are able to create embryoid body-like structures in vitro and to show typical expression profiles of embryonic and primordial germ cells.

    PubMed

    Antonucci, Ivana; Di Pietro, Roberta; Alfonsi, Melissa; Centurione, Maria Antonietta; Centurione, Lucia; Sancilio, Silvia; Pelagatti, Francesca; D'Amico, Maria Angela; Di Baldassarre, Angela; Piattelli, Adriano; Tetè, Stefano; Palka, Giandomenico; Borlongan, Cesar V; Stuppia, Liborio

    2014-01-01

    Human amniotic fluid-derived stem cells (AFSCs) represent a novel class of broadly multipotent stem cells sharing characteristics of both embryonic and adult stem cells. However, both the origin of these cells and their actual properties in terms of pluripotent differentiation potential are still debated. In order to verify the presence of features of pluripotency in human second trimester AFSCs, we have investigated the ability of these cells to form in vitro three-dimensional aggregates, known as embryoid bodies (EBs), and to express specific genes of embryonic stem cells (ESCs) and primordial germ cells (PGCs). EBs were obtained after 5 days of AFSC culture in suspension and showed positivity for alkaline phosphatase (AP) staining and for specific markers of pluripotency (OCT4 and SOX2). Moreover, EB-derived cells showed the expression of specific transcripts of the three germ layers. RT-PCR analysis, carried out at different culture times (second, third, fourth, fifth, and eighth passages), revealed the presence of specific markers of ESCs (such as FGF4 and DAPPA4), as well as of markers typical of PGCs and, in particular, genes involved in early stages of germ cell development (Fragilis, Stella, Vasa, c-Kit, Rnf17). Finally, the expression of genes related to the control of DNA methylation (DNMT3A, DNMT3b1, DNMT1, DNMT3L, MBD1, MBD2, MBD3, MDB4, MeCP2), as well as the lack of inactivation of the X-chromosome in female samples, was also demonstrated. Taken together, these data provide further evidence for the presence of common features among human AFSCs, PGCs, and ESCs.

  3. Transplantation of amniotic membrane and limbal autograft for patients with recurrent pterygium associated with symblepharon

    PubMed Central

    Shimazaki, J.; Shinozaki, N.; Tsubota, K.

    1998-01-01

    AIM—Treatment of recurrent pterygium associated with symblepharon requires both suppression of fibrosis and reconstruction of limbal barrier. To achieve this, human amniotic membrane was transplanted and limbal autografts performed.
METHODS—Four patients with severe symblepharon resulting from multiple surgeries for pterygium were treated. Human amniotic membrane was obtained at caesarean section and preserved until surgery. After excision of the fibrous tissues, the amniotic membrane was placed on the sclera, and a limbal autograft transplantation was performed using limbal tissues taken from the affected eye.
RESULTS—Recurrence of symblepharon was not observed in any of the patients and significant suppression of the subconjunctival fibrosis was achieved. Ocular movement improved in all cases. Complete remission of pterygium regrowth occurred in three cases, and a slight (about 1 mm) recurrence occurred in one case. The limbal donor site showed the presence of mild depressions without the formation of pseudopterygium.
CONCLUSION—Transplantation of human amniotic membrane with a limbal autograft appears to be a promising surgical treatment for reconstructing the ocular surface in patients with recurrent pterygium associated with symblepharon.

 Keywords: pterygium; limbal transplantation; amniotic membrane; symblepharon PMID:9602618

  4. The presence of an embryonic opercular flap in amniotes.

    PubMed

    Richardson, Jo; Shono, Takanori; Okabe, Masataka; Graham, Anthony

    2012-01-22

    The operculum is a large flap consisting of several flat bones found on the side of the head of bony fish. During development, the opercular bones form within the second pharyngeal arch, which expands posteriorly and comes to cover the gill-bearing arches. With the evolution of the tetrapods and the assumption of a terrestrial lifestyle, it was believed that the operculum was lost. Here, we demonstrate that an embryonic operculum persists in amniotes and that its early development is homologous with that of teleosts. As in zebrafish, the second pharyngeal arch of the chick embryo grows disproportionately and comes to cover the posterior arches. We show that the developing second pharyngeal arch in both chick and zebrafish embryos express orthologous genes and require shh signalling for caudal expansion. In amniotes, however, the caudal edge of the expanded second arch fuses to the surface of the neck. We have detailed how this process occurs and also demonstrated a requirement for thyroid signalling here. Our results thus demonstrate the persistence of an embryonic opercular flap in amniotes, that its fusion mirrors aspects of amphibian metamorphosis and gives insights into the origin of branchial cleft anomalies in humans.

  5. Autophagic and apoptotic cell death in amniotic epithelial cells.

    PubMed

    Shen, Z-Y; Li, E-M; Lu, S-Q; Shen, J; Cai, Y-M; Wu, Y-E; Zheng, R-M; Tan, L-J; Xu, L-Y

    2008-11-01

    The aim of this paper is to determine if autophagic cell death is associated with apoptosis and whether it participates in the process of term amniotic rupture. Forty pieces of fresh term amnions, including twenty from a position near the margin of the placentas and twenty from the margin of the naturally ruptured part of the placentas in term gestation were collected, respectively. The amnions were examined by scanning electron microscopy (SEM) and amniotic epithelial (AE) cells were examined by transmission electron microscopy (TEM). Autophagic and apoptotic cell death (PCD) were assayed by laser scanning confocal microscopy (LSCM) or flow cytometry using monodansylcadaverin (MDC) and propidium iodide (PI) stain. BCL(2) and BAX were examined by immunoblotting. Under SEM the amniotic epithelia appeared normal in the position near the placenta. They had an atrophied appearance in the margin of their natural broken parts. In the AE cells PCD was divided into three subtypes by TEM: autophagic cell death with positive stains of MDC and PI; apoptotic cell death; and the mixed type. Quantitative detection showed that there were more death cells, including autophagic and apoptotic, in the AE cells near the ruptured parts than near the placentas. An increased expression of BAX and a decreased expression of BCL(2) protein in the AE cells near the broken margin were observed. Apoptotic and autophagic cell death by the intrinsic pathway are the basic event in the AE cell and they are involved in the cause of membrane rupture of the human amnion in term gestation.

  6. Multilineage potential research of bovine amniotic fluid mesenchymal stem cells.

    PubMed

    Gao, Yuhua; Zhu, Zhiqiang; Zhao, Yuhua; Hua, Jinlian; Ma, Yuehui; Guan, Weijun

    2014-01-01

    The use of amnion and amniotic fluid (AF) are abundant sources of mesenchymal stem cells (MSCs) that can be harvested at low cost and do not pose ethical conflicts. In human and veterinary research, stem cells derived from these tissues are promising candidates for disease treatment, specifically for their plasticity, their reduced immunogenicity, and high anti-inflammatory potential. This work aimed to obtain and characterize bovine amniotic fluid mesenchymal stem cells (AFMSC). The bovine AF from the amniotic cavity of pregnant gilts in the early stages of gestation (3- and 4-m-old bovine embryos) was collected. AFMSCs exhibit a fibroblastic-like morphology only starting from the fourth passage, being heterogeneous during the primary culture. Immunofluorescence results showed that AFMSCs were positive for β-integrin, CD44, CD73 and CD166, but negative for CD34, CD45. Meanwhile, AFMSCs expressed ES cell markers, such as Oct4, and when appropriately induced, are capable of differentiating into ectodermal and mesodermal lineages. This study reinforces the emerging importance of these cells as ideal tools in veterinary medicine; future studies aimed at a deeper evaluation of their immunological properties will allow a better understanding of their role in cellular therapy. PMID:24590129

  7. Multilineage potential research of bovine amniotic fluid mesenchymal stem cells.

    PubMed

    Gao, Yuhua; Zhu, Zhiqiang; Zhao, Yuhua; Hua, Jinlian; Ma, Yuehui; Guan, Weijun

    2014-02-28

    The use of amnion and amniotic fluid (AF) are abundant sources of mesenchymal stem cells (MSCs) that can be harvested at low cost and do not pose ethical conflicts. In human and veterinary research, stem cells derived from these tissues are promising candidates for disease treatment, specifically for their plasticity, their reduced immunogenicity, and high anti-inflammatory potential. This work aimed to obtain and characterize bovine amniotic fluid mesenchymal stem cells (AFMSC). The bovine AF from the amniotic cavity of pregnant gilts in the early stages of gestation (3- and 4-m-old bovine embryos) was collected. AFMSCs exhibit a fibroblastic-like morphology only starting from the fourth passage, being heterogeneous during the primary culture. Immunofluorescence results showed that AFMSCs were positive for β-integrin, CD44, CD73 and CD166, but negative for CD34, CD45. Meanwhile, AFMSCs expressed ES cell markers, such as Oct4, and when appropriately induced, are capable of differentiating into ectodermal and mesodermal lineages. This study reinforces the emerging importance of these cells as ideal tools in veterinary medicine; future studies aimed at a deeper evaluation of their immunological properties will allow a better understanding of their role in cellular therapy.

  8. A prospective study of 20 foot and ankle wounds treated with cryopreserved amniotic membrane and fluid allograft.

    PubMed

    Werber, Bruce; Martin, Erin

    2013-01-01

    We reviewed the background information and previous clinical studies that considered the use of allogeneic amniotic tissue and fluid (granulized amniotic membrane and amniotic fluid) in the treatment of chronic diabetic foot wounds. This innovation represents a relatively new approach to wound management by delivering a unique allograft of live human cells in a nonimmunogenic structural tissue matrix. Developed to fill soft tissue defects and bone voids and to convey antimicrobial and anti-inflammatory capabilities, granulized amniotic membrane and amniotic fluid does not require fetal death, because its procurement is performed with maternal consent during birth. In the present investigation, 20 chronic wounds (20 patients) that had been treated with standard wound therapy for a mean of 36.6 ± 31.58 weeks and with a mean baseline area of 10.15 ± 19.54 cm(2) were followed up during a 12-week observation period or until they healed. A total of 18 of the wounds (90%) healed during the 12-week observation period, and none of the wounds progressed to amputation. From our experience with the patients in the present case series, we believe that granulized amniotic membrane and amniotic fluid represents a useful option for the treatment of chronic diabetic foot wounds.

  9. miR-145 modulates lncRNA-ROR and Sox2 expression to maintain human amniotic epithelial stem cell pluripotency and β islet-like cell differentiation efficiency.

    PubMed

    Zou, Gang; Liu, Te; Guo, Lihe; Huang, Yongyi; Feng, Ya; Huang, Qin; Duan, Tao

    2016-10-10

    In this study, we observed a great reduction in the expression of the endogenous long noncoding RNA ROR (lncRNA-ROR) and the stem cell transcription factor Sox2, in contrast to a marked increase in miR-145 expression, during the course of in vitro induced differentiation of human amniotic epithelial stem cells (HuAECs). Bioinformatics analysis and the luciferase reporter assay revealed binding of miR-145 to specific sites in lncRNA-ROR and Sox2, silencing their expression. Overexpression of a lncRNA-ROR-specific siRNA effectively downregulated the expression levels of Sox2 and other stem cell markers in HuAECs while weakening the efficiency of HuAEC differentiation into β islet-like cells. Moreover, the in vitro response of HuAEC-derived β islet-like cells to extracellular stimuli and C-peptide release by these cells were markedly weakened in the siRNA-ROR transfection group. Furthermore, the in vivo expression of β islet-like cell biomarkers was substantially reduced in HuAECs in the siRNA-ROR transfection group, and their in vivo β islet-like cell differentiation and insulin release capacities were reduced in a streptozocin-induced diabetic rat model. The experimental results indicate that lncRNA-ROR effectively maintains Sox2 gene expression through competitive binding to miR-145, achieving pluripotency maintenance in HuAECs and regulation of their directed β islet-like cell differentiation efficiency.

  10. In vivo differentiation of human amniotic epithelial cells into cardiomyocyte-like cells and cell transplantation effect on myocardial infarction in rats: comparison with cord blood and adipose tissue-derived mesenchymal stem cells.

    PubMed

    Fang, Cheng-Hu; Jin, Jiyong; Joe, Jun-Ho; Song, Yi-Sun; So, Byung-Im; Lim, Sang Moo; Cheon, Gi Jeong; Woo, Sang-Keun; Ra, Jeong-Chan; Lee, Young-Yiul; Kim, Kyung-Soo

    2012-01-01

    Human amniotic epithelial cells (h-AECs), which have various merits as a cell source for cell therapy, are known to differentiate into cardiomyocytes in vitro. However, the ability of h-AECs to differentiate into cardiomyocytes in vivo and their cell transplantation effects on myocardial infarction are still unknown. In this study, we assessed whether h-AECs could differentiate into cardiomyocytes in vivo and whether h-AECs transplantation can decrease infarct size and improve cardiac function, in comparison to transplantation of cord blood-derived mesenchymal stem cells (MSCs) or adipose tissue-derived MSCs. For our study, we injected h-AECs, cord blood-derived MSCs, adipose tissue-derived MSCs, and saline into areas of myocardial infarction in athymic nude rats. After 4 weeks, 3% of the surviving h-AECs expressed myosin heavy chain, a marker specific to the myocardium. Compared with the saline group, all cell-implanted groups showed a higher ejection fraction, lower infarct area by positron emission tomography and histology, and more abundant myocardial gene and protein expression in the infarct area. We showed that h-AECs can differentiate into cardiomyocyte-like cells, decrease infarct size, and improve cardiac function in vivo. The beneficial effects of h-AECs were comparable to those of cord blood and adipose tissue-derived MSCs. These results support the need for further studies of h-AECs as a cell source for myocardial regeneration due to their plentiful availability, low immunity, and lack of ethical issues related to their use.

  11. In vivo differentiation of human amniotic epithelial cells into cardiomyocyte-like cells and cell transplantation effect on myocardial infarction in rats: comparison with cord blood and adipose tissue-derived mesenchymal stem cells.

    PubMed

    Fang, Cheng-Hu; Jin, Jiyong; Joe, Jun-Ho; Song, Yi-Sun; So, Byung-Im; Lim, Sang Moo; Cheon, Gi Jeong; Woo, Sang-Keun; Ra, Jeong-Chan; Lee, Young-Yiul; Kim, Kyung-Soo

    2012-01-01

    Human amniotic epithelial cells (h-AECs), which have various merits as a cell source for cell therapy, are known to differentiate into cardiomyocytes in vitro. However, the ability of h-AECs to differentiate into cardiomyocytes in vivo and their cell transplantation effects on myocardial infarction are still unknown. In this study, we assessed whether h-AECs could differentiate into cardiomyocytes in vivo and whether h-AECs transplantation can decrease infarct size and improve cardiac function, in comparison to transplantation of cord blood-derived mesenchymal stem cells (MSCs) or adipose tissue-derived MSCs. For our study, we injected h-AECs, cord blood-derived MSCs, adipose tissue-derived MSCs, and saline into areas of myocardial infarction in athymic nude rats. After 4 weeks, 3% of the surviving h-AECs expressed myosin heavy chain, a marker specific to the myocardium. Compared with the saline group, all cell-implanted groups showed a higher ejection fraction, lower infarct area by positron emission tomography and histology, and more abundant myocardial gene and protein expression in the infarct area. We showed that h-AECs can differentiate into cardiomyocyte-like cells, decrease infarct size, and improve cardiac function in vivo. The beneficial effects of h-AECs were comparable to those of cord blood and adipose tissue-derived MSCs. These results support the need for further studies of h-AECs as a cell source for myocardial regeneration due to their plentiful availability, low immunity, and lack of ethical issues related to their use. PMID:22776022

  12. Acellular Dermal Matrix in Rotator Cuff Surgery.

    PubMed

    Cooper, Joseph; Mirzayan, Raffy

    2016-01-01

    The success of rotator cuff repair (RCR) surgery can be measured clinically (validated outcome scores, range of motion) as well as structurally (re-tear rates using imaging studies). Regardless of repair type or technique, most studies have shown that patients do well clinically. However, multiple studies have also shown that structurally, the failure rate can be very high. A variety of factors, including poor tendon quality, age over 63 years, smoking, advanced fatty infiltration into the muscle, and the inability of the tendon to heal to bone, have been implicated as the cause of the high re-tear rate in RCRs. The suture-tendon interface is felt to be the weakest link in the RCR construct, and suture pullout through the tendon is believed to be the most common method of failure. This review of the published literature seeks to determine if there is support for augmentation of RCR with acellular dermal matrices to strengthen the suture-tendon interface and reduce the re-tear rate. PMID:27552454

  13. Challenges to acellular biological scaffold mediated skeletal muscle tissue regeneration.

    PubMed

    Corona, Benjamin T; Greising, Sarah M

    2016-10-01

    Volumetric muscle loss (VML) injuries present a complex and heterogeneous clinical problem that results in a chronic loss of muscle tissue and strength. The primary limitation to muscle tissue regeneration after VML injury is the frank loss of all native muscle constituents in the defect, especially satellite cells and the basal lamina. Recent advancements in regenerative medicine have set forth encouraging and emerging translational and therapeutic options for these devastating injuries including the surgical implantation of acellular biological scaffolds. While these biomaterials can modulate the wound environment, the existing data do not support their capacity to promote appreciable muscle fiber regeneration that can contribute to skeletal muscle tissue functional improvements. An apparent restriction of endogenous satellite cell (i.e., pax7(+)) migration to acellular biological scaffolds likely underlies this deficiency. This work critically evaluates the role of an acellular biological scaffold in orchestrating skeletal muscle tissue regeneration, specifically when used as a regenerative medicine approach for VML injury. PMID:27472161

  14. Challenges to acellular biological scaffold mediated skeletal muscle tissue regeneration.

    PubMed

    Corona, Benjamin T; Greising, Sarah M

    2016-10-01

    Volumetric muscle loss (VML) injuries present a complex and heterogeneous clinical problem that results in a chronic loss of muscle tissue and strength. The primary limitation to muscle tissue regeneration after VML injury is the frank loss of all native muscle constituents in the defect, especially satellite cells and the basal lamina. Recent advancements in regenerative medicine have set forth encouraging and emerging translational and therapeutic options for these devastating injuries including the surgical implantation of acellular biological scaffolds. While these biomaterials can modulate the wound environment, the existing data do not support their capacity to promote appreciable muscle fiber regeneration that can contribute to skeletal muscle tissue functional improvements. An apparent restriction of endogenous satellite cell (i.e., pax7(+)) migration to acellular biological scaffolds likely underlies this deficiency. This work critically evaluates the role of an acellular biological scaffold in orchestrating skeletal muscle tissue regeneration, specifically when used as a regenerative medicine approach for VML injury.

  15. Isolation and morphological characterization of ovine amniotic fluid mesenchymal stem cells

    PubMed Central

    Tian, Yunyun; Tao, Li; Zhao, Siriguleng; Tai, Dapeng; Liu, Dongjun; Liu, Pengxia

    2015-01-01

    Mesenchymal stem cells (MSCs) are one of the most promising cell populations for tissue engineering and regenerative medicine. Of utmost importance to MSC research is identification of MSC sources that are easily obtainable and stable. Several studies have shown that MSCs can be isolated from amniotic fluid. The sheep is one of the main types of farm animal, and it has many biophysical and biochemical similarities to humans. Here, we obtained MSCs from ovine amniotic fluid and determined the expansion capacity, surface and intracellular marker expression, karyotype, and multilineage differentiation ability of these ovine amniotic fluid mesenchymal stem cells (oAF-MSCs). Moreover, expression levels of differentiation markers were measured using reverse transcription-qPCR (RT-qPCR). Our phenotypic analysis shows that the isolated oAF-MSCs are indeed MSCs. PMID:26616638

  16. Characterization of amniotic stem cells.

    PubMed

    Koike, Chika; Zhou, Kaixuan; Takeda, Yuji; Fathy, Moustafa; Okabe, Motonori; Yoshida, Toshiko; Nakamura, Yukio; Kato, Yukio; Nikaido, Toshio

    2014-08-01

    The amnion membrane is developed from embryo-derived cells, and amniotic cells have been shown to exhibit multidifferentiation potential. These cells represent a desirable source for stem cells for a variety of reasons. However, to date very few molecular analyses of amnion-derived cells have been reported, and efficient markers for isolating the stem cells remain unclear. This paper assesses the characterization of amnion-derived cells as stem cells by examining stemness marker expressions for amnion-derived epithelial cells and mesenchymal cells by flow cytometry, immunocytochemistry, and quantitative PCR. Flow cytometry revealed that amnion epithelial cells expressed CD133, CD 271, and TRA-1-60, whereas mecenchymal cells expressed CD44, CD73, CD90, and CD105. Immunohistochemistry showed that both cells expressed the stemness markers Oct3/4, Sox2, Klf4, and SSEA4. Stemness genes' expression in amnion epithelial cells, mesenchymal cells, fibroblast, bone marrow-derived mesenchymal stem cells (MSCs), and induced pluripotent stem cells (iPSCs) was compared by quantitative reverse-transcription polymerase chain reaction (RT-PCR). Amnion-derived epithelial cells and mesenchymal cells expressed Oct3/4, Nanog, and Klf4 more than bone marrow-derived MSCs. The sorted TRA1-60-positive cells expressed Oct3/4, Nanog, and Klf4 more than unsorted cells or TRA1-60-negative cells. TRA1-60 can be a marker for isolating amnion epithelial stem cells.

  17. Studies in porphyria. IV. Expression of the gene defect of acute intermittent porphyria in cultured human skin fibroblasts and amniotic cells: prenatal diagnosis of the porphyric trait

    PubMed Central

    1975-01-01

    The gene lesion of the porphyrin-heme synthetic pathway in acute intermittent porphyria (AIP) is reflected in a deficient level of activity of the cytosol enzyme uroporphyrinogen I synthetase (URO-S). A marked URO-S deficiency has been demonstrated in the liver and in circulating erythrocytes of individuals with both active and latent AIP. This enzymic abnormality accounts for the excessive production and excretion into urine of the porphyrin precursors, lamda-aminolevulinic acid (ALA) and porphobilinogen (PBG) in AIP subjects. In this study, utilizing cell culture techniques, a marked URO-S deficiency has also been demonstrated in skin fibroblasts from AIP patients and in cells derived through aminocentesis from an approximately 17-wk old fetus. The prenatal diagnosis of the AIP trait in this fetus was confirmed postnatally by the demonstration in the child of a deficient level of erythrocyte URO-S activity which was comparable to those found in her AIP mother and affected sibling and which was approximately one-half the levels characterizing her normal father and aunt and a second unaffected sibling. The identification of the URO-S deficiency in cultured human fibroblasts from AIP patients was facilitated by a newly developed, sensitive assay for the enzyme activity. In this assay, the ability of such cells to convert ALA to protoporphyrin was quantitated; in the sequence of reactions involved in this transformation, URO-S is limiting so that the gene defect of AIP could be simply and precisely determined by appropriate spectrofluorometry of cell extracts. The technique described has distinct advantages over the direct enzymatic assay for URO-S activity in cultured human skin fibroblasts and permits clear differentiation of AIP carrier from normal individuals. PMID:1165472

  18. Studies in porphyria. IV. Expression of the gene defect of acute intermittent porphyria in cultured human skin fibroblasts and amniotic cells: prenatal diagnosis of the porphyric trait.

    PubMed

    Sassa, S; Solish, G; Levere, R D; Kappas, A

    1975-09-01

    The gene lesion of the porphyrin-heme synthetic pathway in acute intermittent porphyria (AIP) is reflected in a deficient level of activity of the cytosol enzyme uroporphyrinogen I synthetase (URO-S). A marked URO-S deficiency has been demonstrated in the liver and in circulating erythrocytes of individuals with both active and latent AIP. This enzymic abnormality accounts for the excessive production and excretion into urine of the porphyrin precursors, lamda-aminolevulinic acid (ALA) and porphobilinogen (PBG) in AIP subjects. In this study, utilizing cell culture techniques, a marked URO-S deficiency has also been demonstrated in skin fibroblasts from AIP patients and in cells derived through aminocentesis from an approximately 17-wk old fetus. The prenatal diagnosis of the AIP trait in this fetus was confirmed postnatally by the demonstration in the child of a deficient level of erythrocyte URO-S activity which was comparable to those found in her AIP mother and affected sibling and which was approximately one-half the levels characterizing her normal father and aunt and a second unaffected sibling. The identification of the URO-S deficiency in cultured human fibroblasts from AIP patients was facilitated by a newly developed, sensitive assay for the enzyme activity. In this assay, the ability of such cells to convert ALA to protoporphyrin was quantitated; in the sequence of reactions involved in this transformation, URO-S is limiting so that the gene defect of AIP could be simply and precisely determined by appropriate spectrofluorometry of cell extracts. The technique described has distinct advantages over the direct enzymatic assay for URO-S activity in cultured human skin fibroblasts and permits clear differentiation of AIP carrier from normal individuals.

  19. Chondrogenic differentiation of amniotic fluid-derived stem cells.

    PubMed

    Kolambkar, Yash M; Peister, Alexandra; Soker, Shay; Atala, Anthony; Guldberg, Robert E

    2007-10-01

    For regenerating damaged articular cartilage, it is necessary to identify an appropriate cell source that is easily accessible, can be expanded to large numbers, and has chondrogenic potential. Amniotic fluid-derived stem (AFS) cells have recently been isolated from human and rodent amniotic fluid and shown to be highly proliferative and broadly pluripotent. The purpose of this study was to investigate the chondrogenic potential of human AFS cells in pellet and alginate hydrogel cultures. Human AFS cells were expanded in various media conditions, and cultured for three weeks with growth factor supplementation. There was increased production of sulfated glycosaminoglycan (sGAG) and type II collagen in response to transforming growth factor-beta (TGF-beta) supplementation, with TGF-beta1 producing greater increases than TGF-beta3. Modification of expansion media supplements and addition of insulin-like growth factor-1 during pellet culture further increased sGAG/DNA over TGF-beta1 supplementation alone. Compared to bone marrow-derived mesenchymal stem cells, the AFS cells produced less cartilaginous matrix after three weeks of TGF-beta1 supplementation in pellet culture. Even so, this study demonstrates that AFS cells have the potential to differentiate along the chondrogenic lineage, thus establishing the feasibility of using these cells for cartilage repair applications. PMID:17668282

  20. Amniotic Band Syndrome - A Dreaded Condition

    PubMed Central

    Renukadevi, T.K.

    2016-01-01

    Amniotic band syndrome is a unique condition in which amnion a normal structure causes complications. A case of second gravid, obese who is a known diabetic came to OPD at 13 weeks pregnancy for regular antenatal check up. A routine ultrasonogram was advised in which multiple anomalies were noted and the diagnosis of amniotic band syndrome was made. The parents were counseled for medical termination of pregnancy and after obtaining the consent termination were performed and the parents were asked to postpone the next pregnancy for minimum 6 months. This anomaly as seen in this patient could be due to risk factors like diabetes and obesity. PMID:26894130

  1. Amniotic Band Syndrome - A Dreaded Condition.

    PubMed

    R, Durga; Renukadevi, T K

    2016-01-01

    Amniotic band syndrome is a unique condition in which amnion a normal structure causes complications. A case of second gravid, obese who is a known diabetic came to OPD at 13 weeks pregnancy for regular antenatal check up. A routine ultrasonogram was advised in which multiple anomalies were noted and the diagnosis of amniotic band syndrome was made. The parents were counseled for medical termination of pregnancy and after obtaining the consent termination were performed and the parents were asked to postpone the next pregnancy for minimum 6 months. This anomaly as seen in this patient could be due to risk factors like diabetes and obesity.

  2. Evaluation of lymphangiogenesis in acellular dermal matrix

    PubMed Central

    Cherubino, Mario; Pellegatta, Igor; Tamborini, Federico; Cerati, Michele; Sessa, Fausto; Valdatta, Luigi

    2014-01-01

    Introduction: Much attention has been directed towards understanding the phenomena of angiogenesis and lymphangiogenesis in wound healing. Thanks to the manifold dermal substitute available nowadays, wound treatment has improved greatly. Many studies have been published about angiogenesis and cell invasion in INTEGRA®. On the other hand, the development of the lymphatic network in acellular dermal matrix (ADM) is a more obscure matter. In this article, we aim to characterize the different phases of host cell invasion in ADM. Special attention was given to lymphangiogenic aspects. Materials and Methods: Among 57 rats selected to analyse the role of ADM in lymphangiogenesis, we created four groups. We performed an excision procedure on both thighs of these rats: On the left one we did not perform any action except repairing the borders of the wound; while on the right one we used INTEGRA® implant. The excision biopsy was performed at four different times: First group after 7 days, second after 14 days, third after 21 days and fourth after 28 days. For our microscopic evaluation, we used the classical staining technique of haematoxylin and eosin and a semi-quantitative method in order to evaluate cellularity counts. To assess angiogenesis and lymphangiogenesis development we employed PROX-1 Ab and CD31/PECAM for immunohistochemical analysis. Results: We found remarkable wound contraction in defects that healed by secondary intention while minor wound contraction was observed in defects treated with ADM. At day 7, optical microscopy revealed a more plentiful cellularity in the granulation tissue compared with the dermal regeneration matrix. The immunohistochemical process highlighted vascular and lymphatic cells in both groups. After 14 days a high grade of fibrosis was noticeable in the non-treated group. At day 21, both lymphatic and vascular endothelial cells were better developed in the group with a dermal matrix application. At day 28, lymphatic endothelial

  3. Acellular Nerve Allografts in Peripheral Nerve Regeneration: A Comparative Study

    PubMed Central

    Moore, Amy M.; MacEwan, Matthew; Santosa, Katherine B.; Chenard, Kristofer E.; Ray, Wilson Z.; Hunter, Daniel A.; Mackinnon, Susan E.; Johnson, Philip J.

    2011-01-01

    Background Processed nerve allografts offer a promising alternative to nerve autografts in the surgical management of peripheral nerve injuries where short deficits exist. Methods Three established models of acellular nerve allograft (cold-preserved, detergent-processed, and AxoGen® -processed nerve allografts) were compared to nerve isografts and silicone nerve guidance conduits in a 14 mm rat sciatic nerve defect. Results All acellular nerve grafts were superior to silicone nerve conduits in support of nerve regeneration. Detergent-processed allografts were similar to isografts at 6 weeks post-operatively, while AxoGen®-processed and cold-preserved allografts supported significantly fewer regenerating nerve fibers. Measurement of muscle force confirmed that detergent-processed allografts promoted isograft-equivalent levels of motor recovery 16 weeks post-operatively. All acellular allografts promoted greater amounts of motor recovery compared to silicone conduits. Conclusions These findings provide evidence that differential processing for removal of cellular constituents in preparing acellular nerve allografts affects recovery in vivo. PMID:21660979

  4. Development and Characterization of Acellular Extracellular Matrix Scaffolds from Porcine Menisci for Use in Cartilage Tissue Engineering

    PubMed Central

    Chen, Ying-Chen; Chen, Ray-Neng; Jhan, Hua-Jing; Liu, Der-Zen; Ho, Hsiu-O; Mao, Yong; Kohn, Joachim

    2015-01-01

    Given the growing number of arthritis patients and the limitations of current treatments, there is great urgency to explore cartilage substitutes by tissue engineering. In this study, we developed a novel decellularization method for menisci to prepare acellular extracellular matrix (ECM) scaffolds with minimal adverse effects on the ECM. Among all the acid treatments, formic acid treatment removed most of the cellular contents and preserved the highest ECM contents in the decellularized porcine menisci. Compared with fresh porcine menisci, the content of DNA decreased to 4.10%±0.03%, and there was no significant damage to glycosaminoglycan (GAG) or collagen. Histological staining also confirmed the presence of ECM and the absence of cellularity. In addition, a highly hydrophilic scaffold with three-dimensional interconnected porous structure was fabricated from decellularized menisci tissue. Human chondrocytes showed enhanced cell proliferation and synthesis of chondrocyte ECM including type II collagen and GAG when cultured in this acellular scaffold. Moreover, the scaffold effectively supported chondrogenesis of human bone marrow-derived mesenchymal stem cells. Finally, in vivo implantation was conducted in rats to assess the biocompatibility of the scaffolds. No significant inflammatory response was observed. The acellular ECM scaffold provided a native environment for cells with diverse physiological functions to promote cell proliferation and new tissue formation. This study reported a novel way to prepare decellularized meniscus tissue and demonstrated the potential as scaffolds to support cartilage repair. PMID:25919905

  5. Development and Characterization of Acellular Extracellular Matrix Scaffolds from Porcine Menisci for Use in Cartilage Tissue Engineering.

    PubMed

    Chen, Ying-Chen; Chen, Ray-Neng; Jhan, Hua-Jing; Liu, Der-Zen; Ho, Hsiu-O; Mao, Yong; Kohn, Joachim; Sheu, Ming-Thau

    2015-09-01

    Given the growing number of arthritis patients and the limitations of current treatments, there is great urgency to explore cartilage substitutes by tissue engineering. In this study, we developed a novel decellularization method for menisci to prepare acellular extracellular matrix (ECM) scaffolds with minimal adverse effects on the ECM. Among all the acid treatments, formic acid treatment removed most of the cellular contents and preserved the highest ECM contents in the decellularized porcine menisci. Compared with fresh porcine menisci, the content of DNA decreased to 4.10%±0.03%, and there was no significant damage to glycosaminoglycan (GAG) or collagen. Histological staining also confirmed the presence of ECM and the absence of cellularity. In addition, a highly hydrophilic scaffold with three-dimensional interconnected porous structure was fabricated from decellularized menisci tissue. Human chondrocytes showed enhanced cell proliferation and synthesis of chondrocyte ECM including type II collagen and GAG when cultured in this acellular scaffold. Moreover, the scaffold effectively supported chondrogenesis of human bone marrow-derived mesenchymal stem cells. Finally, in vivo implantation was conducted in rats to assess the biocompatibility of the scaffolds. No significant inflammatory response was observed. The acellular ECM scaffold provided a native environment for cells with diverse physiological functions to promote cell proliferation and new tissue formation. This study reported a novel way to prepare decellularized meniscus tissue and demonstrated the potential as scaffolds to support cartilage repair. PMID:25919905

  6. Adjuvant neurotrophic factors in peripheral nerve repair with chondroitin sulfate proteoglycan-reduced acellular nerve allografts

    PubMed Central

    Boyer, Richard B.; Sexton, Kevin W.; Rodriguez-Feo, Charles L.; Nookala, Ratnam; Pollins, Alonda C.; Cardwell, Nancy L.; Tisdale, Keonna Y.; Nanney, Lillian B.; Shack, R. Bruce; Thayer, Wesley P.

    2014-01-01

    Background Acellular nerve allografts are now standard tools in peripheral nerve repair due to decreased donor site morbidity and operative time savings. Preparation of nerve allografts involves several steps of decellularization and modification of extracellular matrix to remove chondroitin sulfate proteoglycans (CSPGs), which have been shown to inhibit neurite outgrowth through a poorly understood mechanism involving RhoA and ECM-integrin interactions. Chondroitinase ABC (ChABC) is an enzyme that degrades CSPG molecules and has been shown to promote neurite outgrowth following injury of the central and peripheral nervous systems. Variable results following chondroitinase ABC treatment make it difficult to predict the effects of this drug in human nerve allografts, especially in the presence of native extracellular signaling molecules. Several studies have shown cross-talk between neurotrophic factor and CSPG signaling pathways, but their interaction remains poorly understood. In this study, we examined the adjuvant effects of nerve growth factor (NGF) and glial cell line-derived neurotrophic factor (GDNF) on neurite outgrowth post-injury in CSPG-reduced substrates and acellular nerve allografts. Materials and Methods E12 chicken DRG explants were cultured in medium containing ChABC, ChABC + NGF, ChABC + GDNF or control media. Explants were imaged at 3 d and neurite outgrowths measured. The rat sciatic nerve injury model involved a 1-cm sciatic nerve gap that was microsurgically repaired with ChABC pre-treated acellular nerve allografts. Prior to implantation, nerve allografts were incubated in NGF, GDNF or sterile water. Nerve histology was evaluated at 5d and 8wk post-injury. Results The addition of GDNF in vitro produced significant increase in sensory neurite length at 3 d compared to ChABC alone (P < 0.01), while NGF was not significantly different from control. In vivo adjuvant NGF produced increases in total myelinated axon count (P < 0.005) and motor axon

  7. The amniotic membrane as a source of stem cells.

    PubMed

    Insausti, Carmen L; Blanquer, Miguel; Bleda, Patricia; Iniesta, Paqui; Majado, María J; Castellanos, Gregorio; Moraleda, José M

    2010-01-01

    Cellular therapy has emerged as a new potential tool for curing a wide range of degenerative diseases and tissue necrosis. Embryonic stem cells possess potential for differentiation into a wide range of cell lineages, but the ethical issues associated with establishment of this human cell line have to be resolved prior to any use. The bone marrow (BM) is the usual source of adult stem cells for hematopoietic stem cell transplants and cellular therapy, but the BM harvest is a surgical procedure that requires general anesthesia or sedation, and there seems to be a reduction of the proliferative potential and differentiation capacity of the marrow mesenchymal stem cells in older donors. For these reasons there is an increasing interest in other sources of stem cells from adult and fetal tissues. The amniotic membrane (AM) or amnion is a tissue of particular interest because its cells possess characteristics of stem cells with multipotent differentiation ability, and because of low immunogenicity and easy procurement from the placenta, which is a discarded tissue after parturition, thus avoiding the current controversies associated with the use of human embryonic stem cells. Therefore, amniotic membrane has been proposed as a good candidate to be used in cellular therapy and regenerative medicine.

  8. Inhibitor of intramembranous absorption in ovine amniotic fluid.

    PubMed

    Brace, Robert A; Cheung, Cecilia Y; Anderson, Debra F

    2014-02-01

    Intramembranous absorption increases during intra-amniotic infusion of physiological saline solutions. The increase may be due partly to the concomitant elevation in fetal urine production as fetal urine contains a stimulator of intramembranous absorption. In this study, we hypothesized that the increase in intramembranous absorption during intra-amniotic infusion is due, in part, to dilution of a nonrenal inhibitor of intramembranous absorption that is present in amniotic fluid. In late-gestation fetal sheep, amniotic fluid volume and the four primary amniotic inflows and outflows were determined over 2-day intervals under three conditions: 1) control conditions when fetal urine entered the amniotic sac, 2) during intra-amniotic infusion of 2 l/day of lactated Ringer solution when urine entered the amniotic sac, and 3) during the same intra-amniotic infusion when fetal urine was continuously replaced with lactated Ringer solution. Amniotic fluid volume, fetal urine production, swallowed volume, and intramembranous absorption rate increased during the infusions independent of fetal urine entry into the amniotic sac or its replacement. Lung liquid secretion rate was unchanged during infusion. Because fetal membrane stretch has been shown not to be involved and because urine replacement did not alter the response, we conclude that the increase in intramembranous absorption that occurs during intra-amniotic infusions is due primarily to dilution of a nonrenal inhibitor of intramembranous absorption that is normally present in amniotic fluid. This result combined with our previous study suggests that a nonrenal inhibitor(s) together with a renal stimulator(s) interact to regulate intramembranous absorption rate and, hence, amniotic fluid volume.

  9. Inhibitor of intramembranous absorption in ovine amniotic fluid

    PubMed Central

    Cheung, Cecilia Y.; Anderson, Debra F.

    2013-01-01

    Intramembranous absorption increases during intra-amniotic infusion of physiological saline solutions. The increase may be due partly to the concomitant elevation in fetal urine production as fetal urine contains a stimulator of intramembranous absorption. In this study, we hypothesized that the increase in intramembranous absorption during intra-amniotic infusion is due, in part, to dilution of a nonrenal inhibitor of intramembranous absorption that is present in amniotic fluid. In late-gestation fetal sheep, amniotic fluid volume and the four primary amniotic inflows and outflows were determined over 2-day intervals under three conditions: 1) control conditions when fetal urine entered the amniotic sac, 2) during intra-amniotic infusion of 2 l/day of lactated Ringer solution when urine entered the amniotic sac, and 3) during the same intra-amniotic infusion when fetal urine was continuously replaced with lactated Ringer solution. Amniotic fluid volume, fetal urine production, swallowed volume, and intramembranous absorption rate increased during the infusions independent of fetal urine entry into the amniotic sac or its replacement. Lung liquid secretion rate was unchanged during infusion. Because fetal membrane stretch has been shown not to be involved and because urine replacement did not alter the response, we conclude that the increase in intramembranous absorption that occurs during intra-amniotic infusions is due primarily to dilution of a nonrenal inhibitor of intramembranous absorption that is normally present in amniotic fluid. This result combined with our previous study suggests that a nonrenal inhibitor(s) together with a renal stimulator(s) interact to regulate intramembranous absorption rate and, hence, amniotic fluid volume. PMID:24381178

  10. Differentiation of mesenchymal stem cells into neuronal cells on fetal bovine acellular dermal matrix as a tissue engineered nerve scaffold.

    PubMed

    Feng, Yuping; Wang, Jiao; Ling, Shixin; Li, Zhuo; Li, Mingsheng; Li, Qiongyi; Ma, Zongren; Yu, Sijiu

    2014-11-15

    The purpose of this study was to assess fetal bovine acellular dermal matrix as a scaffold for supporting the differentiation of bone marrow mesenchymal stem cells into neural cells following induction with neural differentiation medium. We performed long-term, continuous observation of cell morphology, growth, differentiation, and neuronal development using several microscopy techniques in conjunction with immunohistochemistry. We examined specific neuronal proteins and Nissl bodies involved in the differentiation process in order to determine the neuronal differentiation of bone marrow mesenchymal stem cells. The results show that bone marrow mesenchymal stem cells that differentiate on fetal bovine acellular dermal matrix display neuronal morphology with unipolar and bi/multipolar neurite elongations that express neuronal-specific proteins, including βIII tubulin. The bone marrow mesenchymal stem cells grown on fetal bovine acellular dermal matrix and induced for long periods of time with neural differentiation medium differentiated into a multilayered neural network-like structure with long nerve fibers that was composed of several parallel microfibers and neuronal cells, forming a complete neural circuit with dendrite-dendrite to axon-dendrite to dendrite-axon synapses. In addition, growth cones with filopodia were observed using scanning electron microscopy. Paraffin sectioning showed differentiated bone marrow mesenchymal stem cells with the typical features of neuronal phenotype, such as a large, round nucleus and a cytoplasm full of Nissl bodies. The data suggest that the biological scaffold fetal bovine acellular dermal matrix is capable of supporting human bone marrow mesenchymal stem cell differentiation into functional neurons and the subsequent formation of tissue engineered nerve.

  11. Use of hyperdry amniotic membrane in operations for cleft palate: a study in rats.

    PubMed

    Tsuno, Hiroaki; Noguchi, Makoto; Okabe, Motonori; Tomihara, Kei; Yoshida, Toshiko; Nikaido, Toshio

    2015-04-01

    The growth of maxillary bone and the development of dentition are often impaired in patients who have had pushback operations for repair of a cleft palate. There has been considerable discussion about the most suitable technique or material used in such repairs to resolve the problem. Hyperdry amniotic membrane, a new preservable material derived from human amnion, has recently been introduced in several procedures. We have evaluated its use during pushback surgery in animal studies to try to correct the inhibition of growth and development of the maxilla. Mucosal defects were created in 3-week-old rats, and then covered with hyperdry amniotic membrane or not. Healing was assessed by histological and morphological examination at 1 week and 7 weeks postoperatively. In the group treated with hyperdry amniotic membrane, submucosal tissue was reconstructed successfully during the early postoperative period. Lateral palatal growth was not inhibited as much, and medial inclination of the teeth was less, after a period of growth using this material. The results suggest that hyperdry amniotic membrane is a suitable new dressing material for use in the treatment of cleft palate.

  12. Buffalo (Bubalus bubalis) term amniotic-membrane-derived cells exhibited mesenchymal stem cells characteristics in vitro.

    PubMed

    Ghosh, Kaushalya; Kumar, Rajesh; Singh, Jarnail; Gahlawat, S K; Kumar, Dharmendra; Selokar, Naresh Lalaji; Yadav, S P; Gulati, B R; Yadav, P S

    2015-10-01

    Recent studies suggested that placentae amniotic membrane is a valuable source of stem cells in human as well as in livestock species. Advantages of amnion over other sources of stem cells included abundant availability, ethically non-objectionable and non-invasive source. The aim of the present study was the isolation, culture and characterization of amniotic-membrane-derived mesenchymal stem cells from term placentae collected postpartum in buffalo. We have observed that both presumptive epithelial-like and fibroblast-like cells were cultured and maintained from term amnion. These cells were shown the positive expression of pluripotency markers (OCT-4, SOX-2, NANOG, TERT), mesenchymal stem cell markers (CD29, CD44, CD105) and negative for haematopoietic marker (CD34) genes at different passages. In addition, these cells were also positive for alkaline phosphatase staining. Stem-ness potential of any stem cells is determined by their potential to differentiate into specific lineages of cell type. In the present study, we have successfully differentiated the amniotic-membrane-derived cells into adipogenic, chondrogenic and osteogenic lineages of cells in vitro. In conclusion, the results of this study demonstrate that amniotic-membrane-derived cells expressed pluripotent and mesenchymal stem cells markers and have propensity to differentiate into cells of mesenchymal lineage cell type upon directed differentiation in vitro.

  13. BIOCOMPATIBILITY OF ACELLULAR DERMAL MATRIX GRAFT EVALUATED IN CULTURE OF MURINE MACROPHAGES

    PubMed Central

    Vendramini, Ana Paula; Melo, Rafaela Fernanda; Marcantonio, Rosemary Adriana Chiérici; Carlos, Iracilda Zepone

    2006-01-01

    The acellular dermal matrix allograft has been used as an alternative to autogenous palatal mucosal graft. The aim of this study was the evaluation of the biocompatibility of an acellular dermal matrix (AlloDerm®) in culture of macrophages. For hydrogen peroxidase determination we used the method of Pick & Kesari, and the Griess method for nitric oxide determination,. Statistical analysis showed no significant difference (p ≤ 0,05) in the release of nitric oxide and hydrogen peroxide by the macrophages exposed to acellular dermal matrix and the negative control. The results suggest that acellular dermal matrix did not activate the cell inflammatory response. PMID:19089033

  14. Infection in the Nasal Tip Caused by Acellular Dermal Matrix.

    PubMed

    Lee, Kun Hee

    2015-12-01

    A 19-year-old female patient visited our clinic for rhinoplasty. She complained about her low take-off point, which was apparent in profile view, and wanted slight tip projection. She refused additional cartilage harvesting from ears or ribs but consented to the use of homologous tissue, including acellular dermal matrix, for her dorsum and tip. Septoturbinoplasty was performed, and only a very small amount of septal cartilage could be harvested. It was used as both the columellar strut and the alar rim graft. Nasal dorsum and tip were augmented with acellular dermal matrix. Three months postoperatively, she experienced a few episodes of edema and redness on her nasal tip, followed by pus exudation from the nasal skin. Six months postoperatively, she underwent revision rhinoplasty for removal of inflamed grafts, and onlay tip graft with homologous rib cartilage was performed. Nasal dorsum or tip grafts are an integral part of Asian rhinoplasty. Autogenous tissue is the gold standard for grafting materials. However, the limited availability of autogenous tissue and the preference of patients and surgeons for artificial surgical implants make Asian rhinoplasty challenging. Unavailability of autogenous cartilage and patient refusal of artificial implants led to the use of acellular dermal matrix (ADM) in the nasal dorsum and tip for this case. This is the first report of postoperative complication because of infection rather than absorption after ADM use.

  15. Infection in the Nasal Tip Caused by Acellular Dermal Matrix.

    PubMed

    Lee, Kun Hee

    2015-12-01

    A 19-year-old female patient visited our clinic for rhinoplasty. She complained about her low take-off point, which was apparent in profile view, and wanted slight tip projection. She refused additional cartilage harvesting from ears or ribs but consented to the use of homologous tissue, including acellular dermal matrix, for her dorsum and tip. Septoturbinoplasty was performed, and only a very small amount of septal cartilage could be harvested. It was used as both the columellar strut and the alar rim graft. Nasal dorsum and tip were augmented with acellular dermal matrix. Three months postoperatively, she experienced a few episodes of edema and redness on her nasal tip, followed by pus exudation from the nasal skin. Six months postoperatively, she underwent revision rhinoplasty for removal of inflamed grafts, and onlay tip graft with homologous rib cartilage was performed. Nasal dorsum or tip grafts are an integral part of Asian rhinoplasty. Autogenous tissue is the gold standard for grafting materials. However, the limited availability of autogenous tissue and the preference of patients and surgeons for artificial surgical implants make Asian rhinoplasty challenging. Unavailability of autogenous cartilage and patient refusal of artificial implants led to the use of acellular dermal matrix (ADM) in the nasal dorsum and tip for this case. This is the first report of postoperative complication because of infection rather than absorption after ADM use. PMID:26894006

  16. Amniotic fluid embolism after intrauterine fetal demise.

    PubMed

    Kristensen, Karl; Langdana, Fali; Clentworth, Howard; Hansby, Chu; Dalley, Paul

    2016-01-01

    We present a case of the successful treatment of severe amniotic fluid embolism in a 41-year-old woman undergoing emergency caesarean section at 36 weeks of gestation for placental abruption and intrauterine fetal demise. The treatment included prolonged cardiopulmonary resuscitation, emergency hysterectomy, re-operation with intra-abdominal packing and intra-aortic balloon pump insertion. The patient made a remarkable recovery and to date has minimal residual morbidity. Amniotic fluid embolism syndrome (AFES) is a rare and often fatal obstetric condition that remains one of the main causes of maternal mortality in developed countries. The incidence varies from 2 to 6 per 100,000 and suggested mortality rates exceed 60%.1-2 The classic triad of sudden hypoxia, hypotension and coagulopathy with acute onset during labour or immediately after delivery forms the hallmark of the AFES diagnosis, however AFES is primarily a clinical diagnosis of exclusion. We present a case of successful maternal outcome following severe amniotic fluid embolism after placental abruption and intrauterine fetal demise. PMID:27607089

  17. Interactions among pulmonary surfactant, vernix caseosa, and intestinal enterocytes: intra-amniotic administration of fluorescently liposomes to pregnant rabbits.

    PubMed

    Nishijima, Koji; Shukunami, Ken-ichi; Yoshinari, Hideo; Takahashi, Jin; Maeda, Hideyuki; Takagi, Hitoshi; Kotsuji, Fumikazu

    2012-08-01

    Although vernix caseosa is known to be a natural biofilm at birth, human pulmonary surfactant commences to remove the vernix from fetal skin into the amniotic fluid at gestational week 34, i.e., well before delivery. To explain this paradox, we first produced two types of fluorescently labeled liposomes displaying morphology similar to that of pulmonary surfactant and vernix caseosa complexes. We then continuously administered these liposomes into the amniotic fluid space of pregnant rabbits. In addition, we produced pulmonary surfactant and vernix caseosa complexes and administered them into the amniotic fluid space of pregnant rabbits. The intra-amniotic infused fluorescently labeled liposomes were absorbed into the fetal intestinal epithelium. However, the liposomes were not transported to the livers of fetal rabbits. We also revealed that continuous administration of micelles derived from pulmonary surfactants and vernix caseosa protected the small intestine of the rabbit fetus from damage due to surgical intervention. Our results indicate that pulmonary surfactant and vernix caseosa complexes in swallowed amniotic fluid might locally influence fetal intestinal enterocytes. Although the present studies are primarily observational and further studies are needed, our findings elucidate the physiological interactions among pulmonary, dermal-epidermal, and gastrointestinal developmental processes.

  18. [About the estimation of fetal risk by evalution of the serum -- amniotic liqour -- ratio of HPL during the pregnancy (author's transl)].

    PubMed

    Altmann, P; Kucera, H; Spona, J

    1975-08-01

    Radioimmunoassay of human placental lactogen (HPL) is a valuable diagnostic tool for the early recognition of placental dysfunction. But, the rather wide normal range hampers HPL estimations to be used as a good prognostic tool. However, the simultaneous determination of HPL in maternal serum and in amniotic fluid allows a better diagnosis of the fetal well-being. Results of amniotic fluid HPL levels are expressed as percent of maternal serum levels. In this pilot study it was shown that this ratio of maternal serum HPL level to amniotic fluid level is below 10% in normal pregnancies. The ratio is between 10 to 20% in subjects where fetal distress was recorded, and dystrophic babies were observed. The value is above 20% in cases where intrauterine fetal deaths were found. This good correlation of the HPL ratio in maternal serum and amniotic fluid with the outcome of pregnancy offers a new means of using HPL estimation as prognostic tool.

  19. Developmental and Evolutionary Origins of the Amniote Phallus.

    PubMed

    Gredler, Marissa L

    2016-10-01

    An intromittent phallus is used for sperm transfer in most amniote taxa; however, there is extensive variation in external genital morphology within and among the major amniote clades. Amniote phalluses vary in number (paired, single, or rudimentary), spermatic canal morphology (closed tube or open sulcus), and mode of transition between resting and tumescent states (inflation, rotation, eversion, or muscle relaxation). In a phylogenetic context, these varying adult anatomies preclude a clear interpretation for the evolutionary history of amniote external genitalia; as such, multiple hypotheses have been presented for the origin(s) of the amniote phallus. In combination with historic embryological studies, recent comparative developmental analyses have uncovered evidence that, despite extensive morphological variation in adult anatomy, embryonic patterning of the external genitalia is similar among amniotes and begins with emergence of paired swellings adjacent to the cloaca. External genital development in mammals, squamates (lizards, snakes, and amphisbaenians), Rhyncocephalians (tuataras), turtles, crocodilians (alligators, crocodiles, and gharials), and birds proceeds by iterative sequences of budding and fusion events, initiated by emergence of paired swellings adjacent to the embryonic cloaca. Conservation of the embryonic origins, morphogenetic processes, and molecular genetic mechanisms involved in external genital development across Amniota supports derivation from the common ancestor of amniotes, and suggests that lineage-specific divergence of later patterning events underlies the variation observed in extant adult amniote phallus morphology. PMID:27549197

  20. CA 125 in tissues and amniotic fluid during pregnancy.

    PubMed

    Quirk, J G; Brunson, G L; Long, C A; Bannon, G A; Sanders, M M; O'Brien, T J

    1988-09-01

    CA 125 was assayed in amniotic fluid and tissue extracts by immunoradiometric assay, and immunohistochemical studies were performed on paraffin-embedded sections of endometrium, decidua, and fetal membranes with the monoclonal antibody OC 125 used as primary antibody. The concentration of CA 125 in amniotic fluid changes during pregnancy so that levels of 800 to 1000 U/ml are found before 12 weeks. Thereafter, levels of 4000 to 10,000 U/ml are detected routinely. As term approaches, amniotic fluid CA 125 concentrations fall to a range of 1000 to 2000 U/ml. Levels of CA 125 in tissue extracts of secretory endometrium and decidua were 65,000 and 29,500 U/gm of tissue, respectively. CA 125 was readily detected on the apical surfaces of glandular epithelium and in the secretions of endometrial glands obtained throughout the menstrual cycle. It was also detected in the lumina of decidualized glands throughout pregnancy. No antigen was detectable within glandular epithelial cells. We have previously reported high concentrations of CA 125 in chorionic tissue extracts (42,000 U/gm) and low concentrations in amniotic tissue extracts (275 U/gm). In contrast to those findings, immunohistochemical techniques detected CA 125 within the intercellular canaliculi that surround amniotic epithelial cells but not in chorion. We conclude that the likely source of amniotic fluid CA 125 is the decidua and that it gains access to the amniotic fluid via the intercellular canalicular system that traverses the amniotic epithelium.

  1. Phase II trial of whole-cell pertussis vaccine vs an acellular vaccine containing agglutinogens.

    PubMed

    Miller, E; Ashworth, L A; Robinson, A; Waight, P A; Irons, L I

    1991-01-12

    An acellular pertussis vaccine containing agglutinogens 2 and 3, pertussis toxin, and filamentous haemagglutinin was developed by the Centre for Applied Microbiology and Research in the UK. 188 infants were entered into a randomised blind trial and received either the acellular or a whole-cell vaccine, combined with diphtheria and tetanus toxoids, in a 3, 5, and 8-10 month schedule. Local reactions were similar in the two groups but significantly fewer infants had systemic symptoms after the acellular vaccine. Mean log-antibody titres to the agglutinogen and toxin components were higher with the acellular than with the whole-cell vaccine. Persistence of antibodies one year after the third dose was also better in the acellular group. PMID:1670725

  2. Optimizing amniotic membrane tissue banking protocols for ophthalmic use.

    PubMed

    Hettiarachchi, D; Dissanayake, V H W; Goonasekera, H W W

    2016-09-01

    Amniotic membrane (AM) due to its anti-inflammatory, anti-scarring and anti-angiogenic properties is used as corneal and wound grafts. When developing AM tissue banks, cell viability, membrane morphology and genomic stability should be preserved following cryopreservation. To analyze the changes rendered to the AM during the process of cryopreservation by comparing different combinations of standard cryopreservation media; fetal bovine serum (FBS), dimethyl sulfoxide (DMSO), Dulbecco's modified eagle's medium (DMEM) and glycerol at -80 °C and at -196 °C for a period of 6 weeks and at 4 °C in 70 % alcohol for 6 weeks. Following informed consent, placentae of healthy term pregnancies delivered by elective Cesarean section were collected and AM separated into 5 × 5 cm size sections and under sterile conditions stored in 9:1 DMSO:FBS and 1:1 DMEM:Glycerol at -196 and -80 °C for 6 weeks. Similar sections were also stored at 4 °C in 70 % alcohol for 6 weeks. After storage periods following were assessed; AM epithelial cell viability by trypan blue vital stain, epithelial cell proliferation capacity by cell doubling time, membrane morphology by haematoxylin and eosin (H&E) stain and genomic stability by conventional G-banded karyotyping. Human amniotic epithelial cells were cultured in DMEM and 10 % FBS in humidified atmosphere of 5 % carbon dioxide at 37 °C and were characterized using RT-PCR for Octamer-binding protein 4 (Oct-4) and glucose-6-phosphate dehydrogenase (G6PD) genes. All the above parameters were also assessed in fresh AM. AM obtained from 4 term placentae. Mean cell count and mean cell doubling times in days respectively; for fresh AM 3.8 × 10(6); 1.59, after 6 weeks in DMSO:FBS at -196 °C 3.0 × 10(6); 2.38 and at -80 °C 2.1 × 10(6); 1.60, in DMEM:Glycerol at -196 °C 3.6 × 10(6); 2.33 at -80 °C 23 × 10(6); 1.66 and at 4 °C 3.3 × 10(6); 2.14. Histology analysis of the fresh AM showed an intact epithelial

  3. Optimizing amniotic membrane tissue banking protocols for ophthalmic use.

    PubMed

    Hettiarachchi, D; Dissanayake, V H W; Goonasekera, H W W

    2016-09-01

    Amniotic membrane (AM) due to its anti-inflammatory, anti-scarring and anti-angiogenic properties is used as corneal and wound grafts. When developing AM tissue banks, cell viability, membrane morphology and genomic stability should be preserved following cryopreservation. To analyze the changes rendered to the AM during the process of cryopreservation by comparing different combinations of standard cryopreservation media; fetal bovine serum (FBS), dimethyl sulfoxide (DMSO), Dulbecco's modified eagle's medium (DMEM) and glycerol at -80 °C and at -196 °C for a period of 6 weeks and at 4 °C in 70 % alcohol for 6 weeks. Following informed consent, placentae of healthy term pregnancies delivered by elective Cesarean section were collected and AM separated into 5 × 5 cm size sections and under sterile conditions stored in 9:1 DMSO:FBS and 1:1 DMEM:Glycerol at -196 and -80 °C for 6 weeks. Similar sections were also stored at 4 °C in 70 % alcohol for 6 weeks. After storage periods following were assessed; AM epithelial cell viability by trypan blue vital stain, epithelial cell proliferation capacity by cell doubling time, membrane morphology by haematoxylin and eosin (H&E) stain and genomic stability by conventional G-banded karyotyping. Human amniotic epithelial cells were cultured in DMEM and 10 % FBS in humidified atmosphere of 5 % carbon dioxide at 37 °C and were characterized using RT-PCR for Octamer-binding protein 4 (Oct-4) and glucose-6-phosphate dehydrogenase (G6PD) genes. All the above parameters were also assessed in fresh AM. AM obtained from 4 term placentae. Mean cell count and mean cell doubling times in days respectively; for fresh AM 3.8 × 10(6); 1.59, after 6 weeks in DMSO:FBS at -196 °C 3.0 × 10(6); 2.38 and at -80 °C 2.1 × 10(6); 1.60, in DMEM:Glycerol at -196 °C 3.6 × 10(6); 2.33 at -80 °C 23 × 10(6); 1.66 and at 4 °C 3.3 × 10(6); 2.14. Histology analysis of the fresh AM showed an intact epithelial

  4. Whooping cough, twenty years from acellular vaccines introduction.

    PubMed

    Greco, D; Esposito, S; Tozzi, A; Pandolfi, E; Icardi, G; Giammanco, A

    2015-01-01

    Clinical pertussis resulting from infection with B. pertussis is a significant medical and public health problem, despite the huge success of vaccination that has greatly reduced its incidence. The whole cell vaccine had an undeniable success over the last 50 years, but its acceptance was strongly inhibited by fear, only partially justified, of severe side effects, but also, in the Western world, by the difficulty to enter in combination with other vaccines: today multi-vaccine formulations are essential to maintain a high vaccination coverage. The advent of acellular vaccines was greeted with enthusiasm by the public health world: in the Nineties, several controlled vaccine trials were carried out: they demonstrated a high safety and good efficacy of new vaccines. In fact, in the Western world, the acellular vaccines completely replaced the whole cells ones. In the last years, ample evidence on the variety of protection of these vaccines linked to the presence of different antigens of Bordetella pertussis was collected. It also became clear that the protection provided, on average around 80%, leaves every year a significant cohort of vaccinated susceptible even in countries with a vaccination coverage of 95%, such as Italy. Finally, it was shown that, as for the pertussis disease, protection decreases over time, to leave a proportion of adolescents and adults unprotected. Waiting for improved pertussis vaccines, the disease control today requires a different strategy that includes a booster at 5 years for infants, but also boosters for teenagers and young adults, re-vaccination of health care personnel, and possibly of pregnant women and of those who are in contact with infants (cocooning). Finally, the quest for better vaccines inevitably tends towards pertussis acellular vaccines with at least three components, which have demonstrated superior effectiveness and have been largely in use in Italy for fifteen years.

  5. [The dynamics of the amniotic fluid].

    PubMed

    Minh, H N; Douvin, D; Smadja, A; Orcel, L

    1978-01-01

    Morphological study of the ovular membranes showing that under the amnion, until term, there persists perfectly live and active chorial cytotrophoblast covered by a well vascularised and developed parietal decidua. Under the electron microscopy, the apical pole of the amniotic cells as well as their intercellular space are bordered by microvilli. Their basal pole has squat pedicels. The chorion which supports the amnion is avascular. The surface of the cytotrophoblast opposite the chorio-amnion is raised by numerous protuberances in the form of pedicels. The trophoblast is also furrowed by intercellular canals bordered by microvilli and reinforced by desmosomes. Two cellular appearances may be distinguished in the trophoblast: elements similar to the syncytial type with the characteristics of steroid cells and elements of a Langhans type which may play a role in protein synthesis. The decidual cells, in a quinconcial perivascular arrangement, are rich in pinocytotic vesicles. Thus whilst it is undeniable that the foetus plays a certain role in the dynamics of the amniotic fluid by swallowing and urinary excretion, these phenomena are minimal in relation to exchanges through the ovular membranes. Transamniotic passage is the first stage in movement of the fluid. The chorion, by virtue of its plexiform, areolar structure forms the next stop. From the chorion, the circulation continues through the intercellular canals of the trophoblast and the amniotic fluid is thus brought into the well vascularised parietal decidua. In addition to this extracellular circulation, the authors fell that there exists an intracellular passage by uptake and secretion, as evidenced by the high degree of pinocytosis within the cells.

  6. Physiological distal drift in rat molars contributes to acellular cementum formation.

    PubMed

    Tsuchiya, Shinobu; Tsuchiya, Masahiro; Nishioka, Takashi; Suzuki, Osamu; Sasano, Yasuyuki; Igarashi, Kaoru

    2013-08-01

    Occlusal forces may induce the physiological teeth migration in humans, but there is little direct evidence. Rat molars are known to migrate distally during aging, possibly caused by occlusal forces. The purpose of this study was to determine if a reduction in occlusion would decrease teeth migration and affect associated periodontal structures such as cementum. To reduce occlusal forces, the right upper first molar (M1) in juvenile rats was extracted. The transition of the position of upper second molar (M2) and formation of M2 cementum was followed during aging. From the cephalometric analyses, upper M2 was located more anterior compared with the original position with aging after M1 extraction. Associated with this "slowing-down" of the physiological drift, cementum thickness on distal surface, but not on mesial surface, of M2 root was significantly increased. The accumulation of alizarin red as vital stain indicative of calcification, was observed in the distal cementum of M2 root only on the side of M1 extraction. Extraction of M1 that results in less functional loading, distinctly attenuates the physiological drift only in the upper dentition. The decreased physiological drift appears to activate acellular cementum formation only on distal surface of M2 root, perhaps due to reduced mechanical stress associated with the attenuated distal drift. In conclusion, the physiological distal drift in rat molars appears to be largely driven by the occlusal force and also affects the formation of acellular cementum. These findings provide additional direct evidence for an important role of occlusal forces in tooth migration. PMID:23775928

  7. Preclinical evaluations of acellular biological conduits for peripheral nerve regeneration

    PubMed Central

    Liao, I-Chien; Wan, Hua; Qi, Shijie; Cui, Cunqi; Patel, Paarun; Sun, Wendell

    2013-01-01

    Various types of natural biological conduits have been investigated as alternatives to the current surgical standard approach for peripheral nerve injuries. Autologous nerve graft, the current gold standard for peripheral nerve damage, is limited by clinical challenges such as donor-site morbidity and limited availability. The purpose of this study was to evaluate the efficacy of using acellular xenographic conduits (nerve, artery, and dermis) for the repair of a 1.2 cm critical size defect of peripheral nerve in a rodent model. Four months post surgery, the animal group receiving acellular artery as a nerve conduit showed excellent physiological outcome in terms of the prevention of muscle atrophy and foot ulcer. Histological assessment of the bridged site revealed excellent axon regeneration, as opposed to the nonrepaired control group or the group receiving dermal conduit. Finally, the study evaluated the potential improvement via the addition of undifferentiated mesenchymal stem cells into the artery conduit during the bridging procedure. The mesenchymal stem cell–dosed artery conduit group resulted in significantly higher concentration of regenerated axons over artery conduit alone, and exhibited accelerated muscle atrophy rescue. Our results demonstrated that xenographic artery conduits promoted excellent axonal regeneration with highly promising clinical relevance. PMID:23532671

  8. Multilayered implantation using acellular dermal matrix into nude mice.

    PubMed

    Lee, Dong Won; Lee, Myung Chul; Roh, Hyun; Lee, Won Jai

    2014-12-01

    Soft tissue augmentation using acellular dermal matrix has gained popularity to overcome the shortcomings of autogenous and alloplastic materials. Sometimes it needs multilayered stacking to obtain enough volume. In this study, we investigated the efficacy of multilayered implantation using acellular dermal matrix (MatriDerm(®)) for soft tissue augmentation. MatriDerm was implanted subdermally on each side of the dorsum of nude mice (n = 20), stacked two layers thick in the control group and three layers thick in the experimental group. Alterations of thickness, degree of angiogenesis, and collagen and elastin fiber syntheses were observed over 40 days. Three-layered implantation with MatriDerm maintained its volume similarly as in two-layered implantation, although the thickness decreased after 30 days in both groups. At the early stage of implantation, angiogenesis and collagen and elastin fiber syntheses occurred fluently on the central portion, which is the farthest away from the surface in contact with the host tissue. Collagen and elastin fibers became more concentrated over time, and the original structure of MatriDerm could not be maintained due to being replaced with newly formed collagen and elastin fibers 40 days after implantation. Multilayered implantation with MatriDerm is considered appropriate for tissue ingrowth and can be used as a substitute for soft tissue augmentation.

  9. Own Experience From The Use Of A Substitute Of An Allogeneic Acellular Dermal Matrix Revitalized With In Vitro Cultured Skin Cells In Clinical Practice.

    PubMed

    Łabuś, Wojciech; Kawecki, Marek; Glik, Justyna; Maj, Mariusz; Kitala, Diana; Misiuga, Marcelina; Klama-Baryła, Agnieszka; Kraut, Małgorzata; Nowak, Mariusz

    2015-10-01

    As a result of the removal of cells from human allogeneic dermis, a collagen scaffold is obtained, which can be populated de novo with autologous/allogeneic skin cells and transplanted onto the area of skin loss. The optimal method for production of acellular dermal matrices (ADM) has been selected. Three female patients (a mean age of 54 years) were subjected to the transplantation of either autologous or allogeneic keratinocytes and fibroblasts into the holes of acellular dermal matrix (ADM) mesh graft. The method for burn wound treatment based on the use of a viable dermal-epidermal skin substitute (based on ADM and in vitro cultured fibroblasts and keratinocytes) may be the optimal method of burn treatment. PMID:26812752

  10. Own Experience From The Use Of A Substitute Of An Allogeneic Acellular Dermal Matrix Revitalized With In Vitro Cultured Skin Cells In Clinical Practice.

    PubMed

    Łabuś, Wojciech; Kawecki, Marek; Glik, Justyna; Maj, Mariusz; Kitala, Diana; Misiuga, Marcelina; Klama-Baryła, Agnieszka; Kraut, Małgorzata; Nowak, Mariusz

    2015-10-01

    As a result of the removal of cells from human allogeneic dermis, a collagen scaffold is obtained, which can be populated de novo with autologous/allogeneic skin cells and transplanted onto the area of skin loss. The optimal method for production of acellular dermal matrices (ADM) has been selected. Three female patients (a mean age of 54 years) were subjected to the transplantation of either autologous or allogeneic keratinocytes and fibroblasts into the holes of acellular dermal matrix (ADM) mesh graft. The method for burn wound treatment based on the use of a viable dermal-epidermal skin substitute (based on ADM and in vitro cultured fibroblasts and keratinocytes) may be the optimal method of burn treatment.

  11. Dystrophic epidermolysis bullosa associated with amniotic band syndrome.

    PubMed

    Snadecki, Haley; Criscione, Vincent; Jaquith, Alisha; Hay, Beverly; Deng, April; Wiss, Karen

    2014-01-01

    Amniotic band syndrome (ABS) is a term used to describe congenital anomalies that result from the entrapment of a fetus in fibrous bands. We describe two male infants born with features of dystrophic epidermolysis bullosa (DEB) and ABS. These cases add to the few previous reports of simultaneous DEB and ABS. Abnormal type VII collagen in anchoring structures of the amniotic epithelium is a proposed mechanism for loose amniotic bands that entangle the fetus, with an abnormality in the gene that encodes for type VII collagen.

  12. Immunoreactive oestrogens and progesterone in amniotic fluid in twin pregnancies.

    PubMed

    Norman, R J; Joubert, S M

    1985-03-01

    Amniotic fluid concentrations of immunoreactive oestrogens and progesterone were measured at the time of caesarean section in 32 twin pregnancies; 25 women had an elective section and seven were in labour at the time of operation. No significant differences between concentrations in the amniotic fluid of the first and second twin were found in respect of conjugated and unconjugated oestrone, oestradiol, oestriol, oestetrol and unconjugated progesterone either before or during labour. It is unlikely that changes in oestrogens or progesterone in the amniotic fluid are responsible for the selective changes seen in prostaglandins and fetal adrenal steroid during labour in the first twin. PMID:3978052

  13. Atypical presentation of amniotic band sequence.

    PubMed

    Bodamer, O A; Popek, E J; Bacino, C

    2001-04-22

    Amniotic Band Sequence (ABS) is a disruption sequence that results in a variable group of abnormalities secondary to the disruption process and subsequent deformations. The incidence of ABS ranges from 1:1,200 to 1:15,000 live-born, and is even higher in still-born [Froster and Baird, 1993: Am J Med Genet 46:497-500]. The pathophysiology of ABS remains controversial, but a close look to critical periods of embryogenesis and/or organogenesis has helped in understanding pathogenetic mechanisms leading to the ABS disruption. The abnormalities are typically limited to external structures; however, associated internal malformations as seen in the case reported here may occur [Hunter and Carpenter, 1986: Am J Med Genet 24:691-700]. The prognosis depends on the severity of the abnormalities and the involvement of internal organs [Froster and Baird; 1993: Am J Med Genet 46:497-500; Levy, 1998: Ped Rev 19:249].

  14. Hiccups and amniotic fluid regulation in early pregnancy.

    PubMed

    Murchison, Andrew G

    2015-05-01

    Hiccups are an unexplained phenomenon and a subject of medical curiosity. They arise through a reflex arc with central control at the level of the medulla, and their primary physiological effect is the generation of negative intra-thoracic pressure. This paper presents the hypothesis that hiccups serve a purpose during the first half of gestation, when they are most prevalent; namely, that they promote amniotic fluid influx to the primitive gut, allowing fluid to be transferred to the foetal and then maternal vasculature. Furthermore, hiccups could be provoked by increasing amniotic fluid volume and pressure, and act in a regulatory capacity. This hypothesis could be tested by studying foetal movements in the first half of gestation, and assessing whether there is correlation with amniotic fluid flux in the developing gut. Ascertaining whether hiccups increase in frequency with increasing amniotic fluid volume would provide evidence for or against a regulatory function.

  15. Extended Eden model reproduces growth of an acellular slime mold

    NASA Astrophysics Data System (ADS)

    Wagner, Geri; Halvorsrud, Ragnhild; Meakin, Paul

    1999-11-01

    A stochastic growth model was used to simulate the growth of the acellular slime mold Physarum polycephalum on substrates where the nutrients were confined in separate drops. Growth of Physarum on such substrates was previously studied experimentally and found to produce a range of different growth patterns [Phys. Rev. E 57, 941 (1998)]. The model represented the aging of cluster sites and differed from the original Eden model in that the occupation probability of perimeter sites depended on the time of occupation of adjacent cluster sites. This feature led to a bias in the selection of growth directions. A moderate degree of persistence was found to be crucial to reproduce the biological growth patterns under various conditions. Persistence in growth combined quick propagation in heterogeneous environments with a high probability of locating sources of nutrients.

  16. Protection against pertussis by Takeda's acellular pertussis vaccine: household contact studies in Kawasaki City, Japan.

    PubMed

    Kato, T; Kaku, H; Arimoto, Y

    1988-01-01

    To evaluate the vaccine efficacy of an acellular pertussis vaccine which has been in clinical use in Japan since 1981, a retrospective study was performed by a questionnaire survey of secondary pertussis attacks through family contact in 146 children with pertussis diagnosed in the period from January 1981 through May 1988. In this study, Takeda's acellular vaccine which contains a high level of FHA, low level of PT and a small amount of agglutinogen, was evaluated. Secondary pertussis attacks through family contact were found in 17 of 27 siblings (62.9%) not immunized with pertussis vaccine. On the other hand, 26 siblings immunized with Takeda's acellular vaccine were exposed to pertussis through family contact and a secondary attack was seen in only one of them (3.8%). The present study revealed an efficacy rate of 93.9% for Takeda's acellular pertussis vaccine. PMID:3078808

  17. Protection against pertussis by acellular pertussis vaccines (Takeda, Japan): household contact studies in Kawasaki City, Japan.

    PubMed

    Kato, T; Goshima, T; Nakajima, N; Kaku, H; Arimoto, Y; Hayashi, F

    1989-12-01

    To evaluate the vaccine efficacy of an acellular pertussis vaccine which has been in clinical use in Japan since 1981, a retrospective study was performed by a questionnaire survey of secondary pertussis attacks through family contact in 146 children with pertussis diagnosed in the period from January 1981 through May 1988. In this study, acellular vaccine made by Takeda Pharmaceutical Company, which contains a high level of FHA (filamentous hemagglutinin), a low level of PT (pertussis toxin) and a small amount of agglutinogen, was evaluated. Secondary pertussis attacks through family contact were found in 17 of 29 siblings (58.6%) not immunized with pertussis vaccine. On the other hand, 27 siblings immunized with Takeda's acellular vaccine were exposed to pertussis through family contact and a secondary attack was seen in only one of them (3.7%). The present study revealed an efficacy rate of 93.7% for Takeda's acellular pertussis vaccine. PMID:2516396

  18. [Protection against pertussis by Japanese T type acellular pertussis vaccine: household contact study in Kawasaki City].

    PubMed

    Kato, T; Matsuyoshi, S; Goshima, T; Nakajima, N; Yamamoto, H; Arimoto, Y; Kaku, H; Hayashi, F

    1989-09-01

    To evaluate the vaccine efficacy of acellular pertussis vaccine which has been in clinical use in Japan since 1981, a retrospective study was made by a questionnaire from secondary pertussis attack through family contact in 149 children with pertussis diagnosed in the period from January 1981 through May 1988. In this study, Takeda's acellular vaccine which contains a high level of FHA, low level of PT and a small amount of agglutinogen, was evaluated. Secondary pertussis attacks through family contact were found in 17 of 29 siblings (58.6%) not immunized with pertussis vaccine. On the other hand of the siblings immunized with Takeda's acellular vaccine 27 were exposed to pertussis through family contact and a secondary attack was seen in only one of them (3.4%). The present study revealed an efficacy rate of 94.2% for the Takeda's acellular pertussis vaccine. PMID:2509597

  19. AMNIOTIC FLUID HEAT SHOCK PROTEIN 70 CONCENTRATION IN HISTOLOGIC CHORIOAMNIONITIS, TERM AND PRETERM PARTURITION

    PubMed Central

    Chaiworapongsa, Tinnakorn; Erez, Offer; Kusanovic, Juan Pedro; Vaisbuch, Edi; Mazaki-Tovi, Shali; Gotsch, Francesca; Than, Nandor Gabor; Mittal, Pooja; Kim, Yeon Mee; Camacho, Natalia; Edwin, Samuel; Gomez, Ricardo; Hassan, Sonia S.; Romero, Roberto

    2008-01-01

    Objective Heat shock protein (HSP) 70, a conserved member of the stress protein family, is produced in almost all cell types in response to a wide range of stressful stimuli and their production has a survival value. Evidence suggests that extra-cellular HSP70 is involved in the activation of the innate and adaptive immune response. Furthermore, increased mRNA expression of HSP 70 was observed in human fetal membranes following endotoxin stimulation. This study was conducted to determine the changes in amniotic fluid HSP70 concentrations during pregnancy, term and preterm parturition, intra-amniotic infection (IAI), and histologic chorioamnionitis. Study design A cross-sectional study was conducted in 376 pregnant women in the following groups: 1) women with a normal pregnancy that were classified in the following categories: a) women in the mid-trimester (14–18 weeks) who underwent amniocentesis for genetic indications and delivered normal infants at term (n=72); b) women at term not in labor (n=23); and c) those at term in labor (n=48); 2) women with spontaneous preterm labor and intact membranes that were subdivided into the following categories: a) preterm labor who delivered at term without IAI (n=42), b) preterm labor who delivered preterm without IAI (n=57), and c) preterm labor and delivery with IAI (n=30); and 3) women with preterm prelabor rupture of membranes (PROM) with (n=50) and without (n=54) IAI. Among patients with preterm labor with intact membranes and preterm PROM who delivered within 72 hours of amniocentesis, placenta, umbilical cord and chorioamniotic membranes were collected and assessed for the presence or absence of acute inflammatory lesions in the extra-placental membranes (histologic chorioamnionitis) and/or umbilical cords (funisitis). HSP70 concentrations in amniotic fluid were determined using a sensitive and specific immunoassay. Non-parametric statistics were used for analysis. A p value <0.05 was considered statistically

  20. Brainless but Multi-Headed: Decision Making by the Acellular Slime Mould Physarum polycephalum.

    PubMed

    Beekman, Madeleine; Latty, Tanya

    2015-11-20

    Because of its peculiar biology and the ease with which it can be cultured, the acellular slime mould Physarum polycephalum has long been a model organism in a range of disciplines. Due to its macroscopic, syncytial nature, it is no surprise that it has been a favourite amongst cell biologists. Its inclusion in the experimental tool kit of behavioural ecologists is much more recent. These recent studies have certainly paid off. They have shown that, for an organism that lacks a brain or central nervous system, P. polycephalum shows rather complex behaviour. For example, it is capable of finding the shortest path through a maze, it can construct networks as efficient as those designed by humans, it can solve computationally difficult puzzles, it makes multi-objective foraging decisions, it balances its nutrient intake and it even behaves irrationally. Are the slime mould's achievements simply "cute", worthy of mentioning in passing but nothing to take too seriously? Or do they hint at the fundamental processes underlying all decision making? We will address this question after reviewing the decision-making abilities of the slime mould.

  1. Brainless but Multi-Headed: Decision Making by the Acellular Slime Mould Physarum polycephalum.

    PubMed

    Beekman, Madeleine; Latty, Tanya

    2015-11-20

    Because of its peculiar biology and the ease with which it can be cultured, the acellular slime mould Physarum polycephalum has long been a model organism in a range of disciplines. Due to its macroscopic, syncytial nature, it is no surprise that it has been a favourite amongst cell biologists. Its inclusion in the experimental tool kit of behavioural ecologists is much more recent. These recent studies have certainly paid off. They have shown that, for an organism that lacks a brain or central nervous system, P. polycephalum shows rather complex behaviour. For example, it is capable of finding the shortest path through a maze, it can construct networks as efficient as those designed by humans, it can solve computationally difficult puzzles, it makes multi-objective foraging decisions, it balances its nutrient intake and it even behaves irrationally. Are the slime mould's achievements simply "cute", worthy of mentioning in passing but nothing to take too seriously? Or do they hint at the fundamental processes underlying all decision making? We will address this question after reviewing the decision-making abilities of the slime mould. PMID:26189159

  2. Stem cells from amniotic fluid--Potential for regenerative medicine.

    PubMed

    Loukogeorgakis, Stavros P; De Coppi, Paolo

    2016-02-01

    Regenerative medicine has recently been established as an emerging field focussing on repair, replacement or regeneration of cells, tissues and whole organs. The significant recent advances in the field have intensified the search for novel sources of stem cells with potential for therapy. Recently, researchers have identified the amniotic fluid as an untapped source of stem cells that are multipotent, possess immunomodulatory properties and do not have the ethical and legal limitations of embryonic stem cells. Stem cells from the amniotic fluid have been shown to differentiate into cell lineages representing all three embryonic germ layers without generating tumours, which make them an ideal candidate for tissue engineering applications. In addition, their ability to engraft in injured organs and modulate immune and repair responses of host tissues suggest that transplantation of such cells may be useful for the treatment of various degenerative and inflammatory diseases affecting major tissues/organs. This review summarises the evidence on amniotic fluid cells over the past 15 years and explores the potential therapeutic applications of amniotic fluid stem cells and amniotic fluid mesenchymal stem cells.

  3. The Amniotic Membrane: Development and Potential Applications - A Review.

    PubMed

    Favaron, P O; Carvalho, R C; Borghesi, J; Anunciação, A R A; Miglino, M A

    2015-12-01

    Foetal membranes are essential tissues for embryonic development, playing important roles related to protection, breathing, nutrition and excretion. The amnion is the innermost extraembryonic membrane, which surrounds the foetus, forming an amniotic sac that contains the amniotic fluid (AF). In recent years, the amniotic membrane has emerged as a potential tool for clinical applications and has been primarily used in medicine in order to stimulate the healing of skin and corneal diseases. It has also been used in vaginal reconstructive surgery, repair of abdominal hernia, prevention of surgical adhesions and pericardium closure. More recently, it has been used in regenerative medicine because the amniotic-derived stem cells as well as AF-derived cells exhibit cellular plasticity, angiogenic, cytoprotective, immunosuppressive properties, antitumoural potential and the ability to generate induced pluripotent stem cells. These features make them a promising source of stem cells for cell therapy and tissue engineering. In this review, we discussed the development of the amnion, AF and amniotic cavity in different species, as well as the applicability of stem cells from the amnion and AF in cellular therapy.

  4. Identification of the cold-insoluble globulin of plasma in amniotic fluid.

    PubMed

    Chen, A B; Mosesson, M W; Solish, G I

    1976-08-01

    A fetal and adult plasma protein known as the cold-insoluble globulin (CIg) of plasma has been identified in amniotic fluid. Its concentration relative to the total protein in amniotic fluid is several times higher than that in adult or fetal plasma, suggesting that it arises from amniotic tissues.

  5. Comparative biological activities of acellular pertussis vaccines produced by Kitasato.

    PubMed

    Watanabe, M; Izumiya, K; Sato, T; Yoshino, K; Nakagawa, N; Ohoishi, M; Hoshino, M

    1991-04-01

    The quality of 14 lots of acellular pertussis-diphtheria-tetanus (AC-PDT) vaccines manufactured by the Kitasato Institute during the period 1987-1990 were investigated. The geometric means of HSU, LPU, and BWDU were 0.078, 0.257, and 7.33 per ml respectively. The potency was higher than 14 IU per ml. These results indicated the consistency of the Kitasato AC-PDT vaccines. The antibody response to the AC-PDT vaccines was measured in primary and secondary vaccinated mice by ELISA. IgG antibody response to FHA and PT was obtained in all immunized mice (P less than 0.001) after the primary injection. In contrast, IgG antibody response to fimbriae 2 showed a significant titer rise (P less than 0.001) after the booster injection. The results indicated that the Kitasato AC-P vaccines consisted of protein, PT and FHA as the major antigens, and a little agglutinogen as the minor antigen. PMID:1798236

  6. Characterization of co-purified acellular pertussis vaccines.

    PubMed

    Xu, Yinghua; Tan, Yajun; Asokanathan, Catpagavalli; Zhang, Shumin; Xing, Dorothy; Wang, Junzhi

    2015-01-01

    Whole-cell pertussis vaccines (WPVs) have been completely replaced by the co-purified acellular vaccines (APVs) in China. To date few laboratory studies were reported for co-purified APVs in terms of their antigenic composition and protective immune responses. To further understand the antigenic composition in co-purified APVs, in the present study 2-dimensional gel electrophoresis-based proteomic technology was used to analyze the composition of co-purified APVs. The results showed that besides the main antigens pertussis toxin (PT) and filamentous hemagglutinin (FHA), co-purified APVs also contained pertactin (PRN), fimbriae (FIM) 2and3 and other minor protein antigens. Of the 9 proteins identified, 3 were differentially presented in products from manufacturer 1 and manufacturer 2. Compared with WPVs and purified APVs, co-purified APVs induced a mixed Th1/Th2 immune response with more toward to a Th1 response than the purified APVs in this study. These results hint that different immune mechanisms might be involved in protection induced by co-purified and purified APVs.

  7. Characterization of co-purified acellular pertussis vaccines

    PubMed Central

    Xu, Yinghua; Tan, Yajun; Asokanathan, Catpagavalli; Zhang, Shumin; Xing, Dorothy; Wang, Junzhi

    2015-01-01

    Whole-cell pertussis vaccines (WPVs) have been completely replaced by the co-purified acellular vaccines (APVs) in China. To date few laboratory studies were reported for co-purified APVs in terms of their antigenic composition and protective immune responses. To further understand the antigenic composition in co-purified APVs, in the present study 2-dimensional gel electrophoresis-based proteomic technology was used to analyze the composition of co-purified APVs. The results showed that besides the main antigens pertussis toxin (PT) and filamentous hemagglutinin (FHA), co-purified APVs also contained pertactin (PRN), fimbriae (FIM) 2and3 and other minor protein antigens. Of the 9 proteins identified, 3 were differentially presented in products from manufacturer 1 and manufacturer 2. Compared with WPVs and purified APVs, co-purified APVs induced a mixed Th1/Th2 immune response with more toward to a Th1 response than the purified APVs in this study. These results hint that different immune mechanisms might be involved in protection induced by co-purified and purified APVs. PMID:25610957

  8. The potential of amniotic membrane/amnion-derived cells for regeneration of various tissues.

    PubMed

    Toda, Ayaka; Okabe, Motonori; Yoshida, Toshiko; Nikaido, Toshio

    2007-11-01

    Regenerative medicine is a new field based on the use of stem cells to generate biological substitutes and improve tissue functions, restoring damaged tissue with high proliferability and differentiability. It is of interest as a potential alternative to complicated tissue/organ transplantation. Recently, amnion-derived cells have been reported to have multipotent differentiation ability, and these cells have attracted attention as a cell source for cell-transplantation therapy. The amnion possesses considerable advantageous characteristics: the isolated cells can differentiate into all three germ layers; they have low immunogenicity and anti-inflammatory functions; and they do not require the sacrifice of human embryos for their isolation, thus avoiding the current controversies associated with the use of human embryonic stem cells. Moreover, we developed human amniotic cell-sheets using a novel culture surface coated with a noncytotoxic, temperature-responsive elastic protein-based polymer. We also generated a "hyper-dry-amnion", which has already been applied clinically in the ophthalmological field. Compared to cryopreserved fresh amnion, "hyper-dry-amnion" is easy to handle and has started to bring good results to patients. These materials from the amnion are also expected to open a new field in tissue engineering. Thus, amnion, which had been discarded after parturition, has started to be appreciated as an attractive material in the field of regenerative medicine. In this review, the most recent and relevant clinical and experimental data about the use of amniotic membrane and cells derived from it are described.

  9. Effect of Amniotic Fluid Stem Cells and Amniotic Fluid Cells on the Wound Healing Process in a White Rat Model

    PubMed Central

    Choi, Dong Sik; Cho, Young Kyoo; Kim, Taek Kyun; Lee, Jeong Woo; Choi, Kang Young; Chung, Ho Yun; Cho, Byung Chae; Byun, Jin Suk

    2013-01-01

    Background Amniotic-fluid-derived stem cells and amniocytes have recently been determined to have wound healing effects, but their mechanism is not yet clearly understood. In this study, the effects of amniotic fluid stem cells and amniocytes on wound healing were investigated through animal experiments. Methods On the back of Sprague-Dawley rats, four circular full-thickness skin wounds 2 cm in diameter were created. The wounds were classified into the following four types: a control group using Tegaderm disc wound dressings and experimental groups using collagen discs, amniotic fluid stem cell discs, and amniocyte discs. The wounds were assessed through macroscopic histological examination and immunohistochemistry over a period of time. Results The amniotic fluid stem cell and amniocyte groups showed higher wound healing rates compared with the control group; histologically, the inflammatory cell invasion disappeared more quickly in these groups, and there was more significant angiogenesis. In particular, these groups had significant promotion of epithelial cell reproduction, collagen fiber formation, and angiogenesis during the initial 10 days of the wound healing process. The potency of transforming growth factor-β and fibronectin in the experimental group was much greater than that in the control group in the early stage of the wound healing process. In later stages, however, no significant difference was observed. Conclusions The amniotic fluid stem cells and amniocytes were confirmed to have accelerated the inflammatory stage to contribute to an enhanced cure rate and shortened wound healing period. Therefore, they hold promise as wound treatment agents. PMID:24086800

  10. Angiogenic response induced by acellular brain scaffolds grafted onto the chick embryo chorioallantoic membrane.

    PubMed

    Ribatti, Domenico; Conconi, Maria Teresa; Nico, Beatrice; Baiguera, Silvia; Corsi, Patrizia; Parnigotto, Pier Paolo; Nussdorfer, Gastone G

    2003-10-31

    The repair and regeneration of injured tissues and organs depend on the re-establishment of the blood flow needed for cellular infiltration and metabolic support. Among the various materials used in tissue reconstruction, acellular scaffolds have recently been utilized. In this study, we investigated the angiogenic response induced by acellular brain scaffolds implanted in vivo onto the chick embryo chorioallantoic membrane (CAM), a useful model for such investigations. The results show that acellular brain scaffolds are able to induce a strong angiogenic response, comparable to that of fibroblast growth factor-2 (FGF-2), a well known angiogenic cytokine. The response may be considered dependent on a direct angiogenic effect exerted by the scaffold, because no inflammatory infiltrate was detectable in CAM's mesenchyme beneath the implant. Acellular brain scaffolds might induce the release of endogenous angiogenic factors, such as FGF-2 and vascular endothelial growth factor (VEGF) released from the extracellular matrix of the developing CAM. In addition, the angiogenic response may depend, in part, also on the presence in the acellular matrix of transforming growth factor beta 1 (TGFbeta1).

  11. Incidence, diagnosis and pathophysiology of amniotic fluid embolism.

    PubMed

    Ito, F; Akasaka, J; Koike, N; Uekuri, C; Shigemitsu, A; Kobayashi, H

    2014-10-01

    Amniotic fluid embolism (AFE) is a rare clinical entity, sometimes fatal. A review was conducted to describe the frequency, diagnosis and pathophysiology of AFE. The reported incidences ranged from 1.9 cases per 100,000 maternities (UK) to 6.1 per 100,000 maternities (Australia), which can vary considerably, depending on the period, region of study and the definition. Although the development of amniotic fluid-specific markers would have an impact on early diagnosis, definition of AFE based on these markers is not widely accepted. To date, immunological mechanisms, amniotic fluid-dependent anaphylactic reaction and complement activation, have been proposed as potential pathogenetic and pathophysiological mechanisms. Immune cell activation induced through complement activation may be associated with the mechanism that immediately initiates maternal death, only in susceptible individuals. This review will focus on advances in the field of AFE biology and discuss the prevalence, diagnosis and pathophysiology of AFE.

  12. Intra-amniotic Infection Caused by Capnocytophaga Species

    PubMed Central

    Kawazoe, Kyoko; Sato, Yasumasa; Izumi, Koji; Tamaya, Teruhiko

    1996-01-01

    Background: Capnocytophaga species are common oral pathogens and infrequent causes of systemic infection in patients with compromised host. The isolation of this organism suggests an oral source of infection. Case: A 32-year-old woman was admitted at 23 weeks gestation in preterm premature rupture of the membranes. She subsequently developed signs of clinical intra-amniotic infection, including fever, fetal tachycardia, and uterine tenderness. Bacteriologic studies of the amniotic fluid by trans-abdominal amniocentesis and subchorionic placental cultures yielded Capnocytophaga species. On review of the patient's history, a temporal relation was noted between orogenital contact and onset of clinical infection. Thorough evaluation of the patient, including dental examination with periodontal cultures, did not reveal an obvious source of infection. However, significant periodontal disease was identified in her partner and Capnocytophaga species were isolated from her partner. Conclusion: This case suggests that intra-amniotic infection may have been due to an ascending infection after orogenital contact. PMID:18476112

  13. Toward consistent terminology: assessment and reporting of amniotic fluid volume.

    PubMed

    Moise, Kenneth J

    2013-10-01

    Amniotic fluid is typically measured by ultrasound using the amniotic fluid index (AFI) or the maximum vertical pocket (MVP). Although both parameters correlate poorly with the actual amniotic fluid volume measured with dye-dilution methods, cross-sectional studies have been used to establish gestational norms. The current acceptable definition of polyhydramnios in the late second and the third trimester in both singleton and multiple gestations is a MVP > 8 cm, while the definition of oligohydramnios is a MVP < 2 cm. The pocket to be measured should exclude the umbilical cord or fetal parts. Randomized clinical trials have indicated that defining oligohydramnios as a MVP < 2 cm will result in fewer obstetrical interventions and similar perinatal outcomes when compared to an AFI < 5 cm.

  14. Ex vivo evaluation of acellular and cellular collagen-glycosaminoglycan flowable matrices.

    PubMed

    Hodgkinson, Tom; Bayat, Ardeshir

    2015-08-01

    Collagen-glycosaminoglycan flowable matrices (CGFM) are increasingly finding utility in a diversifying number of cutaneous surgical procedures. Cellular in-growth and vascularisation of CGFM remain rate-limiting steps, increasing cost and decreasing efficacy. Through in vitro and ex vivo culture methods, this study investigated the improvement of injectable CGFM by the incorporation of hyaluronan (HA) and viable human cells (primary human dermal fibroblasts (PHDFs) and bone marrow-derived mesenchymal stem cells (BM-MSCs)). Ex vivo investigations included the development and evaluation of a human cutaneous wound healing model for the comparison of dermal substitutes. Cells mixed into the Integra Flowable Wound Matrix (IFWM), a commercially available CGFM, were confirmed to be viable and proliferative through MTT assays (p  <  0.05). PHDFs proliferated with greater rapidity than BM-MSCs up to 1 week in culture (p  <  0.05), with PHDF proliferation further enhanced by HA supplementation (p  <  0.05). After scaffold mixing, gene expression was not significantly altered (qRT-PCR). PHDF and BM-MSC incorporation into ex vivo wound models significantly increased re-epithelialisation rate, with maximal effects observed for BM-MSC supplemented IFWM. HA supplementation to PHDF populated IFWM increased re-epithelialisation but had no significant effect on BM-MSC populated IFWM. In conclusion, when combined with PHDF, HA increased re-epithelialisation in IFWM. BM-MSC incorporation significantly improved re-epithelialisation in ex vivo models over acellular and PHDF populated scaffolds. Viable cell incorporation into IFWM has potential to significantly benefit wound healing in chronic and acute cutaneous injuries by allowing a point-of-care matrix to be formed from autologous or allogenic cells and bioactive molecules. PMID:26181360

  15. Case report: a fatal case of amniotic fluid embolism.

    PubMed

    Azzan, B B

    1979-01-01

    A primigravida was induced for PET, the liquor was meconium stained; she was put on oxytocin in-fussion and developed hypertonic uterine action. She then had an amniotic fluid embolism which presented clinically as profound shock, dyspnoea, tachycardia, cyanosis, hypotension and pyrexia. The patient was delivered by vacuum extraction. The picture was further complicated by pulmonary oedema intravascular microcoagulation and anuria. She deteriorated rapidly and died despite treatment with double strength plasma (in the absence of fibrinogen), massive hydrocortiosone therapy, blood transfusion amd sub-total hysterectomy. Post mortem findings in the lungs confirmed amniotic fluid embolism.

  16. Tissue engineering of the small intestine by acellular collagen sponge scaffold grafting.

    PubMed

    Hori, Y; Nakamura, T; Matsumoto, K; Kurokawa, Y; Satomi, S; Shimizu, Y

    2001-01-01

    Tissue engineering of the small intestine will prove a great benefit to patients suffering from short bowel disease. However cell seeding in tissue engineering, such as fetal cell use, is accompanied by problems of ethical issues, rejection, and short supply. To overcome these problems, we carried out an experimental study on tissue engineering of the small intestine by acellular collagen sponge scaffold grafting. We resected the 5 cm long jejunum from beagle dogs and reconstructed it by acellular collagen sponge grafting with a silicon tube stent. The graft was covered with the omentum. At 1 month after operation, the silicon stent was removed endoscopically. Animals were sacrificed 1 and 4 months after operation, and were examined microscopically. Neo-intestinal regeneration was observed and the intestinal mucosa covered the luminal side of the regenerated intestine across the anastomosis. Thus, the small intestine was regenerated by tissue engineering technology using an acellular collagen sponge scaffold.

  17. Pertactin deficient Bordetella pertussis present a better fitness in mice immunized with an acellular pertussis vaccine.

    PubMed

    Hegerle, N; Dore, G; Guiso, N

    2014-11-20

    Bordetella pertussis is the etiologic agent of whooping cough and has been the target of vaccination for over fifty years. The latest strategies include the use of acellular pertussis vaccines that induce specific immunity against few virulence factors amongst which pertactin is included in three and five component acellular pertussis vaccines. Recently, it has been reported that B. pertussis clinical isolates loose the production of this adhesin in regions reaching high vaccine coverage with vaccines targeting this virulence factor. We here demonstrate that isolates not producing pertactin are capable of sustaining longer infection as compared to pertactin producing isolates in an in vivo model of acellular pertussis immunization. Loosing pertactin production might thus provide a selective advantage to these isolates in this background, which could account for the upraise in prevalence of these pertactin deficient isolates in the population.

  18. Acellular comet assay: a tool for assessing variables influencing the alkaline comet assay.

    PubMed

    Kennedy, Erin K; McNamee, James P; Prud'homme Lalonde, Louise; Jones, Trevor; Wilkinson, Diana

    2012-01-01

    In this study, an acellular modification to the alkaline comet assay to further evaluate key variables within the assay that may influence the outcome of genotoxicity studies is described. This acellular comet assay can detect differences of 0.2 Gy of (60)Co gamma-ray radiation between 0 and 1 Gy and differences of 1 Gy between 0 and 8 Gy; thus, this assay is applicable for a wide range of DNA damage levels. It is also shown that DNA damage from different radiation energies was not significantly different from (60)Co gamma-ray. This assay displayed a statistical increase in DNA damage due to uncontrolled exposure to natural light; however, the slope of the dose-response curve for light-exposed samples was similar to that for samples protected from light. A comparison of the alkaline comet assay with the acellular comet assay allowed for the intrinsic repair capacity of the alkaline comet assay to be quantified.

  19. Diversity in primary palate ontogeny of amniotes revealed with 3D imaging.

    PubMed

    Abramyan, John; Thivichon-Prince, Beatrice; Richman, Joy Marion

    2015-05-01

    The amniote primary palate encompasses the upper lip and the nasal cavities. During embryonic development, the primary palate forms from the fusion of the maxillary, medial nasal and lateral nasal prominences. In mammals, as the primary palate fuses, the nasal and oral cavities become completely separated. Subsequently, the tissue demarcating the future internal nares (choanae) thins and becomes the bucconasal membrane, which eventually ruptures and allows for the essential connection of the oral and nasal cavities to form. In reptiles (including birds), the other major amniote group, primary palate ontogeny is poorly studied with respect to prominence fusion, especially the formation of a bucconasal membrane. Using 3D optical projection tomography, we found that the prominences that initiate primary palate formation are similar between mammals and crocodilians but distinct from turtles and lizards, which are in turn similar to each other. Chickens are distinct from all non-avian lineages and instead resemble human embryos in this aspect. The majority of reptiles maintain a communication between the oral and nasal cavities via the choanae during primary palate formation. However, crocodiles appear to have a transient separation between the oral and nasal cavities. Furthermore, the three lizard species examined here, exhibit temporary closure of their external nares via fusion of the lateral nasal prominences with the frontonasal mass, subsequently reopening them just before hatching. The mechanism of the persistent choanal opening was examined in chicken embryos. The mesenchyme posterior/dorsal to the choana had a significant decline in proliferation index, whereas the mesenchyme of the facial processes remained high. This differential proliferation allows the choana to form a channel between the oral and nasal cavities as the facial prominences grow and fuse around it. Our data show that primary palate ontogeny has been modified extensively to support the array of

  20. Autologous transplantation of amniotic fluid-derived mesenchymal stem cells into sheep fetuses.

    PubMed

    Shaw, S W Steven; Bollini, Sveva; Nader, Khalil Abi; Gastaldello, Annalisa; Gastadello, Annalisa; Mehta, Vedanta; Filppi, Elisa; Cananzi, Mara; Gaspar, H Bobby; Qasim, Waseem; De Coppi, Paolo; David, Anna L

    2011-01-01

    Long-term engraftment and phenotype correction has been difficult to achieve in humans after in utero stem cell transplantation mainly because of allogeneic rejection. Autologous cells could be obtained during gestation from the amniotic fluid with minimal risk for the fetus and the mother. Using a sheep model, we explored the possibility of using amniotic fluid mesenchymal stem cells (AFMSCs) for autologous in utero stem cell/gene therapy. We collected amniotic fluid (AF) under ultrasound-guided amniocentesis in early gestation pregnant sheep (n = 9, 58 days of gestation, term = 145 days). AFMSCs were isolated and expanded in all sampled fetal sheep. Those cells were transduced using an HIV vector encoding enhanced green fluorescent protein (GFP) with 63.2% (range 38.3-96.2%) transduction efficiency rate. After expansion, transduced AFMSCs were injected into the peritoneal cavity of each donor fetal sheep at 76 days under ultrasound guidance. One ewe miscarried twin fetuses after amniocentesis. Intraperitoneal injection was successful in the remaining 7 fetal sheep giving a 78% survival for the full procedure. Tissues were sampled at postmortem examination 2 weeks later. PCR analysis detected GFP-positive cells in fetal tissues including liver, heart, placenta, membrane, umbilical cord, adrenal gland, and muscle. GFP protein was detected in these tissues by Western blotting and further confirmed by cytofluorimetric and immunofluorescence analyses. This is the first demonstration of autologous stem cell transplantation in the fetus using AFMSCs. Autologous cells derived from AF showed widespread organ migration and could offer an alternative way to ameliorate prenatal congenital disease.

  1. [Pertussis vaccines: acellular versus whole cell. Perhaps a return to the past?].

    PubMed

    Cofré, José

    2015-10-01

    The resurgence of pertussis in the world and in our country has questioned the effectiveness of cellular and acellular vaccines. The reason why pertussis has not been controlled or eliminated after 70 years of implementation of the vaccination is probably multifactorial. This article, on the basis of questions and answers, describes the benefits and limitations of both cellular and acellular vaccines and suggests new strategies of vaccination in childhood. It is a fact that the currently applied vaccination does not eliminate the circulation of Bordetella pertussis in the community. Perhaps the introduction of vaccines with live B. pertussis, inhalation, will be able to eliminate the disease around the world. PMID:26633113

  2. Histochemical and morphological studies on a new type of acellular cartilage.

    PubMed

    Junqueira, L C; Toledo, O M; Montes, G S

    1983-01-01

    A new type of cartilage was found participating in a valve-like system inside the conus arteriosus of the fresh water sting ray, Potamotrygon sp.. This cartilage possesses no chondrocytes and its matrix is perforated by vascular channels that ramify dendritically forming canaliculi. The acellular cartilage does not possess perichondrium but, rather, it is attached to a basement membrane-like structure. The cartilaginous matrix contains collagen fibrils that strongly interact with the chondroitin sulfate of the ground substance. The histochemical and biochemical findings suggest that not all of the glycosaminoglycans present in the acellular cartilage are bound to protein cores to form proteoglycans. PMID:6407464

  3. [Amniotic fluid embolism as a cause of maternal death].

    PubMed

    Nadeev, A P; Zhukova, V A; Ageeva, T A; Drobinskaya, A N; Travin, M A; Karpov, M A; Savchenko, S V; Chikinev, Yu V; Polyakevich, A S

    2015-01-01

    This article reports the case of death of a puerperal woman resulting from amniotic fluid embolism. The diagnosis was established based on the results of the pathohistological study that revealed the presence of mucoproteides and epithelial scales in pulmonary blood vessels and capillaries. PMID:26856060

  4. [Amniotic fluid embolism as a cause of maternal death].

    PubMed

    Nadeev, A P; Zhukova, V A; Ageeva, T A; Drobinskaya, A N; Travin, M A; Karpov, M A; Savchenko, S V; Chikinev, Yu V; Polyakevich, A S

    2015-01-01

    This article reports the case of death of a puerperal woman resulting from amniotic fluid embolism. The diagnosis was established based on the results of the pathohistological study that revealed the presence of mucoproteides and epithelial scales in pulmonary blood vessels and capillaries.

  5. Biochemical markers of trisomy 21 in amniotic fluid.

    PubMed

    Spencer, K; Muller, F; Aitken, D A

    1997-01-01

    In a study of amniotic fluid from 91 Down's syndrome cases and 240 controls, we have shown that the median values of four biochemical markers (AFP, total hCG, free beta hCG, and unconjugated oestriol) in the amniotic fluid of pregnancies affected by Down's syndrome on the whole reflect those observed in the maternal serum of affected cases. The median MOM for AFP was lower than average (0.56), as was that for unconjugated oestriol (0.55), whilst those for total hCG (1.82) and free beta hCG (2.10) were increased on average. The width of the distribution of marker levels in amniotic fluid is similar to that in serum for free beta hCG and total hCG but between 1.5 and 2 times wider for unconjugated oestriol and AFP. Analysis of data by fetal sex showed a significantly higher median MOM in female control cases compared with male controls for the analytes free beta hCG, total hCG, and unconjugated oestriol, but not for AFP. Amongst the Down's syndrome cases, this trend was not statistically significant and we cannot confirm a previous study which reported that elevated levels of amniotic fluid total and free beta hCG were associated only with female fetuses. PMID:9021826

  6. Alteration of the amniotic fluid and neonatal outcome.

    PubMed

    Volante, Enrico; Gramellini, Dandolo; Moretti, Sabrina; Kaihura, Christine; Bevilacqua, Giulio

    2004-01-01

    Quantitative and qualitative alterations of the amniotic fluid complicate 7% of the pregnancies. Polyhydramnios complicates 1-3% while oligohydramnios involves 3-5% of the pregnancies. The most common causes of polyhydramnios are fetal abnormalities, maternal diabetes and twin pregnancies, but are idiopathic in the 60%. Perinatal mortality has been reported to range between 10-30% while the risk of preterm birth reaches up to 22% in pregnancies complicated by polyhydramnios. The neonatal outcome, in cases where polyhydramnios is due to fetal-neonatal abnormalities, depends on the underlying pathology. Polyhydramnios due to defects in intestinal canalisation in particular, has been correlated to good neonatal prognosis. In our experience no early postoperative deaths occurred in a group of 16 newborns consequtively admitted to our unit in the last two years, with abnormalities of the gastrointestinal tract with need of surgery within the second week of life. Most cases of oligohydramnios are due to premature rupture of membranes, other causes are fetal abnormalities, such as urinary tract malformations, or chromosomopaties and drugs e.g. NSAID's. Oligohydramnios of mild entities is often associated to preterm birth, fetal growth restriction. In some cases of oligohydramnios, neonatal survival is highly conditioned by pulmonary hypoplasia which develops with rates that range between 13 and 21%. Neonatal prognosis is often disastrous in cases with severe oligohydramnios, which however could be improved by amnioinfusion, which restores an amniotic fluid volume sufficient in reducing the adverse environmental effects and in prolonging, where possible, pregnancy. Beside the quantity also the quality of the amniotic fluid may be related to the neonatal outcome. Finding of some inflammatory factors (interleukines) in the amniotic fluid seems to be significantly correlated to periventricular leucomalacia (PVL), cerebral paralysis and long-term neurological abnormalities

  7. Immunogenicity and safety of a monovalent, multicomponent acellular pertussis vaccine in 15 month-6-year-old German children. Monovalent Acellular Pertussis Vaccine Study Group.

    PubMed

    Stehr, K; Heininger, U; Uhlenbusch, R; Angersbach, P; Hackell, J; Eckhardt, T

    1995-03-01

    Immunization against pertussis has been re-recommended for healthy children in Germany in 1991. In addition the former restriction of immunizing only in the first 2 years of life was abolished. In children born before 1991 immunization rates against pertussis were 15% or less. With the new recommendations physicians are now faced with an increasing demand of parents for catch-up vaccinations in these children. Since they were immunized against diphtheria and tetanus previously monovalent pertussis vaccines are needed for this indication. Therefore a monovalent, multicomponent acellular pertussis vaccine was studied in 249 German children 15 months to 6 years of age. Three doses were administered at 6-10 week intervals. Reactogenicity and antibody responses against the vaccine antigens pertussis toxin (PT), filamentous haemagglutinin (FHA), 69-kd antigen (pertactin) and fimbriae-2 (agglutinogen) were investigated. Local and systemic reactions were minimal in frequency and severity. Antibody responses against all vaccine antigens were pronounced with 93%-100% of vaccinees demonstrating at least four fold titre rises above pre-immunization after the third dose. These findings indicate that this monovalent, multicomponent acellular pertussis vaccine with excellent immunogenicity and low reactogenicity is an appropriate candidate for closing immunization gaps in older children in countries with previously low vaccination rates against pertussis. Based on the results of this study the monovalent acellular pertussis vaccine was licensed in Germany in January 1994. PMID:7758519

  8. Beta Cells Secrete Significant and Regulated Levels of Insulin for Long Periods when Seeded onto Acellular Micro-Scaffolds.

    PubMed

    Sionov, Ronit Vogt; Finesilver, Gershon; Sapozhnikov, Lena; Soroker, Avigail; Zlotkin-Rivkin, Efrat; Saad, Yocheved; Kahana, Meygal; Bodaker, Matan; Alpert, Evgenia; Mitrani, Eduardo

    2015-11-01

    The aim of this work is to obtain significant and regulated insulin secretion from human beta cells ex vivo. Long-term culture of human pancreatic islets and attempts at expanding human islet cells normally result in loss of beta-cell phenotype. We propose that to obtain proper ex vivo beta cell function, there is a need to develop three-dimensional structures that mimic the natural islet tissue microenvironment. We here describe the preparation of endocrine micro-pancreata (EMPs) that are made up of acellular organ-derived micro-scaffolds seeded with human intact or enzymatically dissociated islets. We show that EMPs constructed by seeding whole islets, freshly enzymatically-dissociated islets or even dissociated islets grown first in standard monolayer cultures express high levels of key beta-cell specific genes and secrete quantities of insulin per cell similar to freshly isolated human islets in a glucose-regulated manner for more than 3 months in vitro. PMID:26416226

  9. Data on clinical significance of second trimester inflammatory biomarkers in the amniotic fluid in predicting preterm delivery.

    PubMed

    Kesrouani, Assaad; Chalhoub, Elie; El Rassy, Elie; Germanos, Mirna; Khazzaka, Aline; Rizkallah, Jamale; Attieh, Elie; Aouad, Norma

    2016-12-01

    In this article second trimester amniotic fluid biomarkers are measured for correlation with preterm delivery. One additional milliliter of amniotic fluid is collected during amniocentesis for dosages of IL-6, MMP-9, CRP and glucose levels, along with maternal serum CRP and glucose. MMP-9 and Il-6 levels were measured with the corresponding Human Quantikine(R) ELISA Kit (R&D systems) according to the instructions provided by the manufacturer. Cut-off values for AF MMP-9 and IL-6 were fixed by the kit sensitivity thresholds. Data includes ROC curves for glucose (Fig. 1), IL-6 (Fig. 2) and MMP-9 (Fig. 3), aiming to search for sensitivity and specificity in the prediction of premature delivery. Statistical analyses are performed with SPSS v20.0 software. Statistical significance is determined using the Mann-Whitney and one way ANOVA test. The association with preterm delivery is performed using a two proportions test. Correlations are measured using the Pearson׳'s coefficient. A p value<0.05 is considered statistically significant. The data is presented in the figures provided. Data relied on a previous publication "Prediction of preterm delivery by second trimester inflammatory biomarkers in the amniotic fluid" (A. Kesrouani, E. Chalhoub, E. El Rassy, M. Germanos, A. Khazzaka, J. Rizkallah, E. Attieh, N. Aouad, 2016) [1].

  10. Data on clinical significance of second trimester inflammatory biomarkers in the amniotic fluid in predicting preterm delivery.

    PubMed

    Kesrouani, Assaad; Chalhoub, Elie; El Rassy, Elie; Germanos, Mirna; Khazzaka, Aline; Rizkallah, Jamale; Attieh, Elie; Aouad, Norma

    2016-12-01

    In this article second trimester amniotic fluid biomarkers are measured for correlation with preterm delivery. One additional milliliter of amniotic fluid is collected during amniocentesis for dosages of IL-6, MMP-9, CRP and glucose levels, along with maternal serum CRP and glucose. MMP-9 and Il-6 levels were measured with the corresponding Human Quantikine(R) ELISA Kit (R&D systems) according to the instructions provided by the manufacturer. Cut-off values for AF MMP-9 and IL-6 were fixed by the kit sensitivity thresholds. Data includes ROC curves for glucose (Fig. 1), IL-6 (Fig. 2) and MMP-9 (Fig. 3), aiming to search for sensitivity and specificity in the prediction of premature delivery. Statistical analyses are performed with SPSS v20.0 software. Statistical significance is determined using the Mann-Whitney and one way ANOVA test. The association with preterm delivery is performed using a two proportions test. Correlations are measured using the Pearson׳'s coefficient. A p value<0.05 is considered statistically significant. The data is presented in the figures provided. Data relied on a previous publication "Prediction of preterm delivery by second trimester inflammatory biomarkers in the amniotic fluid" (A. Kesrouani, E. Chalhoub, E. El Rassy, M. Germanos, A. Khazzaka, J. Rizkallah, E. Attieh, N. Aouad, 2016) [1]. PMID:27626053

  11. Measurement of pesticides and other toxicants in amniotic fluid as a potential biomarker of prenatal exposure: a validation study.

    PubMed Central

    Bradman, Asa; Barr, Dana B; Claus Henn, Birgit G; Drumheller, Timothy; Curry, Cynthia; Eskenazi, Brenda

    2003-01-01

    Prenatal pesticide exposures may adversely affect children's health. However, exposure and health research is hampered by the lack of reliable fetal exposure data. No studies have been published that report measurements of commonly used nonpersistent pesticides in human amniotic fluid, although recent studies of pesticides in urine from pregnant women and in meconium indicate that fetuses are exposed to these chemicals. Amniotic fluid collected during amniocentesis is the only medium available to characterize direct fetal exposures early in pregnancy (approximately 18 weeks of gestation). As a first step in validating this exposure biomarker, we collected 100 amniotic fluid samples slated for disposal and evaluated analytical methods to measure organophosphate and carbamate pesticides and metabolites, synthetic pyrethroid metabolites, herbicides, and chlorinated phenolic compounds. The following six phenols were detected (detection frequency): 1- and 2-naphthol (70%), 2,5-dichlorophenol (55%), carbofuranphenol (5%), ortho-phenylphenol (30%), and pentachlorophenol (15%), with geometric mean concentrations of 0.72, 0.39, 0.12, 0.13, and 0.23 microg/L, respectively, for positive values. The organophosphate metabolites diethylphosphate and dimethylphosphate were detected in two (10%) samples, and dimethylthiophosphate was detected in one (5%) sample, with geometric mean concentrations of 0.31, 0.32, and 0.43 microg/L, respectively, for positive values. These levels are low compared with levels reported in urine, blood, and meconium in other studies, but indicate direct exposures to the young fetus, possibly during critical periods of development. Results of this pilot study suggest that amniotic fluid offers a unique opportunity to investigate fetal exposures and health risks. PMID:14594631

  12. A novel approach for noninvasive drug delivery and sensing through the amniotic sac.

    PubMed

    Azagury, Aharon; Amar-Lewis, Eliz; Mann, Ella; Goldbart, Riki; Traitel, Tamar; Jelinek, Raz; Hallak, Mordechai; Kost, Joseph

    2014-06-10

    Current invasive prenatal tests (amniocentesis and chorionic villus sampling) are known for their risk to the fetus. In the last decade, the use and awareness of these prenatal tests have increased, resulting in growing demand for a safe, non-invasive, and accurate prenatal test. Chemical penetration enhancers (CPEs) have long been used to increase transport phenomena across skin and other membranes (e.g., tympanic membrane). The amniotic sac membrane is called the chorioamnion (CA) membrane and serves as the physical barrier between the fetus and the mother. In this research, the effect of CPEs on human CA mass transport was evaluated both in vitro and ex vivo. The results show that the tested CPEs exhibit an enhancing effect on CA mass transport. Based on the permeability results, two mechanisms of action were suggested: "extractors" and "fluidizers". Fourier transform infrared (FTIR) and rapid colorimetric screening measurements supported the mechanisms, based on which, more potent compounds were designed and tested for their enhancing effect. The enhancing mass transport effect of CPEs on CA membrane may be used both for sampling of cell-free DNA and for noninvasively administering drugs and other biological agents to the amniotic sac. PMID:24685707

  13. A novel approach for noninvasive drug delivery and sensing through the amniotic sac.

    PubMed

    Azagury, Aharon; Amar-Lewis, Eliz; Mann, Ella; Goldbart, Riki; Traitel, Tamar; Jelinek, Raz; Hallak, Mordechai; Kost, Joseph

    2014-06-10

    Current invasive prenatal tests (amniocentesis and chorionic villus sampling) are known for their risk to the fetus. In the last decade, the use and awareness of these prenatal tests have increased, resulting in growing demand for a safe, non-invasive, and accurate prenatal test. Chemical penetration enhancers (CPEs) have long been used to increase transport phenomena across skin and other membranes (e.g., tympanic membrane). The amniotic sac membrane is called the chorioamnion (CA) membrane and serves as the physical barrier between the fetus and the mother. In this research, the effect of CPEs on human CA mass transport was evaluated both in vitro and ex vivo. The results show that the tested CPEs exhibit an enhancing effect on CA mass transport. Based on the permeability results, two mechanisms of action were suggested: "extractors" and "fluidizers". Fourier transform infrared (FTIR) and rapid colorimetric screening measurements supported the mechanisms, based on which, more potent compounds were designed and tested for their enhancing effect. The enhancing mass transport effect of CPEs on CA membrane may be used both for sampling of cell-free DNA and for noninvasively administering drugs and other biological agents to the amniotic sac.

  14. Isolation and characterization of equine amniotic membrane-derived mesenchymal stem cells.

    PubMed

    Seo, Min-Soo; Park, Sang-Bum; Kim, Hyung-Sik; Kang, Jun-gu; Chae, Joon-Seok; Kang, Kyung-Sun

    2013-01-01

    Recent studies have shown that mesenchymal stem cells (MSCs) are able to differentiate into multi-lineage cells such as adipocytes, chondroblasts, and osteoblasts. Amniotic membrane from whole placenta is a good source of stem cells in humans. This membrane can potentially be used for wound healing and corneal surface reconstruction. Moreover, it can be easily obtained after delivery and is usually discarded as classified waste. In the present study, we successfully isolated and characterized equine amniotic membrane-derived mesenchymal stem cells (eAM-MSCs) that were cultured and maintained in low glucose Dulbecco's modified Eagle's medium. The proliferation of eAM-MSCs was measured based on the cumulative population doubling level (CPDL). Immunophenotyping of eAM-MSCs by flow cytometry showed that the major population was of mesenchymal origin. To confirm differentiation potential, a multi-lineage differentiation assay was conducted. We found that under appropriate conditions, eAM-MSCs are capable of multi-lineage differentiation. Our results indicated that eAM-MSCs may be a good source of stem cells, making them potentially useful for veterinary regenerative medicine and cell-based therapy.

  15. Midtrimester intra-amniotic sludge and the risk of spontaneous preterm birth.

    PubMed

    Himaya, Eric; Rhalmi, Nadia; Girard, Mario; Tétu, Amélie; Desgagné, Josée; Abdous, Belkacem; Gekas, Jean; Giguère, Yves; Bujold, Emmanuel

    2011-12-01

    We examined the association between midtrimester intra-amniotic sludge and spontaneous preterm birth (PTB) in asymptomatic women undergoing amniocentesis. We performed a prospective cohort study of women having an amniocentesis for fetal karyotyping between 14 and 24 weeks' gestation. Cervical length and the presence of amniotic sludge were assessed by transvaginal ultrasound. Amniotic fluid concentrations of matrix metalloproteinase-8, glucose and lactate were measured. Early (<32 weeks) and late (32 to 36 weeks) preterm premature rupture of membranes (PPROM) and spontaneous PTB constituted primary outcomes. Nonparametric analyses were conducted. Three hundred ten women, including 94 (30%) with free-floating echogenic particles and 16 (5%) with dense amniotic sludge, were recruited. Dense amniotic sludge was linked with early (13%) but not with late (0%) primary outcome ( P < 0.01). Two women with combined dense amniotic sludge and short cervix delivered 4 and 10 weeks later (at 20 and 25 weeks, respectively) and had a higher median amniotic lactate concentration than controls ( P < 0.05). A third woman with dense amniotic sludge at 15 weeks was diagnosed with a short cervix and an intra-amniotic infection at 22 weeks that was eradicated with intravenous antibiotics. Midtrimester dense amniotic sludge is associated with early PPROM and spontaneous PTB.

  16. Repair of a Gingival Fenestration Using an Acellular Dermal Matrix Allograft.

    PubMed

    Breault, Lawrence G; Brentson, Raquel C; Fowler, Edward B; Bisch, Frederick C

    2016-01-01

    A case report illustrating the successful treatment of a gingival fenestration with an acellular dermal matrix (ADM) allograft. After 2½ months of healing, the ADM was completely integrated into the soft tissues of the mandibular anterior gingiva with complete resolution of the gingival fenestration, resulting in excellent gingival esthetics. PMID:26874103

  17. Amniotic mesenchymal stem cells display neurovascular tropism and aid in the recovery of injured peripheral nerves.

    PubMed

    Li, YongNan; Guo, Longzhe; Ahn, Hyun Sook; Kim, Moo Hyun; Kim, Sung-Whan

    2014-06-01

    Recently, we reported that human amniotic membrane-derived mesenchymal stem cells (AMMs) possess great angiogenic potential. In this study, we determined whether local injection of AMMs ameliorates peripheral neuropathy. AMMs were transplanted into injured sciatic nerves. AMM injection promoted significant recovery of motor nerve conduction velocity and voltage amplitude compared to human adipose-derived mesenchymal stem cells. AMM implantation also augmented blood perfusion and increased intraneural vascularity. Whole-mount fluorescent imaging analysis demonstrated that AMMs exhibited higher engraftment and endothelial incorporation abilities in the sciatic nerve. In addition, the higher expression of pro-angiogenic factors was detected in AMMs injected into the peripheral nerve. Therefore, these data provide novel therapeutic and mechanistic insights into stem cell biology, and AMM transplantation may represent an alternative therapeutic option for treating peripheral neuropathy.

  18. Midkine and Pleiotrophin Concentrations in Amniotic Fluid in Healthy and Complicated Pregnancies

    PubMed Central

    Chaemsaithong, Piya; Yan, Gai; Peran, Ivana; Wellstein, Anton; Romero, Roberto; Baron, Jeffrey

    2016-01-01

    Background Midkine (MDK) and pleiotrophin (PTN) are heparin-binding growth factors that, in rodents, are highly expressed in early life and decrease to undetectable levels by adulthood. The potential roles of MDK and PTN in human growth and development are not completely elucidated. Method and Findings To delineate the role of MDK and PTN in human development, we developed high sensitivity assays to measure their concentrations in amniotic fluid (AF) at various gestational ages in both healthy and complicated pregnancies. We found that both of these growth factors could be readily measured in AF and that the concentrations were higher than most cytokines previously reported in AF. Conclusion The concentration of MDK but not that of PTN declined with gestational age. Both MDK and PTN concentrations were found to be lower in pregnancies that were complicated by chorioamnionitis at term, raising the possibility that these growth factors might be useful as markers for infection. PMID:27089523

  19. Development and characterization of acellular porcine pulmonary valve scaffolds for tissue engineering.

    PubMed

    Luo, Ji; Korossis, Sotirios A; Wilshaw, Stacy-Paul; Jennings, Louise M; Fisher, John; Ingham, Eileen

    2014-11-01

    Currently available replacement heart valves all have limitations. This study aimed to produce and characterize an acellular, biocompatible porcine pulmonary root conduit for reconstruction of the right ventricular outflow tract e.g., during Ross procedure. A process for the decellularization of porcine pulmonary roots was developed incorporating trypsin treatment of the adventitial surface of the scraped pulmonary artery and sequential treatment with hypotonic Tris buffer (HTB; 10 mM Tris pH 8.0, 0.1% (w/v) EDTA, and 10 KIU aprotinin), 0.1% (w/v) sodium dodecyl sulfate in HTB, two cycles of DNase and RNase, and sterilization with 0.1% (v/v) peracetic acid. Histology confirmed an absence of cells and retention of the gross histoarchitecture. Immunohistochemistry further confirmed cell removal and partial retention of the extracellular matrix, but a loss of collagen type IV. DNA levels were reduced by more than 96% throughout all regions of the acellular tissue and no functional genes were detected using polymerase chain reaction. Total collagen levels were retained but there was a significant loss of glycosaminoglycans following decellularization. The biomechanical, hydrodynamic, and leaflet kinematics properties were minimally affected by the process. Both immunohistochemical labeling and antibody absorption assay confirmed a lack of α-gal epitopes in the acellular porcine pulmonary roots and in vitro biocompatibility studies indicated that acellular leaflets and pulmonary arteries were not cytotoxic. Overall the acellular porcine pulmonary roots have excellent potential for development of a tissue substitute for right ventricular outflow tract reconstruction e.g., during the Ross procedure. PMID:24786313

  20. Amniotic membrane properties and current practice of amniotic membrane use in ophthalmology in Slovenia.

    PubMed

    Cirman, Tina; Beltram, Matej; Schollmayer, Petra; Rožman, Primož; Kreft, Mateja Erdani

    2014-06-01

    Amniotic membrane (AM) is the innermost, multilayered part of the placenta. When harvested, processed and stored properly, its properties, stemming from AM biological composition, make it a useful tissue for ophthalmic surgery. AM was shown to have several beneficial effects: it promotes epithelization, has antimicrobial effects, decreases inflammation, fibrosis and neovascularization. Many case reports and case series as well as practical experience (e.g. reconstruction of conjunctival and corneal defects, treatment of corneal ulcers) demonstrated the beneficial effect of AM for different ophthalmological indications. The combination of the above mentioned beneficial effects and reasonable mechanical properties are also the reason why AM is used as a substrate for ex vivo expansion of epithelial progenitor cells. Recently, amnion-derived cells, which also have stem cell characteristics, have been proposed as potential contributors to cell-based treatment of ocular surface disease. However, the use of AM remains one of the least standardized methods in ophthalmic surgery. In this review, the various properties of AM and its current clinical use in ophthalmology in Slovenia are discussed.

  1. Prognostic potential of amniotic fluid analysis at birth on canine neonatal outcomes.

    PubMed

    Groppetti, D; Martino, P A; Ravasio, G; Bronzo, V; Pecile, A

    2015-12-01

    Glucose, lactate and cortisol concentrations in amniotic fluid were measured at birth in 95 pups and related to neonatal viability based on Apgar scoring and to neonatal mortality. Neither amniotic parameters nor neonatal mortality were associated with the Apgar score. Stillborn pups showed high lactate (P < 0.001) and cortisol (P < 0.05) but low glucose amniotic concentrations (P < 0.001). No amniotic fluid differences were observed between normal and malformed pups. Amniotic glucose (P < 0.001), lactate (P < 0.05) and cortisol (P < 0.05) concentrations were higher in pups delivered by vaginal parturition than by Caesarean section. Birth weight was higher in live pups than in pups dying within 48 h (P < 0.05). Although these are preliminary results, the analysis of amniotic fluid collected at birth could be a valuable predictor of neonatal outcomes in dogs.

  2. Amniotic fluid-borne hepatocyte growth factor protects rat pups against experimental necrotizing enterocolitis.

    PubMed

    Jain, Sunil K; Baggerman, Eric W; Mohankumar, Krishnan; Namachivayam, Kopperuncholan; Jagadeeswaran, Ramasamy; Reyes, Victor E; Maheshwari, Akhil

    2014-03-01

    Fetal swallowing of amniotic fluid, which contains numerous cytokines and growth factors, plays a key role in gut mucosal development. Preterm birth interrupts this exposure to amniotic fluid-borne growth factors, possibly contributing to the increased risk of necrotizing enterocolitis (NEC) in premature infants. We hypothesized that supplementation of formula feeds with amniotic fluid can provide amniotic fluid-borne growth factors and prevent experimental NEC in rat pups. We compared NEC-like injury in rat pups fed with infant formula vs. formula supplemented either with 30% amniotic fluid or recombinant hepatocyte growth factor (HGF). Cytokines/growth factors in amniotic fluid were measured by immunoassays. Amniotic fluid and HGF effects on enterocyte migration, proliferation, and survival were measured in cultured IEC6 intestinal epithelial cells. Finally, we used an antibody array to investigate receptor tyrosine kinase (RTK) activation and immunoblots to measure phosphoinositide 3-kinase (PI3K) signaling. Amniotic fluid supplementation in oral feeds protected rat pups against NEC-like injury. HGF was the most abundant growth factor in rat amniotic fluid in our panel of analytes. Amniotic fluid increased cell migration, proliferation, and cell survival in vitro. These effects were reproduced by HGF and blocked by anti-HGF antibody or a PI3K inhibitor. HGF transactivated several RTKs in IEC6 cells, indicating that its effects extended to multiple signaling pathways. Finally, similar to amniotic fluid, recombinant HGF also reduced the frequency and severity of NEC-like injury in rat pups. Amniotic fluid supplementation protects rat pups against experimental NEC, which is mediated, at least in part, by HGF.

  3. Amniotic fluid-borne hepatocyte growth factor protects rat pups against experimental necrotizing enterocolitis

    PubMed Central

    Baggerman, Eric W.; MohanKumar, Krishnan; Namachivayam, Kopperuncholan; Jagadeeswaran, Ramasamy; Reyes, Victor E.; Maheshwari, Akhil

    2014-01-01

    Fetal swallowing of amniotic fluid, which contains numerous cytokines and growth factors, plays a key role in gut mucosal development. Preterm birth interrupts this exposure to amniotic fluid-borne growth factors, possibly contributing to the increased risk of necrotizing enterocolitis (NEC) in premature infants. We hypothesized that supplementation of formula feeds with amniotic fluid can provide amniotic fluid-borne growth factors and prevent experimental NEC in rat pups. We compared NEC-like injury in rat pups fed with infant formula vs. formula supplemented either with 30% amniotic fluid or recombinant hepatocyte growth factor (HGF). Cytokines/growth factors in amniotic fluid were measured by immunoassays. Amniotic fluid and HGF effects on enterocyte migration, proliferation, and survival were measured in cultured IEC6 intestinal epithelial cells. Finally, we used an antibody array to investigate receptor tyrosine kinase (RTK) activation and immunoblots to measure phosphoinositide 3-kinase (PI3K) signaling. Amniotic fluid supplementation in oral feeds protected rat pups against NEC-like injury. HGF was the most abundant growth factor in rat amniotic fluid in our panel of analytes. Amniotic fluid increased cell migration, proliferation, and cell survival in vitro. These effects were reproduced by HGF and blocked by anti-HGF antibody or a PI3K inhibitor. HGF transactivated several RTKs in IEC6 cells, indicating that its effects extended to multiple signaling pathways. Finally, similar to amniotic fluid, recombinant HGF also reduced the frequency and severity of NEC-like injury in rat pups. Amniotic fluid supplementation protects rat pups against experimental NEC, which is mediated, at least in part, by HGF. PMID:24407592

  4. Are some chromosomes particularly good at sex? Insights from amniotes.

    PubMed

    O'Meally, Denis; Ezaz, Tariq; Georges, Arthur; Sarre, Stephen D; Graves, Jennifer A Marshall

    2012-01-01

    Several recent studies have produced comparative maps of genes on amniote sex chromosomes, revealing homology of gene content and arrangement across lineages as divergent as mammals and lizards. For example, the chicken Z chromosome, which shares homology with the sex chromosomes of all birds, monotremes, and a gecko, is a striking example of stability of genome organization and retention, or independent acquisition, of function in sex determination. In other lineages, such as snakes and therian mammals, well conserved but independently evolved sex chromosome systems have arisen. Among lizards, novel sex chromosomes appear frequently, even in congeneric species. Here, we review recent gene mapping data, examine the evolutionary relationships of amniote sex chromosomes and argue that gene content can predispose some chromosomes to a specialized role in sex determination.

  5. An enzymic radiochemical method for determining phosphatidylglycerol in amniotic fluid

    SciTech Connect

    Siegel, L.; Walker, S.I.; Robin, N.I.

    1983-05-01

    We describe an enzymic quantification of phosphatidylglycerol in amniotic fluid. Phosphatidylglycerol is hydrolyzed in alkali and the resulting glycerol is then enzymatically phosphorylated with adenosine 5'-(gamma-/sup 32/P)triphosphate to yield glycero(/sup 32/P)phosphate. After removal of excess (gamma-/sup 32/P)ATP by charcoal, the radioactivity of the glycerophosphate is measured in a liquid scintillation counter. Triglyceride in the amniotic fluid is hydrolyzed by lipase before extraction and thus does not interfere with the analysis. This method is specific for phosphatidylglycerol. Preliminary studies suggest that a phosphatidylglycerol value greater than or equal to 10 nmol/mL correlates with fetal lung maturity.

  6. Use of an acellular flowable dermal replacement scaffold on lower extremity sinus tract wounds: a retrospective series.

    PubMed

    Brigido, Stephen A; Schwartz, Edward; McCarroll, Raymond; Hardin-Young, Janet

    2009-04-01

    A novel injectable human dermal matrix has been developed for the treatment of complex diabetic sinus tract wounds. Bioengineered grafts are commercially available that have been somewhat effective in treating chronic wounds such as diabetic foot ulcers; however, these bioengineered grafts are only available in sheet form. These therapies are less effective in treating complex or irregularly shaped wounds that demonstrate tunnels or extensions into deep soft tissue. One acellular graft (GRAFTJACKET, Matrix, Wright Medical Technology, Arlington, Tennessee) that has been shown to effectively treat open wounds is also available in a micronized form (GRAFTJACKET Xpress Scaffold, Wright Medical Technology). This human dermal graft forms a flowable soft tissue scaffold that can be delivered via syringe into tunneling wounds. In this retrospective series, 12 patients with deep tunneling wounds were treated with GRAFTJACKET Xpress Scaffold and followed for 12 weeks. Complete wound healing was achieved in 10 of 12 patients within the 12-week evaluation. The average time to complete healing was 8.5 weeks, whereas the average time to depth healing was 7.8 weeks. The data from the study suggest that this injectable human dermal matrix has unique properties that allow it to facilitate healing of complex tunneling diabetic foot ulcers. The material is easy to prepare and inject into the wound, thereby preventing the necessity of extensive surgical exposure. The matrix supports neo-subcutaneous tissue formation and allows the body to rapidly repair these wounds.

  7. 21 CFR 862.1455 - Lecithin/sphingomyelin ratio in amniotic fluid test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... lecithin/sphingomyelin ratio in amniotic fluid. Lecithin and sphingomyelin are phospholipids (fats or fat-like substances containing phosphorus). Measurements of the lecithin/sphingomyelin ratio in...

  8. 21 CFR 862.1455 - Lecithin/sphingomyelin ratio in amniotic fluid test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... lecithin/sphingomyelin ratio in amniotic fluid. Lecithin and sphingomyelin are phospholipids (fats or fat-like substances containing phosphorus). Measurements of the lecithin/sphingomyelin ratio in...

  9. 21 CFR 862.1455 - Lecithin/sphingomyelin ratio in amniotic fluid test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... lecithin/sphingomyelin ratio in amniotic fluid. Lecithin and sphingomyelin are phospholipids (fats or fat-like substances containing phosphorus). Measurements of the lecithin/sphingomyelin ratio in...

  10. The Immunosuppressive Activity of Amniotic Membrane Mesenchymal Stem Cells on T Lymphocytes

    PubMed Central

    Alikarami, Fatemeh; Yari, Fatemeh; Amirizadeh, Naser; Nikougoftar, Mahin; Jalili, Mohammad Ali

    2015-01-01

    Background: Mesenchymal Stem Cells (MSCs) are isolated from different sources like placenta. The placenta and its membranes like Amniotic Membrane (AM) are readily available and easy to work with. There is only limited knowledge on the immunomodulatory properties of human Amniotic Membrane-derived Mesenchymal Stem Cells (hAM-MSCs). The aim of this study was to survey the suppressive activity of hAM-MSCs on T lymphocytes in vitro. Methods: Human AMs were obtained after caesarean section births from healthy women. After enzymatic digestion, cells were cultured and hAM-MSCs were obtained. In addition, human T lymphocytes were isolated and co-cultured with hAM-MSCs for 72 hr in the presence or absence of phytohemagglutinin (PHA). Subsequently, proliferation of T cells was analyzed using BrdU and subsequently flow cytometry technique. Besides, the production of IL-4 and IFN-γ was examined by ELISA method. Additionally, the expression of activation markers (CD38, HLA-DR) was studied on T lymphocytes by flow cytometry technique. Results: It was revealed that hAM-MSCs could significantly suppress the proliferation of T lymphocytes (p≤0.01) and significantly decrease the production of IFN-γ by T cells (p<0.05). hAM-MSCs also down regulated the expression of activation markers on the surface of T lymphocytes, CD38 and HLA-DR. The difference was significant between the case and control samples (p<0.05). All the comparisons were carried out between the case (Tcell+PHA+hAM-MSCs) and control (Tcell+PHA) groups. Conclusion: In conclusion, hAM-MSCs could inhibit the (mitogen-activated) T cells even in the absence of blood monocytes. Besides, hAM-MSCs-mediated inhibition of T lymphocytes was combined with down regulation of activation markers. PMID:26306147

  11. Immunomodulatory effects of amniotic membrane matrix incorporated into collagen scaffolds.

    PubMed

    Hortensius, Rebecca A; Ebens, Jill H; Harley, Brendan A C

    2016-06-01

    Adult tendon wound repair is characterized by the formation of disorganized collagen matrix which leads to decreases in mechanical properties and scar formation. Studies have linked this scar formation to the inflammatory phase of wound healing. Instructive biomaterials designed for tendon regeneration are often designed to provide both structural and cellular support. In order to facilitate regeneration, success may be found by tempering the body's inflammatory response. This work combines collagen-glycosaminoglycan scaffolds, previously developed for tissue regeneration, with matrix materials (hyaluronic acid and amniotic membrane) that have been shown to promote healing and decreased scar formation in skin studies. The results presented show that scaffolds containing amniotic membrane matrix have significantly increased mechanical properties and that tendon cells within these scaffolds have increased metabolic activity even when the media is supplemented with the pro-inflammatory cytokine interleukin-1 beta. Collagen scaffolds containing hyaluronic acid or amniotic membrane also temper the expression of genes associated with the inflammatory response in normal tendon healing (TNF-α, COLI, MMP-3). These results suggest that alterations to scaffold composition, to include matrix known to decrease scar formation in vivo, can modify the inflammatory response in tenocytes. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1332-1342, 2016.

  12. Therapeutic outcomes of transplantation of amniotic fluid-derived stem cells in experimental ischemic stroke

    PubMed Central

    Tajiri, Naoki; Acosta, Sandra; Portillo-Gonzales, Gabriel S.; Aguirre, Daniela; Reyes, Stephanny; Lozano, Diego; Pabon, Mibel; Dela Peña, Ike; Ji, Xunming; Yasuhara, Takao; Date, Isao; Solomita, Marianna A.; Antonucci, Ivana; Stuppia, Liborio; Kaneko, Yuji; Borlongan, Cesar V.

    2014-01-01

    Accumulating preclinical evidence suggests the use of amnion as a source of stem cells for investigations of basic science concepts related to developmental cell biology, but also for stem cells’ therapeutic applications in treating human disorders. We previously reported isolation of viable rat amniotic fluid-derived stem (AFS) cells. Subsequently, we recently reported the therapeutic benefits of intravenous transplantation of AFS cells in a rodent model of ischemic stroke. Parallel lines of investigations have provided safety and efficacy of stem cell therapy for treating stroke and other neurological disorders. This review article highlights the need for investigations of mechanisms underlying AFS cells’ therapeutic benefits and discusses lab-to-clinic translational gating items in an effort to optimize the clinical application of the cell transplantation for stroke. PMID:25165432

  13. Upregulation of Nanog and Sox-2 genes following ectopic expression of Oct-4 in amniotic fluid mesenchymal stem cells.

    PubMed

    Wang, Kai-Hung; Kao, An-Pei; Chang, Chia-Cheng; Lin, Ta-Chin; Kuo, Tsung-Cheng

    2015-01-01

    Octamer-binding transcription factor 4 (Oct-4), an important gene regulating stem cell pluripotency, is well-known for its ability to reprogram somatic cells in vitro, either alone or in concert with other factors. The aim of this study was to assess the effect of ectopic expression of Oct human amniotic fluid stem cells. We developed a novel method for isolation of putative human amniotic fluid-derived multipotent stem cells. These cells showing mesenchymal stem cell phenotypes (human amniotic fluid-derived mesenchymal stem cells, hAFMSCs) were transfected with a plasmid carrying genes for Oct-4 and the green fluorescent protein (GFP). The stably transfected cells, hAFMSCs-Oct4/GFP, were selected by using G418 and found to express the GFP reporter gene under the control of Oct-4 promoter. We found that hAFMSCs developed by our method possess very high self-renewal ability (about 78 cumulative population doublings) and multilineage differentiation potency. Significantly, the hAFMSCs-Oct4/GFP cells showed enhanced expression of the three major pluripotency genes Oct-4, Nanog, and Sox-2, and increased colony-forming ability and growth rate compared with the parental hAFMSCs. We demonstrated that the ectopic expression of Oct-4 gene in hAFMSCs with high self-renewal ability could upregulate Nanog and Sox-2 gene expression and enhance cell growth rate and colony-forming efficiency. Therefore, the ectopic expression of Oct-4 could be a strategy to develop pluripotency in hAFMSCs for clinical applications.

  14. Amniotic membrane welded to contact lens by 1470-nm diode laser: a novel method for sutureless amniotic membrane transplantation

    PubMed Central

    Rasier, Rifat; Gulsoy, Murat

    2014-01-01

    AIM To avoid the side effects of the suture usage by welding amniotic membrane (AM) to contact lens (CL) with laser. METHODS AM was taken from pregnant women and cleaned from blood clots with sterile phosphate-buffered physiological saline solution which included antibiotics. Stromal side of the AM was spread inside of the CL and it was welded to CL by 1470 nm diode laser. 600 µm diameter fiber tip of the laser was contacted with the epithelial side of the AM from 4 separate points. After welding excess amniotic membrane around the CL was cut with a scalpel. RESULTS Stromal side of the AM was spread inside of the CL and then with laser fiber, different power levels and exposure times were applied on the epithelium of AM and 340 mW for seven seconds was found optimal. CL and AM attached with the spot welding effect in 4 points by touching fiber tip. CL-AM welded complex did not separated from each other while holding AM that extend beyond the CL with the help of two forceps. CONCLUSION As a conclusion, it was aimed in this study to achieve the success of the conventional amniotic membrane transplantation (AMT)with the easiness of applying a CL and to avoid risks and side effects of corneal or conjunctival suturing. The results showed that the application of the CL–AM complex will be as easy as the application of a CL and lasts shortly. PMID:25540753

  15. Cell-free fetal DNA in amniotic fluid supernatant for prenatal diagnosis.

    PubMed

    Soltani, M; Nemati, M; Maralani, M; Estiar, M A; Andalib, S; Fardiazar, Z; Sakhinia, E

    2016-01-01

    In widespread conviction, amniotic fluid is utilized for prenatal diagnosis. Amniotic fluid supernatant is usually discarded, notwithstanding being a good source of fetal DNA. The aim of the present study was to assess cell-free fetal DNA extracted from amniotic fluid supernatant for application in prenatal diagnosis such as gender determination and early diagnosis of β-thalassemia. Samples of amniotic fluid of 70 pregnant women were collected and went through routine tests along with tests for cell-free fetal DNA from amniotic fluid supernatant. The DNA in the amniotic fluid supernatant was extracted and analyzed for gender determination by PCR and Real-time PCR. ARMS-PCR was applied to test early diagnosis of IVS II-I mutation (common β-thalassemia mutation) and E7V mutation for sickle cell anemia using DNA extracted from the amniotic fluid supernatant. Using the cell-free fetal DNA extracted from the amniotic fluid supernatant, the sensitivity of PCR and Real-time PCR for gender detection was compared with the routine cytogenetic method. The fetus tested for sickle cell anemia and β-thalassemia was observed to be healthy but heterozygous for IVS II-I mutation. The findings indicated that cell-free fetal DNA from amniotic fluid supernatant can be a good source of fetal DNA and be used in early prenatal diagnosis since because of its fast and accurate application. Therefore, it would be suggested that the amniotic fluid supernatant's disposal is prevented because if the tests needs to be repeated, cell-free fetal DNA extracted from the amniotic fluid supernatant can be used as an alternative source for prenatal diagnosis. PMID:27188728

  16. Cell-free fetal DNA in amniotic fluid supernatant for prenatal diagnosis.

    PubMed

    Soltani, M; Nemati, M; Maralani, M; Estiar, M A; Andalib, S; Fardiazar, Z; Sakhinia, E

    2016-04-30

    In widespread conviction, amniotic fluid is utilized for prenatal diagnosis. Amniotic fluid supernatant is usually discarded, notwithstanding being a good source of fetal DNA. The aim of the present study was to assess cell-free fetal DNA extracted from amniotic fluid supernatant for application in prenatal diagnosis such as gender determination and early diagnosis of β-thalassemia. Samples of amniotic fluid of 70 pregnant women were collected and went through routine tests along with tests for cell-free fetal DNA from amniotic fluid supernatant. The DNA in the amniotic fluid supernatant was extracted and analyzed for gender determination by PCR and Real-time PCR. ARMS-PCR was applied to test early diagnosis of IVS II-I mutation (common β-thalassemia mutation) and E7V mutation for sickle cell anemia using DNA extracted from the amniotic fluid supernatant. Using the cell-free fetal DNA extracted from the amniotic fluid supernatant, the sensitivity of PCR and Real-time PCR for gender detection was compared with the routine cytogenetic method. The fetus tested for sickle cell anemia and β-thalassemia was observed to be healthy but heterozygous for IVS II-I mutation. The findings indicated that cell-free fetal DNA from amniotic fluid supernatant can be a good source of fetal DNA and be used in early prenatal diagnosis since because of its fast and accurate application. Therefore, it would be suggested that the amniotic fluid supernatant's disposal is prevented because if the tests needs to be repeated, cell-free fetal DNA extracted from the amniotic fluid supernatant can be used as an alternative source for prenatal diagnosis.

  17. Prosthetic Breast Reconstruction With Acellular Dermal Matrices: Achieving Predictability and Reproducibility.

    PubMed

    Nahabedian, Maurice Y

    2016-05-01

    The use of acellular dermal matrices in the setting of prosthetic breast reconstruction has captured the attention of many plastic surgeons. The regenerative capacity of these materials has provided additional tissue support to the mastectomy skin flaps with the ultimate result of improving surgical and aesthetic outcomes. Despite the benefits, there remains a significant diversity with regard to outcomes with some surgeons reporting increased morbidity. The reasons for this are varied but ultimately related to differences in patient selection and surgical techniques. The purpose of this article is to provide strategies for using acellular dermal matrix to achieve success in a manner that is usually associated with outcomes that are predictable and reproducible. PMID:27579223

  18. Pioneering technique using Acellular Dermal Matrix in the rescue of a radiation ulcer

    PubMed Central

    NASEEM, S.; PATEL, A.D.; DEVALIA, H.

    2016-01-01

    Background Radiotherapy as an adjuvant to mastectomy is integral to the treatment of breast cancer, but can result in skin ulceration. Skin ulceration following radiotherapy is traditionally managed by removing the implant and allowing the skin to heal by secondary intention. Case report A 42-year-old woman underwent radiotherapy following a breast reconstruction. She developed a 2 x 3cm radiation ulcer. The ulcer was managed by removing the implant and performing capsulectomy. A Beckers 50 expander was placed and reinforced with acellular dermal matrix inferolaterally. At follow-up the patient had a good cosmetic outcome. Conclusion Post-radiation skin ulcers present a challenge to treat with no current standardised management. The use of acellular dermal matrix may present a new technique to promote healing in these testing cases. PMID:27142826

  19. Prosthetic Breast Reconstruction With Acellular Dermal Matrices: Achieving Predictability and Reproducibility

    PubMed Central

    2016-01-01

    Summary: The use of acellular dermal matrices in the setting of prosthetic breast reconstruction has captured the attention of many plastic surgeons. The regenerative capacity of these materials has provided additional tissue support to the mastectomy skin flaps with the ultimate result of improving surgical and aesthetic outcomes. Despite the benefits, there remains a significant diversity with regard to outcomes with some surgeons reporting increased morbidity. The reasons for this are varied but ultimately related to differences in patient selection and surgical techniques. The purpose of this article is to provide strategies for using acellular dermal matrix to achieve success in a manner that is usually associated with outcomes that are predictable and reproducible. PMID:27579223

  20. The Cellular Immune Mechanism after Transfer of Chemically Extracted Acellular Nerve Xenografts

    PubMed Central

    Lin, Xingshi; Yang, Ruojia; He, Qing; Ruan, Dike

    2013-01-01

    Severe peripheral nerve defect by injuries causing functional loss require nerve grafting. Autograft has limitations for clinical use because it results in the creation of a new nerve injury and the generation of donor site morbidity. Based on these limitations, nerve allografts and xenografts provide a readily accessible alternative strategy. The aim of the present study was to observe the immune mechanism underlying the rejection of chemically extracted acellular nerve xenografts, and further evaluate immunogenicity of chemically treated acellular nerve grafts for clinical applications. A total of 160 BALB/c mice were randomly divided into a negative contrast group (NC, 40 mice), a fresh autograft group (AG, 40 mice), a fresh xenogeneic nerve group (FXN, 40 mice) and a chemically extracted acellular xenogeneic nerve group (CEXN, 40 mice). Various types of nerve grafts were implanted into the thigh muscle of BALB/C mice in the corresponding groups. At 3, 7, 14 and 28 days post-operation, the mice (10 mice from each group) were sacrificed and their spleens were extracted. The spleens were ground into paste. The erythrocytes and other cells were lysed using distilled water and the T lymphocytes were collected. Fluorescein isothiocyanate (FITC) -labeled monoclonal antibodies (CD3, CD4, CD8, CD25, IL-2, IFN-γ and TNF-α) were then added to the solution. The Fluorescence Activated Cell Sorting (FACS) was used to determine the positivity rate of the cells combined with the monoclonal antibodies above. No significant statistical differences were observed between the CEXN, NC and AG groups, so that no obvious immune rejections were observed among the chemically extracted acellular nerve xenografts. PMID:23874771

  1. Hertwig's epithelial root sheath cell behavior during initial acellular cementogenesis in rat molars.

    PubMed

    Yamamoto, Tsuneyuki; Yamamoto, Tomomaya; Yamada, Tamaki; Hasegawa, Tomoka; Hongo, Hiromi; Oda, Kimimitsu; Amizuka, Norio

    2014-11-01

    This study was designed to examine developing acellular cementum in rat molars by immunohistochemistry, to elucidate (1) how Hertwig's epithelial root sheath disintegrates and (2) whether epithelial sheath cells transform into cementoblasts through epithelial-mesenchymal transition (EMT). Initial acellular cementogenesis was divided into three developmental stages, which can be seen in three different portions of the root: portion 1, where the epithelial sheath is intact; portion 2, where the epithelial sheath becomes fragmented; and portion 3, where acellular cementogenesis begins. Antibodies against three kinds of matrix proteinases, which degrade epithelial sheath-maintaining factors, including basement membrane and desmosomes, were used to investigate proteolytic activity of the epithelial sheath. Tissue non-specific alkaline phosphatase (TNALP) and keratin were used to investigate EMT. Epithelial sheath cells showed immunoreactivity for all three enzymes at fragmentation, which suggests that epithelial sheath disintegration is enzymatically mediated. Dental follicle cells and cementoblasts showed intense immunoreactivity for TNALP, and from portion 1 through to 3, the reaction extended from the alveolar bone-related zone to the root-related zone. Cells possessing keratin/TNALP double immunoreactivity were virtually absent. Keratin-positive epithelial sheath cells showed negligible immunoreactivity for TNALP, and epithelial cells did not appear to migrate to the dental follicle. Together, these findings suggest that a transition phenotype between epithelial cells and cementoblasts does not exist in the developing dental follicle and hence that epithelial sheath cells do not undergo EMT during initial acellular cementogenesis. In brief, this study supports the notion that cementoblasts derive from the dental follicle. PMID:24859538

  2. Mechanical properties of acellular mouse lungs after sterilization by gamma irradiation.

    PubMed

    Uriarte, Juan J; Nonaka, Paula N; Campillo, Noelia; Palma, Renata K; Melo, Esther; de Oliveira, Luis V F; Navajas, Daniel; Farré, Ramon

    2014-12-01

    Lung bioengineering using decellularized organ scaffolds is a potential alternative for lung transplantation. Clinical application will require donor scaffold sterilization. As gamma-irradiation is a conventional method for sterilizing tissue preparations for clinical application, the aim of this study was to evaluate the effects of lung scaffold sterilization by gamma irradiation on the mechanical properties of the acellular lung when subjected to the artificial ventilation maneuvers typical within bioreactors. Twenty-six mouse lungs were decellularized by a sodium dodecyl sulfate detergent protocol. Eight lungs were used as controls and 18 of them were submitted to a 31kGy gamma irradiation sterilization process (9 kept frozen in dry ice and 9 at room temperature). Mechanical properties of acellular lungs were measured before and after irradiation. Lung resistance (RL) and elastance (EL) were computed by linear regression fitting of recorded signals during mechanical ventilation (tracheal pressure, flow and volume). Static (Est) and dynamic (Edyn) elastances were obtained by the end-inspiratory occlusion method. After irradiation lungs presented higher values of resistance and elastance than before irradiation: RL increased by 41.1% (room temperature irradiation) and 32.8% (frozen irradiation) and EL increased by 41.8% (room temperature irradiation) and 31.8% (frozen irradiation). Similar increases were induced by irradiation in Est and Edyn. Scanning electron microscopy showed slight structural changes after irradiation, particularly those kept frozen. Sterilization by gamma irradiation at a conventional dose to ensure sterilization modifies acellular lung mechanics, with potential implications for lung bioengineering. PMID:25241281

  3. Preparation and characterization of an acellular bovine pericardium intended for manufacture of valve bioprostheses.

    PubMed

    Goissis, Gilberto; Giglioti, Aparecida de Fátima; Braile, Domingo Marcolino

    2011-05-01

    Major problems with biological heart valves post-implantation are associated with progressive structural deterioration and calcification attributed to glutaraldehyde processing, dead cells, and cell fragments present in the native tissue. In spite of these problems, glutaraldehyde still is the reagent of choice. The results with acellular matrix xenograft usually prepared by detergent treatment in association with enzymes are rather conflicting because while preserving mechanical properties, tissue morphology and collagen structure are process dependent. This work describes a chemical approach for the preparation of an acellular bovine pericardium matrix intended for the manufacture of heart valve bioprostheses. Cell removal was performed by an alkaline extraction in the presence of calcium salts for periods ranging from 6 to 48 h. The results showed that cell removal was achieved after 12 h, with swelling and negative charge increasing with processing time. Nevertheless, collagen fibril structure, ability to form fibrils, and stability to collagenase were progressive after 24-h processing. There was no denaturation of the collagen matrix. A process is described for the preparation of acellular bovine pericardium matrices with preserved fibril structure and morphology for the manufacture of cardiac valve bioprostheses and may be used in other applications for tissue reconstruction.

  4. Immunolocation of proteoglycans and bone-related noncollagenous glycoproteins in developing acellular cementum of rat molars.

    PubMed

    Yamamoto, T; Domon, T; Takahashi, S; Arambawatta, A K S; Wakita, M

    2004-09-01

    To elucidate the roles of proteoglycans of (PGs), bone sialoprotein (BSP), and osteopontin (OPN) in cementogenesis, their distribution was investigated in developing and established acellular cementum of rat molars by an immunoperoxidase method. To characterize PGs, antibodies against five species of glycosaminoglycans (GAGS), chondroitin-4-sulfate (C4S), chondroitin-6-sulfate (C6S), unsulfated chondroitin (C0S), dermatan sulfate (DS), and keratan sulfate (KS) were used. Routine histological staining was also applied. With onset of dentin mineralization, the initial cementum appeared on the dentin surface as a hematoxylin-stained fibril-poor layer. Subsequently, primitive principal fibers attached to the initial cementum. As the acellular cementum containing extrinsic fibers covered the initial cementum, the intal cementum formed the cemento-dentinal junction. Following immunohistochemistry at the earliest time of cementogenesis, the initial cementum was intensely immunoreactive for C4S, C6S, C0S, BSP, and OPN. After the initial cementum was embedded, neither the cemento-dentinal junction nor the cementum was immunoreactive for any GAG species. However, the cementum was immunoreactive for any GAG species. However, the cementum and cemento-dentinal were consistently immunoreactive for BSP. Although the cemento-dentinal junction was consistently immunoreactive for OPN, the remaining cementum showed no significant immunoreactivity. Thus, initial acellular cementogenesis requires a dense accumulation of PGs, BSP, and OPN, which may be associated with the mineralization process independently of collagen fibrils and initial principal fiber attachment. PMID:15278434

  5. Preparation and characterization of an acellular bovine pericardium intended for manufacture of valve bioprostheses.

    PubMed

    Goissis, Gilberto; Giglioti, Aparecida de Fátima; Braile, Domingo Marcolino

    2011-05-01

    Major problems with biological heart valves post-implantation are associated with progressive structural deterioration and calcification attributed to glutaraldehyde processing, dead cells, and cell fragments present in the native tissue. In spite of these problems, glutaraldehyde still is the reagent of choice. The results with acellular matrix xenograft usually prepared by detergent treatment in association with enzymes are rather conflicting because while preserving mechanical properties, tissue morphology and collagen structure are process dependent. This work describes a chemical approach for the preparation of an acellular bovine pericardium matrix intended for the manufacture of heart valve bioprostheses. Cell removal was performed by an alkaline extraction in the presence of calcium salts for periods ranging from 6 to 48 h. The results showed that cell removal was achieved after 12 h, with swelling and negative charge increasing with processing time. Nevertheless, collagen fibril structure, ability to form fibrils, and stability to collagenase were progressive after 24-h processing. There was no denaturation of the collagen matrix. A process is described for the preparation of acellular bovine pericardium matrices with preserved fibril structure and morphology for the manufacture of cardiac valve bioprostheses and may be used in other applications for tissue reconstruction. PMID:21595716

  6. Management of complex abdominal wall defects using acellular porcine dermal collagen.

    PubMed

    Chavarriaga, Luis Felipe; Lin, Edward; Losken, Albert; Cook, Michael W; Jeansonne, Louis O; White, Brent C; Sweeney, John F; Galloway, John R; Davis, S Scott

    2010-01-01

    Multiple techniques have been used for the repair of complex abdominal wall defects after recurrent incisional hernias with varying rates of success. Primary repair has been associated with high recurrence rates, and prosthetic mesh placement is contraindicated in contaminated surgical fields. The development of biologic prostheses has changed the approach to these difficult problems. This study evaluates the management of complex abdominal wall defects using acellular porcine dermal collagen. Between August 2006 and May 2007, 18 patients underwent abdominal wall reconstruction for complex defects with acellular porcine dermal collagen (CollaMend; Bard Inc., Warwick, RI). Patient demographics, preoperative risk factors, previous herniorrhaphy attempts, postoperative complications, recurrences, and long-term results were retrospectively reviewed. Records were reviewed at a mean follow up of 7.3 months; the recurrence rate was 44.4 per cent. A total of 38.9 per cent (seven of 18) developed a postoperative wound complications, including infection in 22.2 per cent (four of 18). All of the patients with infection required prosthesis removal as a result of encapsulation rather than incorporation of the biologic prosthesis. Acellular porcine dermal collagen has the potential for reconstruction of abdominal wall defects with postoperative wound occurrences comparable with other biologic materials. Encapsulation of the material was a major problem in cases with wound infection that required graft removal rather than local wound measures. Hernia recurrence and dehiscence of the graft were problems in noncompromised surgical fields.

  7. Neonatal Responsiveness to the Odor of Amniotic and Lacteal Fluids: A Test of Perinatal Chemosensory Continuity.

    ERIC Educational Resources Information Center

    Marlier, Luc; Schaal, Benoist; Soussignan, Robert

    1998-01-01

    Studied head-orientation response of breast-feeding neonates in paired-choice odor tests. Found that 2-day olds detected amniotic fluid and colostrum, treating them as similar sensorily and/or hedonically. Four-day olds exhibited a preference for breast milk. Three-day olds oriented longer toward the odor of their own amniotic fluid than alien…

  8. Intra-amniotic Candida albicans infection induces mucosal injury and inflammation in the ovine fetal intestine

    PubMed Central

    Nikiforou, Maria; Jacobs, Esmee M.R.; Kemp, Matthew W.; Hornef, Mathias W.; Payne, Matthew S.; Saito, Masatoshi; Newnham, John P.; Janssen, Leon E.W.; Jobe, Alan H.; Kallapur, Suhas G.; Kramer, Boris W.; Wolfs, Tim G.A.M.

    2016-01-01

    Chorioamnionitis is caused by intrauterine infection with microorganisms including Candida albicans (C.albicans). Chorioamnionitis is associated with postnatal intestinal pathologies including necrotizing enterocolitis. The underlying mechanisms by which intra-amniotic C.albicans infection adversely affects the fetal gut remain unknown. Therefore, we assessed whether intra-amniotic C.albicans infection would cause intestinal inflammation and mucosal injury in an ovine model. Additionally, we tested whether treatment with the fungistatic fluconazole ameliorated the adverse intestinal outcome of intra-amniotic C.albicans infection. Pregnant sheep received intra-amniotic injections with 107 colony-forming units C.albicans or saline at 3 or 5 days before preterm delivery at 122 days of gestation. Fetuses were given intra-amniotic and intra-peritoneal fluconazole treatments 2 days after intra-amniotic administration of C.albicans. Intra-amniotic C.albicans caused intestinal colonization and invasive growth within the fetal gut with mucosal injury and intestinal inflammation, characterized by increased CD3+ lymphocytes, MPO+ cells and elevated TNF-α and IL-17 mRNA levels. Fluconazole treatment in utero decreased intestinal C.albicans colonization, mucosal injury but failed to attenuate intestinal inflammation. Intra-amniotic C.albicans caused intestinal infection, injury and inflammation. Fluconazole treatment decreased mucosal injury but failed to ameliorate C.albicans-mediated mucosal inflammation emphasizing the need to optimize the applied antifungal therapeutic strategy. PMID:27411776

  9. 21 CFR 862.1455 - Lecithin/sphingomyelin ratio in amniotic fluid test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... fluid are used in evaluating fetal maturity. (b) Classification. Class II. ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Lecithin/sphingomyelin ratio in amniotic fluid... Clinical Chemistry Test Systems § 862.1455 Lecithin/sphingomyelin ratio in amniotic fluid test system....

  10. 21 CFR 862.1455 - Lecithin/sphingomyelin ratio in amniotic fluid test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... fluid are used in evaluating fetal maturity. (b) Classification. Class II. ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Lecithin/sphingomyelin ratio in amniotic fluid... Clinical Chemistry Test Systems § 862.1455 Lecithin/sphingomyelin ratio in amniotic fluid test system....

  11. Amniotic fluid sludge as a marker of intra-amniotic infection and histological chorioamnionitis in cervical insufficiency: a report of four cases and literature review.

    PubMed

    Paules, Cristina; Moreno, Esther; Gonzales, Ariel; Fabre, Ernesto; González de Agüero, Rafael; Oros, Daniel

    2016-01-01

    Amniotic fluid sludge (AFS) is defined as the presence of particulate matter in the amniotic fluid in close proximity to the cervix. Although its prevalence is known to correlate with the risk of preterm delivery, initial reports describe a strong association between AFS and microbial invasion of the amniotic cavity (MIAC) and histological chorioamnionitis. However, AFS is also present in uncomplicated pregnancies, and its prevalence appears to increase with gestational age. Recent evidence debates the usefulness of AFS as a marker of early preterm delivery risk. We present four cases with AFS diagnosed by transvaginal ultrasound at admission for cervical insufficiency between 20 and 24 weeks of gestation, with confirmed lower genital tract and intra-amniotic infections by amniocentesis and histological chorioamnionitis and funisitis. Our findings reinforce the presence of AFS as a useful marker of MIAC, chorioamnionitis and funisitis that increase the likelihood of preterm delivery at an extreme gestational age.

  12. Effect of high doses of gamma radiation on the functional characteristics of amniotic membrane

    NASA Astrophysics Data System (ADS)

    Singh, Rita; Purohit, Sumita; Chacharkar, M. P.

    2007-06-01

    The effect of different doses of gamma radiation viz. 25, 36 and 50 kGy on the chemical and functional characteristics of the amniotic membrane was studied. The change in the chemical structure of amniotic membranes at high doses of gamma irradiation was evaluated by means of Infrared (IR) Spectroscopy. The degradation of amnion on irradiation with gamma rays could produce a relative variation in IR absorption troughs. This kind of variation was absent in the samples irradiated to doses of 25, 36 and 50 kGy indicating no qualitative change in the material property of amnion. No significant differences in the water absorption capacity and water vapour transmission rate of amniotic membranes irradiated to different doses were observed. Impermeability of the amniotic membranes to different microorganisms was also not affected at high doses of gamma radiation. Gamma irradiation at doses of 25-50 kGy did not evoke undesirable changes in the functional properties of the amniotic membrane.

  13. An evaluation of serious neurological disorders following immunization: a comparison of whole-cell pertussis and acellular pertussis vaccines.

    PubMed

    Geier, David A; Geier, Mark R

    2004-08-01

    Serious neurological disorders reported following whole-cell pertussis in comparison to acellular pertussis vaccines were evaluated. The Vaccine Adverse Events Reporting System (VAERS) was analyzed for Emergency Department (ED) visits, life-threatening reactions, hospitalizations, disabilities, deaths, seizures, infantile spasms, encephalitis/encephalopathy, autism, Sudden Infant Death Syndrome (SIDS) and speech disorders reported with an initial onset of symptoms within 3 days following whole-cell pertussis and acellular pertussis vaccines among those residing in the US from 1997 to 1999. Controls were employed to evaluate potential biases in VAERS. Evaluations as to whether whole-cell and acellular vaccines were administered to populations of similar age and sex were undertaken because these factors might influence the study's results. Statistical increases were observed for all events examined following whole-cell pertussis vaccination in comparison to acellular pertussis vaccination, excepting cerebellar ataxia. Reporting biases were minimal in VAERS, and whole-cell and acellular pertussis vaccines were administered to populations of similar age and sex. Biologic mechanisms for the increased reactogenicity of whole-cell pertussis vaccines may stem from the fact that whole-cell pertussis vaccines contain 3,000 different proteins, whereas DTaP contains two to five proteins. Whole-cell pertussis vaccine contains known neurotoxins including: endotoxin, pertussis toxin and adenylate cyclase. Our results, and conclusions by the US Institute of Medicine, suggest an association between serious neurological disorders and whole-cell pertussis immunization. In light of the presence of a safer and at least equally efficacious acellular pertussis vaccine alternative, the Japanese and US switch to using acellular pertussis vaccine seems well justified. Other countries using whole-cell pertussis-containing vaccines should consider following suite in the near future.

  14. Amniotic fluid volume and fetal swallowing rate in sheep

    SciTech Connect

    Tomoda, S.; Brace, R.A.; Longo, L.D.

    1985-07-01

    To investigate amniotic fluid (AF) dynamics and volume regulatory mechanisms, the authors measured the concentration of radioiodinated (/sup 125/I) serum albumin (RISA), /sup 51/Cr-labeled red blood cells (Cr-RBC), and /sup 103/Ru-labeled microspheres after injection into the amniotic cavity and determined AF volume and fetal swallowing rate in 22 singleton pregnant sheep. Under normal conditions 2-3 h were required for complete mixing of RISA and Cr-RBC within AF; however, when the fetus was dead only 3-5 h were required. AF volume of 17 sheep on the 5th postoperative day averaged 975 +/- 128 ml by RISA and 986 +/- 130 ml by Cr-RBC. AF volume determined with RISA and Cr-RBC correlated well. In contrast, AF volume measurement with microspheres produced erratic results. The disappearance rate of the labels in 17 ewes on the 5th postoperative day averaged 4.9 +/- 0.7%/h for RISA and 5.5 +/- 0.7 for Cr-RBC, and the calculated rates of fetal swallowing were 935 +/- 78 ml/day by RISA and 1,085 +/- 102 by Cr-RBC. In dead fetuses the disappearance rates were almost zero, suggesting that the labels disappear mainly by swallowing. Absolute volume swallowed and swallowed volume per fetal weight correlated with gestational age. AF volume correlated with fetal weight. Radiolabeled albumin or red blood cells may be used to simultaneously measure amniotic fluid volume and the rate of fetal swallowing. Furthermore it appears that fetal swallowing increases with gestational age.

  15. Evolutionary origin and diversification of epidermal barrier proteins in amniotes.

    PubMed

    Strasser, Bettina; Mlitz, Veronika; Hermann, Marcela; Rice, Robert H; Eigenheer, Richard A; Alibardi, Lorenzo; Tschachler, Erwin; Eckhart, Leopold

    2014-12-01

    The evolution of amniotes has involved major molecular innovations in the epidermis. In particular, distinct structural proteins that undergo covalent cross-linking during cornification of keratinocytes facilitate the formation of mechanically resilient superficial cell layers and help to limit water loss to the environment. Special modes of cornification generate amniote-specific skin appendages such as claws, feathers, and hair. In mammals, many protein substrates of cornification are encoded by a cluster of genes, termed the epidermal differentiation complex (EDC). To provide a basis for hypotheses about the evolution of cornification proteins, we screened for homologs of the EDC in non-mammalian vertebrates. By comparative genomics, de novo gene prediction and gene expression analyses, we show that, in contrast to fish and amphibians, the chicken and the green anole lizard have EDC homologs comprising genes that are specifically expressed in the epidermis and in skin appendages. Our data suggest that an important component of the cornified protein envelope of mammalian keratinocytes, that is, loricrin, has originated in a common ancestor of modern amniotes, perhaps during the acquisition of a fully terrestrial lifestyle. Moreover, we provide evidence that the sauropsid-specific beta-keratins have evolved as a subclass of EDC genes. Based on the comprehensive characterization of the arrangement, exon-intron structures and conserved sequence elements of EDC genes, we propose new scenarios for the evolutionary origin of epidermal barrier proteins via fusion of neighboring S100A and peptidoglycan recognition protein genes, subsequent loss of exons and highly divergent sequence evolution. PMID:25169930

  16. Identification of emergent motion compartments in the amniote embryo

    PubMed Central

    Loganathan, Rajprasad; Little, Charles D; Joshi, Pranav; Filla, Michael B; Cheuvront, Tracey J; Lansford, Rusty; Rongish, Brenda J

    2014-01-01

    Abstract The tissue scale deformations (≥1mm) required to form an amniote embryo are poorly understood. Here, we studied ∼400 μm-sized explant units from gastrulating quail embryos. The explants deformed in a reproducible manner when grown using a novel vitelline membrane-based culture method. Time-lapse recordings of latent embryonic motion patterns were analyzed after disk-shaped tissue explants were excised from three specific regions near the primitive streak: 1) anterolateral epiblast, 2) posterolateral epiblast, and 3) the avian organizer (Hensen's node). The explants were cultured for 8 hours—an interval equivalent to gastrulation. Both the anterolateral and the posterolateral epiblastic explants engaged in concentric radial/centrifugal tissue expansion. In sharp contrast, Hensen's node explants displayed Cartesian-like, elongated, bipolar deformations—a pattern reminiscent of axis elongation. Time-lapse analysis of explant tissue motion patterns indicated that both cellular motility and extracellular matrix fiber (tissue) remodeling take place during the observed morphogenetic deformations. As expected, treatment of tissue explants with a selective Rho-Kinase (p160ROCK) signaling inhibitor, Y27632, completely arrested all morphogenetic movements. Microsurgical experiments revealed that lateral epiblastic tissue was dispensable for the generation of an elongated midline axis— provided that an intact organizer (node) is present. Our computational analyses suggest the possibility of delineating tissue-scale morphogenetic movements at anatomically discrete locations in the embryo. Further, tissue deformation patterns, as well as the mechanical state of the tissue, require normal actomyosin function. We conclude that amniote embryos contain tissue-scale, regionalized morphogenetic motion generators, which can be assessed using our novel computational time-lapse imaging approach. These data and future studies—using explants excised from overlapping

  17. Evolutionary Origin and Diversification of Epidermal Barrier Proteins in Amniotes

    PubMed Central

    Strasser, Bettina; Mlitz, Veronika; Hermann, Marcela; Rice, Robert H.; Eigenheer, Richard A.; Alibardi, Lorenzo; Tschachler, Erwin; Eckhart, Leopold

    2014-01-01

    The evolution of amniotes has involved major molecular innovations in the epidermis. In particular, distinct structural proteins that undergo covalent cross-linking during cornification of keratinocytes facilitate the formation of mechanically resilient superficial cell layers and help to limit water loss to the environment. Special modes of cornification generate amniote-specific skin appendages such as claws, feathers, and hair. In mammals, many protein substrates of cornification are encoded by a cluster of genes, termed the epidermal differentiation complex (EDC). To provide a basis for hypotheses about the evolution of cornification proteins, we screened for homologs of the EDC in non-mammalian vertebrates. By comparative genomics, de novo gene prediction and gene expression analyses, we show that, in contrast to fish and amphibians, the chicken and the green anole lizard have EDC homologs comprising genes that are specifically expressed in the epidermis and in skin appendages. Our data suggest that an important component of the cornified protein envelope of mammalian keratinocytes, that is, loricrin, has originated in a common ancestor of modern amniotes, perhaps during the acquisition of a fully terrestrial lifestyle. Moreover, we provide evidence that the sauropsid-specific beta-keratins have evolved as a subclass of EDC genes. Based on the comprehensive characterization of the arrangement, exon–intron structures and conserved sequence elements of EDC genes, we propose new scenarios for the evolutionary origin of epidermal barrier proteins via fusion of neighboring S100A and peptidoglycan recognition protein genes, subsequent loss of exons and highly divergent sequence evolution. PMID:25169930

  18. Multiple lineages of ancient CR1 retroposons shaped the early genome evolution of amniotes.

    PubMed

    Suh, Alexander; Churakov, Gennady; Ramakodi, Meganathan P; Platt, Roy N; Jurka, Jerzy; Kojima, Kenji K; Caballero, Juan; Smit, Arian F; Vliet, Kent A; Hoffmann, Federico G; Brosius, Jürgen; Green, Richard E; Braun, Edward L; Ray, David A; Schmitz, Jürgen

    2015-01-01

    Chicken repeat 1 (CR1) retroposons are long interspersed elements (LINEs) that are ubiquitous within amniote genomes and constitute the most abundant family of transposed elements in birds, crocodilians, turtles, and snakes. They are also present in mammalian genomes, where they reside as numerous relics of ancient retroposition events. Yet, despite their relevance for understanding amniote genome evolution, the diversity and evolution of CR1 elements has never been studied on an amniote-wide level. We reconstruct the temporal and quantitative activity of CR1 subfamilies via presence/absence analyses across crocodilian phylogeny and comparative analyses of 12 crocodilian genomes, revealing relative genomic stasis of retroposition during genome evolution of extant Crocodylia. Our large-scale phylogenetic analysis of amniote CR1 subfamilies suggests the presence of at least seven ancient CR1 lineages in the amniote ancestor; and amniote-wide analyses of CR1 successions and quantities reveal differential retention (presence of ancient relics or recent activity) of these CR1 lineages across amniote genome evolution. Interestingly, birds and lepidosaurs retained the fewest ancient CR1 lineages among amniotes and also exhibit smaller genome sizes. Our study is the first to analyze CR1 evolution in a genome-wide and amniote-wide context and the data strongly suggest that the ancestral amniote genome contained myriad CR1 elements from multiple ancient lineages, and remnants of these are still detectable in the relatively stable genomes of crocodilians and turtles. Early mammalian genome evolution was thus characterized by a drastic shift from CR1 prevalence to dominance and hyperactivity of L2 LINEs in monotremes and L1 LINEs in therians. PMID:25503085

  19. Sutureless Fixation of Amniotic Membrane for Therapy of Ocular Surface Disorders

    PubMed Central

    Kotomin, Ilya; Valtink, Monika; Hofmann, Kai; Frenzel, Annika; Morawietz, Henning; Werner, Carsten; Funk, Richard H. W.; Engelmann, Katrin

    2015-01-01

    Amniotic membrane is applied to the diseased ocular surface to stimulate wound healing and tissue repair, because it releases supportive growth factors and cytokines. These effects fade within about a week after application, necessitating repeated application. Generally, amniotic membrane is fixed with sutures to the ocular surface, but surgical intervention at the inflamed or diseased site can be detrimental. Therefore, we have developed a system for the mounting of amniotic membrane between two rings for application to a diseased ocular surface without surgical intervention (sutureless amniotic membrane transplantation). With this system, AmnioClip, amniotic membrane can be applied like a large contact lens. First prototypes were tested in an experiment on oneself for wearing comfort. The final system was tested on 7 patients in a pilot study. A possible influence of the ring system on the biological effects of amniotic membrane was analyzed by histochemistry and by analyzing the expression of vascular endothelial growth factor-A (VEGF-A), hepatocyte growth factor (HGF), fibroblast growth factor 2 (FGF 2) and pigment epithelium-derived factor (PEDF) from amniotic membranes before and after therapeutic application. The final product, AmnioClip, showed good tolerance and did not impair the biological effects of amniotic membrane. VEGF-A and PEDF mRNA was expressed in amniotic membrane after storage and mounting before transplantation, but was undetectable after a 7-day application period. Consequently, transplantation of amniotic membranes with AmnioClip provides a sutureless and hence improved therapeutic strategy for corneal surface disorders. Trial Registration ClinicalTrials.gov NCT02168790 PMID:25955359

  20. Amniotic fluid fluorescence polarisation values for assessing fetal lung maturation.

    PubMed Central

    Legge, M; Potter, H C

    1981-01-01

    The fluorescence probe 1,6, diphenyl-1,3,5-hexatriene was used to determine the micro-viscosity of third trimester amniotic fluid samples utilising an Aminco Bowman spectrofluorometer fitted with a polariser prism. The results were related to the lecithin sphingomyelin (L:S) ratio. A fluorescence polarisation value of 0.360 corresponded to an L:S ratio of 2.0 in uncomplicated and diabetic pregnancies. Pregnancies complicated by Rhesus disease gave significantly different fluorescence polarisation values when compared with the uncomplicated pregnancies. PMID:7309891

  1. Evaluating acellular versus cellular perfusate composition during prolonged ex vivo lung perfusion after initial cold ischaemia for 24 hours.

    PubMed

    Becker, Simon; Steinmeyer, Jasmin; Avsar, Murat; Höffler, Klaus; Salman, Jawad; Haverich, Axel; Warnecke, Gregor; Ochs, Matthias; Schnapper, Anke

    2016-01-01

    Normothermic ex vivo lung perfusion (EVLP) has developed as a powerful technique to evaluate particularly marginal donor lungs prior to transplantation. In this study, acellular and cellular perfusate compositions were compared in an identical experimental setting as no consensus has been reached on a preferred technique yet. Porcine lungs underwent EVLP for 12 h on the basis of an acellular or a cellular perfusate composition after 24 h of cold ischaemia as defined organ stress. During perfusion, haemodynamic and respiratory parameters were monitored. After EVLP, the lung condition was assessed by light and transmission electron microscopy. Aerodynamic parameters did not show significant differences between groups and remained within the in vivo range during EVLP. Mean oxygenation indices were 491 ± 39 in the acellular group and 513 ± 53 in the cellular group. Groups only differed significantly in terms of higher pulmonary artery pressure and vascular resistance in the cellular group. Lung histology and ultrastructure were largely well preserved after prolonged EVLP and showed only minor structural alterations which were similarly present in both groups. Prolonged acellular and cellular EVLP for 12 h are both feasible with lungs prechallenged by ischaemic organ stress. Physiological and ultrastructural analysis showed no superiority of either acellular or cellular perfusate composition.

  2. Amniotic fluid embolism: pathophysiology and new strategies for management.

    PubMed

    Kanayama, Naohiro; Tamura, Naoaki

    2014-06-01

    The registry program of amniotic fluid embolism (AFE) in Japan started in 2003. More than 400 hundred clinical diagnosed amniotic fluid embolism has been accumulated. Those data showed that there were two etiologies of AFE: the fetal materials create physical obstructions in the maternal microvessels in various organs, such as the lung; and (ii) the liquids cause an anaphylactoid reaction that leads to pulmonary vasospasm and activation of platelets, white blood cells and/or complements. The clinical findings showed that AFE was characterized mainly by cardiopulmonary collapse, the other involves the presence of disseminated intravascular coagulation (DIC) and atonic bleeding. Zinc coproporphyrin-1, sialyl Tn antigen (STN), complement C3, C4 and interleukin-8 have been used as serum markers of AFE. The levels of zinc coproporphyrin-1 and STN were increased in cardiopulmonary collapse type AFE, and a marked reduction of C3 and C4 was observed in DIC type AFE. At the primary medical institution, initial treatments for shock airway management, vascular management, fluid replacement, administration of anti-DIC therapy such as antithrombin, and administration of fresh frozen plasma should be provided. C1 esterase inhibitor activity in AFE cases was significantly lower than those of normal pregnant women. C1 esterase inhibitor may be a promising candidate of treatment of AFE.

  3. Effects of octreotide acetate and amniotic membrane on wound healing in experimental glaucoma surgery.

    PubMed

    Demir, Tamer; Turgut, Burak; Celiker, Ulku; Ozercan, Ibrahim; Ulas, Fatih; Akyol, Nuray

    2003-09-01

    Wound healing affects the success of glaucoma filtering surgery. Antimetabolites and antifibrotic agents are used in the modulation of surgical trauma. This study is performed to evaluate the effects of amniotic membrane and octreotide acetate on wound healing. Thirty pigmented rabbits were divided into three groups each including 10 animals. Trabeculectomy and topical postoperative prednisolone sodium phosphate four times daily for 7 days were applied to one eye of all the rabbits. After trabeculectomy, octreotide 10 microg three times daily applied topically to the octreotide group for 14 days and amniotic membrane transplantation was performed by suturing amniotic membrane between scleral flap and sclera to the amniotic membrane group. The operated eyes of the rabbits were enucleated on the 14th day of the operation and histopathological specimens were obtained from the bleb sites and they were evaluated by light microscope. Fibroblast and macrophage number per cm2 were counted and the average values were calculated. Compared with the control group, the fibroblasts and macrophages significantly decreased in the other two groups (p < 0.0001, p < 0.0001, respectively). The mean number of fibroblasts was lower than those of the control and amniotic membrane groups (p < 0.0001, p < 0.0001, respectively). Similarly the mean macrophage number was significantly lower in the octreotide group versus the control and amniotic membrane groups (p < 0.0001, p < 0.01, respectively). Octreotide administration and amniotic membrane transplantation might be alternative treatments in modulating the wound healing after trabeculectomy.

  4. Prey processing in amniotes: biomechanical and behavioral patterns of food reduction.

    PubMed

    Reilly, S M; McBrayer, L D; White, T D

    2001-03-01

    In this paper we examine the biomechanics of prey processing behavior in the amniotes. Whether amniotes swallow prey items whole or swallow highly processed slurries or boluses of food, they share a common biomechanical system where hard surfaces (teeth or beaks) are brought together on articulated jaws by the actions of adductor muscles to grasp and process food. How have amniotes modified this basic system to increase the chewing efficiency of the system? To address this question we first examine the primitive condition for prey processing representative of many of the past and present predatory amniotes. Because herbivory is expected to be related to improved prey processing in the jaws we review patterns of food processing mechanics in past and present herbivores. Herbivory has appeared numerous times in amniotes and several solutions to the task of chewing plant matter have appeared. Birds have abandoned jaw chewing in favor of a new way to chew--with the gut--so we will detour from the jaws to examine the appearance of gut chewing in the archosaurs. We will then fill in the gaps among amniote taxa with a look at some new data on patterns of prey processing behavior and jaw mechanics in lizards. Finally, we examine evolutionary patterns of amniote feeding mechanism and how correlates of chewing relate to the need to increase the efficiency of prey processing in order to facilitate increased metabolic rate and activity.

  5. Florid pustular dermatitis of breast: A case report on a unusual complication from acellular dermal matrix use

    PubMed Central

    James, Justin; Jackson, Lee; Saunders, Christobel

    2016-01-01

    Introduction Idiopathic erythematous reaction of the breast (Red breast syndrome) is a known complication following breast reconstruction with acellular dermal matrix. However pustular dermatitis like presentation is not previously known. Presentation of case We present a 42-year-old lady who developed bilateral pustular dermatitis like appearance following breast reconstruction with acellular dermal matrix slings. Though surgical washout was done, both expanders and flex HD could be preserved. Discussion Acellular dermal matrix use is the only possible explanation for such a presentation and this can be considered a variant of red breast syndrome. Conclusion Pustular dermatitis like presentation can be associated with acelluar dermal matrix use and should be considered in similar clinical presentations, since this can avoid unnecessary surgical procedures. PMID:27058152

  6. The potential of mesenchymal stem cells derived from amniotic membrane and amniotic fluid for neuronal regenerative therapy.

    PubMed

    Kim, Eun Young; Lee, Kyung-Bon; Kim, Min Kyu

    2014-03-01

    The mesenchymal stem cells (MSCs), which are derived from the mesoderm, are considered as a readily available source for tissue engineering. They have multipotent differentiation capacity and can be differentiated into various cell types. Many studies have demonstrated that the MSCs identified from amniotic membrane (AM-MSCs) and amniotic fluid (AF-MSCs) are shows advantages for many reasons, including the possibility of noninvasive isolation, multipotency, self-renewal, low immunogenicity, anti-inflammatory and nontumorigenicity properties, and minimal ethical problem. The AF-MSCs and AM-MSCs may be appropriate sources of mesenchymal stem cells for regenerative medicine, as an alternative to embryonic stem cells (ESCs). Recently, regenerative treatments such as tissue engineering and cell transplantation have shown potential in clinical applications for degenerative diseases. Therefore, amnion and MSCs derived from amnion can be applied to cell therapy in neuro-degeneration diseases. In this review, we will describe the potential of AM-MSCs and AF-MSCs, with particular focus on cures for neuronal degenerative diseases.

  7. Adipose tissue regeneration in vivo using micronized acellular allogenic dermis as an injectable scaffold.

    PubMed

    Lee, Hee Young; Yang, Hyun Jin; Rhie, Jong Won; Han, Ki Talk

    2014-10-01

    Over the past few years, the clinical use of injectable artificial materials in plastic surgery has increased. In addition, autologous lipoimplantation is being performed for volume replacement of soft tissue for aesthetic purposes. In this study, acellular allogenic dermis was utilized as a scaffold for the culturing of preadipocytes, confirming the possibility of three-dimensional proliferation of progenitor cells, the eventual differentiation of stromal cells in adipose tissue into the adipocytes, and the in vivo implantation of such adipocytes to form fat tissue. Preadipocytes, recently called ASCs (adipose tissue-derived stromal/stem cells), were cultured in acellular allogenic dermis, successfully attached to the dermal particles in a three-dimensional structure, and proliferated, differentiated, and eventually formed a cluster. For the in vivo implantation, four groups were formed: the first group was cultured within the dermal scaffold for 24 h before implantation (24-h preconditioned group), the second group was induced for differentiation for 10 days before implantation (10-day preconditioned group), the third group was implanted immediately after cell propagation (nonpreconditioned group), and the control group was implanted with only dermal scaffold. In vivo implanted preadipocytes showed great differentiation into adipocytes within the dermal scaffolds. Also, the 10-day preconditioned group showed a greater volume of fat tissue compared to the 24-h preconditioned group. From these results, we confirmed that after a three-dimensional culture in acellular allogenic dermis, implanted preadipocytes formed a greater amount of fat tissue and that this could be a possible effective method for future soft tissue restoration.

  8. Reconstruction of a Recurrent First Dorsal Web Space Defect using Acellular Dermis

    PubMed Central

    Buck, Donald W.; Kloeters, Oliver; Eo, SuRak; Jones, Neil F.

    2007-01-01

    Oncologic defects of the hand can be problematic for the reconstructive surgeon. These defects may require a delay in definitive coverage until clear margins of resection can be obtained, which can result in a prolonged period of painful dressing changes and increased risk of soft-tissue infection. In addition, reconstructive options for oncologic defects are often limited to skin grafting, which can yield functional deficits secondary to contracted healing. Currently, there is no definitive method for preventing skin graft contracture; however, acellular dermis has been proposed as a biomechanical scaffold to enhance subsequent skin graft healing and slow this functionally debilitating process. Here, we present a patient with recurrent melanoma of the first dorsal web space. After re-resection of the melanoma, the 11 cm × 5 cm defect was reconstructed using acellular dermis as temporary coverage to allow ample time for permanent section results. Ten days later, after confirming negative margins of resection, a split-thickness skin graft (STSG) was applied over the vascularized neo-dermis. Follow-up clinical examination and the Disabilities of the Arm, Shoulder, and Hand (DASH) questionnaires were used to assess outcome. At 7 months, the patient had no recurrence of melanoma and a DASH functional reduction of only 6.9%. After approximately 18 months, the patient’s wounds had healed with excellent cosmetic and functional results, without any evidence of a web space contracture. These observations suggest that acellular dermis is a useful adjunct for wound coverage of the hand, particularly in areas of functional importance, such as the first dorsal web space. PMID:18780060

  9. Current opinions on indications and algorithms for acellular dermal matrix use in primary prosthetic breast reconstruction.

    PubMed

    Vu, Michael M; Kim, John Y S

    2015-06-01

    Acellular dermal matrix (ADM) is widely used in primary prosthetic breast reconstruction. Many indications and contraindications to use ADM have been reported in the literature, and their use varies by institution and surgeon. Developing rational, tested algorithms to determine when ADM is appropriate can significantly improve surgical outcomes and reduce costs associated with ADM use. We review the important indications and contraindications, and discuss the algorithms that have been put forth so far. Further research into algorithmic decision-making for ADM use will allow optimized balancing of cost with risk and benefit. PMID:26161304

  10. Current opinions on indications and algorithms for acellular dermal matrix use in primary prosthetic breast reconstruction

    PubMed Central

    Vu, Michael M.

    2015-01-01

    Acellular dermal matrix (ADM) is widely used in primary prosthetic breast reconstruction. Many indications and contraindications to use ADM have been reported in the literature, and their use varies by institution and surgeon. Developing rational, tested algorithms to determine when ADM is appropriate can significantly improve surgical outcomes and reduce costs associated with ADM use. We review the important indications and contraindications, and discuss the algorithms that have been put forth so far. Further research into algorithmic decision-making for ADM use will allow optimized balancing of cost with risk and benefit. PMID:26161304

  11. Inhibition of glycogen synthase kinase-3 (GSK3) promotes the neural differentiation of full-term amniotic fluid-derived stem cells towards neural progenitor cells.

    PubMed

    Gao, Liyang; Zhao, Mingyan; Ye, Wei; Huang, Jinzhi; Chu, Jiaqi; Yan, Shouquan; Wang, Chaojun; Zeng, Rong

    2016-08-01

    The amniotic fluid has a heterogeneous population of cells. Some human amniotic fluid-derived stem (hAFS) cells have been shown to harbor the potential to differentiate into neural cells. However, the neural differentiation efficiency of hAFS cells remains low. In this study, we isolated CD117-positive hAFS cells from amniotic fluid and then examined the pluripotency of these cells through the formation of embryoid bodies (EBs). Additionally, we induced the neural differentiation of these cells using neuroectodermal medium. This study revealed that the GSK3-beta inhibitor SB216763 was able to stimulate the proliferation of CD117-positive hAFS cells without influencing their undifferentiated state. Moreover, SB216763 can efficiently promote the neural differentiation of CD117-positive hAFS cells towards neural progenitor cells in the presence of DMEM/F12 and N2 supplement. These findings provide an easy and low-cost method to maintain the proliferation of hAFS cells, as well as induce an efficacious generation of neural progenitor cells from hAFS cells. Such induction of the neural commitment of hAFS cells may provide an option for the treatment of neurodegenerative diseases by hAFS cells-based therapies.

  12. Using SELDI-TOF Mass Spectrometry on Amniotic Fluid and for Clinical Proteomics and Theranostics in Disorders of Pregnancy

    PubMed Central

    Buhimschi, Irina A.

    2013-01-01

    Clinical proteomics encompasses a multitude of experimental approaches, tools and techniques based on proteomics technology that are directly aimed to accelerate and improve diagnosis and treatment of human diseases. Surface-enhanced laser desorption ionization time-of-flight (SELDI-TOF) mass spectrometry is a variant of matrix-enhanced laser desorption ionization (MALDI) that makes use of chemically-modified surfaces to reduce the complexity of biological samples prior to separation in the mass analyzer. Compared to other proteomic techniques, SELDI has several important advantages such as ability to analyze complex biological samples with minimal pre-processing, ease of handling and high throughput. Importantly, once the biomarker or combination of biomarkers with potential clinical value has been established, validation analyses can be conducted in close proximity to clinical settings which is important for establishing the utility of new diagnostics in clinical decision making and perhaps future theranostic interventions. This chapter provides protocols for experimental design and methodology aimed at 1) UUdiscovering biologically relevant biomarkers in amniotic fluid using SELDI-TOF; 2) validating the clinical utility of the biomarkers as new diagnostics; 3) translating the biomarker findings into pathophysiological phenomena to provide further insight and extend the current understanding of the disease process. Many of the principles described herein for amniotic fluid could be generalized to studies involving other types of biological samples and other clinical questions. PMID:22083824

  13. Prevalence and Clinical Significance of Sterile Intra-amniotic Inflammation in Patients with Preterm Labor and Intact Membranes

    PubMed Central

    Romero, Roberto; Miranda, Jezid; Chaiworapongsa, Tinnakorn; Korzeniewski, Steven J.; Chaemsaithong, Piya; Gotsch, Francesca; Dong, Zhong; Ahmed, Ahmed I.; Yoon, Bo Hyun; Hassan, Sonia S.; Kim, Chong J.; Yeo, Lami

    2014-01-01

    Problem Inflammation and infection play a major role in preterm birth. The purpose of this study was to: 1) determine the prevalence and clinical significance of sterile intra-amniotic inflammation; and 2) examine the relationship between amniotic fluid (AF) concentrations of high mobility group box-1 (HMGB1) and the interval from amniocentesis-to-delivery in patients with sterile intra-amniotic inflammation. Method of Study AF samples obtained from 135 women with preterm labor and intact membranes were analyzed using cultivation techniques as well as broad-range PCR and mass spectrometry (PCR/ESI-MS). Sterile intra-amniotic inflammation was defined when patients with negative AF cultures and without evidence of microbial footprints had intra-amniotic inflammation (AF interleukin-6 ≥ 2.6 ng/mL). Results 1) The frequency of sterile intra-amniotic inflammation was significantly greater than that of microbial-associated intra-amniotic inflammation [26% (35/135) vs. 11% (15/135); (p=0.005)]; 2) patients with sterile intra-amniotic inflammation delivered at comparable gestational ages, had similar rates of acute placental inflammation and adverse neonatal outcomes as patients with microbial-associated intra-amniotic inflammation; and 3) patients with sterile intra-amniotic inflammation and high AF concentrations of HMGB1 (≥ 8.55 ng/mL) delivered earlier than those with low AF concentrations of HMGB1 (p=0.02). Conclusions 1) sterile intra-amniotic inflammation is more frequent than microbial-associated intra-amniotic inflammation; and 2) we propose that danger signals participate in sterile intra-amniotic inflammation in the setting of preterm labor. PMID:25078709

  14. Fetal Renal Structure and the Genesis of Amniotic Fluid Disorders

    PubMed Central

    Naeye, Richard L.; Blanc, William A.

    1972-01-01

    Newborn recipient twins in the transplacental transfusion syndrome have dilated renal tubules, enlarged bladders and an increased urinary output in the early neonatal period, suggesting that increased fetal micturition is responsible for hydramnios in the syndrome. There is the possibility that such micturition contributes to hydramnios in other disorders as well. In the present study, renal tubules were found to be dilated in single-born infants with a diverse group of disorders having hydramnios as a common feature. Many of the neonates had hypoplastic lungs, an abnormality whose role in the hydramnios is undetermined. Renal tubular lumina were of normal size in neonates associated with oligohydramnios due to chronic leak of amniotic fluid. Donor members of parabiotic transplacental transfusion pairs had contracted renal tubules which helps to explain their oligohydramnios. ImagesFig 1Fig 2Fig 3Fig 4 PMID:4558222

  15. [Application of amniotic membrane dressings in patients with skin damage].

    PubMed

    Carrera González, Elier; Noa Hernández, Jose Eduardo; Marín Rojo, Carlos A

    2011-01-01

    The application of amniotic membranes in patients diagnosed with skin damage is a valid treatment option. A care plan following the Virginia Henderson model and NANDA, NOC and NIC taxonomy was applied to 36 patients admitted to the Dr. Miguel Enríquez hospital with different cutaneous lesions. This membrane has already been used for years due to its healing properties. These are attributed to antimicrobial properties reducing infection risk and promoting epithelial activity. They can decrease the need for the use of antibiotics, expendable materials, and can be applied during long periods of healing. This decreases the cost of wide spectrum antibiotic treatments, as well as the time patients spend in hospital. We present the results of this application in cases with several types of skin lesions.

  16. A Mini Overview of Isolation, Characterization and Application of Amniotic Fluid Stem Cells

    PubMed Central

    Gholizadeh-Ghalehaziz, Shiva; Farahzadi, Raheleh; Fathi, Ezzatollah; Pashaiasl, Maryam

    2015-01-01

    Amniotic fluid represents rich sources of stem cells that can be used in treatments for a wide range of diseases. Amniotic fluid- stem cells have properties intermediate between embryonic and adult mesenchymal stem cells which make them particularly attractive for cellular regeneration and tissue engineering. Furthermore, scientists are interested in these cells because they come from the amniotic fluid that is routinely discarded after birth. In this review we give a brief introduction of amniotic fluid followed by a description of the cells present within this fluid and aim to summarize the all existing isolation methods, culturing, characterization and application of these cells. Finally, we elaborate on the differentiation and potential for these cells to promote regeneration of various tissue defects, including fetal tissue, the nervous system, heart, lungs, kidneys, bones, and cartilage in the form of table. PMID:26634059

  17. A Mini Overview of Isolation, Characterization and Application of Amniotic Fluid Stem Cells.

    PubMed

    Gholizadeh-Ghalehaziz, Shiva; Farahzadi, Raheleh; Fathi, Ezzatollah; Pashaiasl, Maryam

    2015-11-01

    Amniotic fluid represents rich sources of stem cells that can be used in treatments for a wide range of diseases. Amniotic fluid- stem cells have properties intermediate between embryonic and adult mesenchymal stem cells which make them particularly attractive for cellular regeneration and tissue engineering. Furthermore, scientists are interested in these cells because they come from the amniotic fluid that is routinely discarded after birth. In this review we give a brief introduction of amniotic fluid followed by a description of the cells present within this fluid and aim to summarize the all existing isolation methods, culturing, characterization and application of these cells. Finally, we elaborate on the differentiation and potential for these cells to promote regeneration of various tissue defects, including fetal tissue, the nervous system, heart, lungs, kidneys, bones, and cartilage in the form of table.

  18. Comparative ontogeny and phylogeny of the upper jaw skeleton in amniotes.

    PubMed

    Richman, Joy M; Buchtová, Marcela; Boughner, Julia C

    2006-05-01

    The morphology, position, and presence of the upper jaw bones vary greatly across amniote taxa. In this review, we compare the development and anatomy of upper jaw bones from the three living amniote groups: reptiles, birds, and mammals. The study of reptiles is particularly important as comparatively little is known about the embryogenesis of the jaw in this group. Our review covers the ontogeny and phylogeny of membranous bones in the face. The aim is to identify conserved embryonic processes that may exist among the three major amniote groups. Finally, we discuss how temporal and spatial regulation of preosseous condensations and ossification centers can lead to variation in the morphology of amniote upper jaw bones. PMID:16496291

  19. Amniotic fluid phospholipids after maternal administration of dexamethasone.

    PubMed

    Farrell, P M; Engle, M J; Zachman, R D; Curet, L B; Morrison, J C; Rao, A V; Poole, W K

    1983-02-15

    The administration of corticosteroids to pregnant women in premature labor can accelerate fetal lung development and potentially prevent neonatal respiratory distress syndrome (RDS). Controversy exists, however, as to whether amniotic fluid phospholipid indices of lung maturation are influenced by such treatment. Without a suitable test for evaluating the fetal response to corticosteroids, there is no method of recognizing whether and when lung development has been stimulated. In an attempt to resolve this issue, we carried out a study of amniotic fluid phospholipids as part of the National Institutes of Health multicenter trial of prenatal corticosteroids. Amniocenteses were performed before the administration of either steroid hormone or placebo and approximately 1 week after a series of four injections was initiated. Analysis of the ratio of lecithin (phosphatidylcholine) to sphingomyelin (L/S ratio) revealed nearly identical values initially and no significant difference in the posttreatment means when 25 steroid-treated pregnancies were compared to 20 control pregnancies. Although there were significant increases in both groups during the interval between amniocenteses, no statistical difference was found in the extent of change in L/S ratios between the two groups, when pretreatment values were compared with those obtained an average of 1 week later. In addition to evaluating L/S ratios, we performed an assessment of phospholipid concentrations in 17 pregnancies before and after administration of dexamethasone. This revealed no detectable phosphatidylglycerol. There were increases in the absolute concentrations of phosphatidylcholine and disaturated phosphatidylcholine, but these changes were relatively modest in magnitude and could be attributable to either advanced gestational age or dexamethasone. Our results demonstrate that current tests of fetal lung maturity do not provide a routine means for prenatal detection of pulmonary maturational responses to

  20. Amniotic fluid as a source of multipotent cells for clinical use.

    PubMed

    Young, Bruce K; Chan, Michael K; Liu, Li; Basch, Ross S

    2016-04-01

    Amniotic fluid cells (AFC) from 2nd trimester amniocentesis have been found to be a source of multipotent stem cells which might overcome the limitations of expansion, histocompatibility, tumorigenesis, and ethical issues associated with using human embryonic cells, umbilical cord, cord blood, bone marrow, and induced pluripotent cells. Previous work by our group and others demonstrated multipotency and the ability to grow well in culture. However, all these studies were done in media containing fetal calf serum. We sought to observe the properties of AFC grown in serum-free media as that would be required for clinical transplantation in humans. Fresh samples were obtained from three patients, and each sample divided into a culture whose cells were not exposed to fetal calf serum, and the other half into a standard culture medium containing fetal calf serum. Doubling time and stem cell marker expression by flow cytometry were assessed. Differentiation to neural, osteoid, and chondrogenic lineages was induced using appropriate media and confirmed by fluorescent microscopy, histology, and immunohistochemistry. There were no statistically significant differences between cells grown serum-free and in standard media in any of these parameters. The data supports the possibility of clinical use of AFC in stem cell transplantation.

  1. [Green amniotic fluid as initial symptom of high intestinal obstruction in infants].

    PubMed

    Swarte, R M; Hack, W W; Roex, A J; Ekkelkamp, S

    1997-01-25

    At the birth of two children the amniotic fluid was green colored. The Apgar scores were good. Because of bilious vomiting and food retention, respectively, an open stomach tube was inserted, out of which bilious stomach contains were drained. The cause of green amniotic fluid was not meconium production or infection with Listeria monocytogenes, but mixing with green bile. At further investigation the children both proved to have a high intestinal obstruction distal of the papilla duodeni major.

  2. Creation and implantation of acellular rat renal ECM-based scaffolds

    PubMed Central

    Peloso, Andrea; Ferrario, Jacopo; Maiga, Benedetta; Benzoni, Ilaria; Bianco, Carolina; Citro, Antonio; Currao, Manuela; Malara, Alessandro; Gaspari, Annalisa; Balduini, Alessandra; Abelli, Massimo; Piemonti, Lorenzo; Dionigi, Paolo; Orlando, Giuseppe; Maestri, Marcello

    2015-01-01

    Abstract Kidney transplantation is the only potentially curative treatment for patient facing end-stage renal disease, and it is now routinely used. Its use is mainly limited by the supply of transplantable donor organs, which far exceeds the demand. Regenerative medicine and tissue engineering offer promising means for overcoming this shortage. In the present study, we developed and validated a protocol for producing acellular rat renal scaffolds. Left kidneys were removed from 26 male Lewis rats (weights: 250–350 g) and decellularized by means of aortic anterograde perfusion with ionic and anionic detergents (Triton X-100 1% and SDS 1%, respectively). 19 scaffolds thus obtained (and contralateral native kidneys as controls) were deeply characterized in order to evaluate the decellularization quality, the preservation of extracellular matrix components and resultant micro-angioarchitecture structure. The other 7 were transplanted into 7 recipient rats that had undergone unilateral nephrectomy. Recipients were sacrificed on post-transplantation day 7 and the scaffolds subjected to histologic studies. The dual-detergent protocol showed, with only 5 h of perfusion per organ, to obtain thoroughly decellularized renal scaffolds consisting almost exclusively of extracellular matrix. Finally the macro- and the microarchitecture of the renal parenchyma were well preserved, and the grafts were implanted with ease. Seven days after transplant, the scaffolds were morphologically intact although all vascular structures were obstructed with thrombi. Production and implantation of acellular rat renal scaffolds is a suitable platform for further studies on regenerative medicine and tissue engineering. PMID:26186418

  3. Aseptic versus Sterile Acellular Dermal Matrices in Breast Reconstruction: An Updated Review

    PubMed Central

    Mendenhall, Shaun D.; Neumeister, Michael W.; Cederna, Paul S.; Momoh, Adeyiza O.

    2016-01-01

    Background: As the use of acellular dermal matrices in breast reconstruction has become more commonplace and efforts are made to improve on postoperative outcomes, the method of acellular dermal matrix (ADM) processing (aseptic versus sterile) has become a subject of interest. This article provides an updated overview of the critical aspects of ADM processing in addition to application of ADMs in single- and two-stage breast reconstruction, a review of the morbidity associated with ADM use, and alternatives. Methods: A literature review was performed in PubMed identifying recent systematic reviews, meta-analyses, and head-to-head comparisons on aseptically processed ADM and sterile-processed ADM in implant-based breast reconstruction. Results: Recent meta-analyses have shown a 2- to 3-fold increase in infections and tissue expander/implant explantation rates and a 3- to 4-fold increase in seroma formation compared with non-ADM reconstruction techniques. Comparisons of aseptic and sterile ADMs in multiple studies have shown no significant difference in infection rates and equivocal findings for other specific complications such as seroma formation. Conclusions: Current evidence on the impact of processing techniques that improve ADM sterility on postoperative morbidity in implant breast reconstruction is unclear. Deficiencies of the available data highlight the need for well-designed, multicenter, randomized controlled studies that will aid in optimizing outcomes in implant-based breast reconstruction. PMID:27536502

  4. Deficiency in acellular cementum and periodontal attachment in bsp null mice.

    PubMed

    Foster, B L; Soenjaya, Y; Nociti, F H; Holm, E; Zerfas, P M; Wimer, H F; Holdsworth, D W; Aubin, J E; Hunter, G K; Goldberg, H A; Somerman, M J

    2013-02-01

    Bone sialoprotein (BSP) is an extracellular matrix protein found in mineralized tissues of the skeleton and dentition. BSP is multifunctional, affecting cell attachment and signaling through an RGD integrin-binding region, and acting as a positive regulator for mineral precipitation by nucleating hydroxyapatite crystals. BSP is present in cementum, the hard tissue covering the tooth root that anchors periodontal ligament (PDL) attachment. To test our hypothesis that BSP plays an important role in cementogenesis, we analyzed tooth development in a Bsp null ((-/-)) mouse model. Developmental analysis by histology, histochemistry, and SEM revealed a significant reduction in acellular cementum formation on Bsp (-/-) mouse molar and incisor roots, and the cementum deposited appeared hypomineralized. Structural defects in cementum-PDL interfaces in Bsp (-/-) mice caused PDL detachment, likely contributing to the high incidence of incisor malocclusion. Loss of BSP caused progressively disorganized PDL and significantly increased epithelial down-growth with aging. Bsp (-/-) mice displayed extensive root and alveolar bone resorption, mediated by increased RANKL and the presence of osteoclasts. Results collected here suggest that BSP plays a non-redundant role in acellular cementum formation, likely involved in initiating mineralization on the root surface. Through its importance to cementum integrity, BSP is essential for periodontal function. PMID:23183644

  5. The acellular matrix (ACM) for bladder tissue engineering: A quantitative magnetic resonance imaging study.

    PubMed

    Cheng, Hai-Ling Margaret; Loai, Yasir; Beaumont, Marine; Farhat, Walid A

    2010-08-01

    Bladder acellular matrices (ACMs) derived from natural tissue are gaining increasing attention for their role in tissue engineering and regeneration. Unlike conventional scaffolds based on biodegradable polymers or gels, ACMs possess native biomechanical and many acquired biologic properties. Efforts to optimize ACM-based scaffolds are ongoing and would be greatly assisted by a noninvasive means to characterize scaffold properties and monitor interaction with cells. MRI is well suited to this role, but research with MRI for scaffold characterization has been limited. This study presents initial results from quantitative MRI measurements for bladder ACM characterization and investigates the effects of incorporating hyaluronic acid, a natural biomaterial useful in tissue-engineering and regeneration. Measured MR relaxation times (T(1), T(2)) and diffusion coefficient were consistent with increased water uptake and glycosaminoglycan content observed on biochemistry in hyaluronic acid ACMs. Multicomponent MRI provided greater specificity, with diffusion data showing an acellular environment and T(2) components distinguishing the separate effects of increased glycosaminoglycans and hydration. These results suggest that quantitative MRI may provide useful information on matrix composition and structure, which is valuable in guiding further development using bladder ACMs for organ regeneration and in strategies involving the use of hyaluronic acid.

  6. Creation and implantation of acellular rat renal ECM-based scaffolds.

    PubMed

    Peloso, Andrea; Ferrario, Jacopo; Maiga, Benedetta; Benzoni, Ilaria; Bianco, Carolina; Citro, Antonio; Currao, Manuela; Malara, Alessandro; Gaspari, Annalisa; Balduini, Alessandra; Abelli, Massimo; Piemonti, Lorenzo; Dionigi, Paolo; Orlando, Giuseppe; Maestri, Marcello

    2015-01-01

    Kidney transplantation is the only potentially curative treatment for patient facing end-stage renal disease, and it is now routinely used. Its use is mainly limited by the supply of transplantable donor organs, which far exceeds the demand. Regenerative medicine and tissue engineering offer promising means for overcoming this shortage. In the present study, we developed and validated a protocol for producing acellular rat renal scaffolds. Left kidneys were removed from 26 male Lewis rats (weights: 250-350 g) and decellularized by means of aortic anterograde perfusion with ionic and anionic detergents (Triton X-100 1% and SDS 1%, respectively). 19 scaffolds thus obtained (and contralateral native kidneys as controls) were deeply characterized in order to evaluate the decellularization quality, the preservation of extracellular matrix components and resultant micro-angioarchitecture structure. The other 7 were transplanted into 7 recipient rats that had undergone unilateral nephrectomy. Recipients were sacrificed on post-transplantation day 7 and the scaffolds subjected to histologic studies. The dual-detergent protocol showed, with only 5 h of perfusion per organ, to obtain thoroughly decellularized renal scaffolds consisting almost exclusively of extracellular matrix. Finally the macro- and the microarchitecture of the renal parenchyma were well preserved, and the grafts were implanted with ease. Seven days after transplant, the scaffolds were morphologically intact although all vascular structures were obstructed with thrombi. Production and implantation of acellular rat renal scaffolds is a suitable platform for further studies on regenerative medicine and tissue engineering. PMID:26186418

  7. Sterile acellular dermal collagen as a treatment for rippling deformity of breast.

    PubMed

    Busse, Brittany; Orbay, Hakan; Sahar, David E

    2014-01-01

    Prosthetic implants are frequently used for breast augmentation and breast reconstruction following mastectomy. Unfortunately, long-term aesthetic results of prosthetic breast restoration may be hindered by complications such as rippling, capsular contracture, and implant malposition. The advent of use of acellular dermal matrices has greatly improved the outcomes of prosthetic breast reconstruction. We describe a case of rippling deformity of breast that was treated using an acellular dermal matrix product, AlloMax. The patient presented with visible rippling of bilateral prosthetic breast implants as well as significant asymmetry of the breasts after multiple excisional biopsies for right breast ductal carcinoma in situ. A 6 × 10 cm piece of AlloMax was placed on the medial aspect of each breast between the implant and the skin flap. Follow-up was performed at 1 week, 3 months, and 1 year following the procedure. The patient recovered well from the surgery and there were no complications. At her first postoperative follow-up the patient was extremely satisfied with the result. At her 3-month and 1-year follow-up she had no recurrence of her previous deformity and no new deformity. PMID:25610697

  8. Waning vaccine immunity in teenagers primed with whole cell and acellular pertussis vaccine: recent epidemiology.

    PubMed

    Sheridan, Sarah L; Frith, Katie; Snelling, Thomas L; Grimwood, Keith; McIntyre, Peter B; Lambert, Stephen B

    2014-09-01

    The recent epidemics of pertussis (whooping cough) in parts of the USA and Australia have led to the largest numbers of annual cases reported in over half a century. These epidemics demonstrated a new pattern, with particularly high rates of disease among pre-adolescents and early adolescents. These high rates of pertussis coincided with the first cohorts vaccinated with purely acellular pertussis vaccine, which replaced whole-cell pertussis (wP) vaccine in the later 1990s in the USA and Australia. Studies undertaken during these epidemics provide new evidence of more rapid waning of acellular pertussis-containing vaccines and longer-term protection from effective wP-containing vaccines. There is evidence that receiving wP as at least the first dose of pertussis-containing vaccine provides greater and more long-lived protection, irrespective of the nature of subsequent doses. This evidence will be reviewed together with the immunobiology associated with both vaccines, and the implications for pertussis control discussed. PMID:25093268

  9. Alternatives to HIST for acellular pertussis vaccines: progress and challenges in replacement

    PubMed Central

    Arciniega, J.; Wagner, L.; Prymula, R.; Sebo, P.; Isbrucker, R.; Descampe, B.; Chapsal, J.M.; Costanzo, A.; Hendriksen, C.; Hoonaker, M.; Nelson, S.; Lidster, K.; Casey, W.; Allen, D.

    2016-01-01

    The ‘International Workshop on Alternatives to the Murine Histamine Sensitization Test for Acellular Pertussis Vaccines: Progress and Challenges in the Replacement of HIST’ was held on 24 August 2014, in Prague, Czech Republic, as a satellite meeting to the 9 th World Congress on Alternatives and Animal Use in the Life Sciences. Participants discussed the progress and challenges associated with the development, validation, and implementation of in vitro assays as replacements for the histamine sensitisation test (HIST) for acellular pertussis vaccines. Discussions focused on the consistency approach, the necessary framework for regulatory acceptance of a harmonised method, and recent international efforts towards the development of in vitro assays to replace the HIST. Workshop participants agreed that acceptable alternatives to the HIST should be based on ADP ribosylation-mediated cell intoxication and therefore that the CHO cell clustering assay, which measures cell intoxication, should be further pursued and developed as a possible replacement for the HIST. Participants also agreed to continue ongoing multinational discussions involving national and international standardisation authorities to reach consensus and to organise collaborative studies in this context for assay characterisation and calibration of reference materials. PMID:27506225

  10. Interposition Porcine Acellular Dermal Matrix Xenograft Successful Alternative in Treatment for Massive Rotator Cuff

    PubMed Central

    Neumann, Julie; Zgonis, Miltiadis H.; Reay, Kathleen Dolores; Mayer, Stephanie W.; Boggess, Blake; Toth, Alison P.

    2016-01-01

    Objectives: Despite advances in the surgical techniques of rotator cuff repair (RCR), the management of massive rotator cuff tears in shoulders without glenohumeral arthritis poses a difficult problem for surgeons. Failure of massive rotator cuff repairs range from 20-90% at one to two years postoperatively using arthrography, ultrasound, or magnetic resonance imaging. Additionally, there are inconsistent outcomes reported with debridement alone of massive rotator cuff tears as well as limitations seen with other current methods of operative intervention including arthroplasty and tendon transfers. The purpose of this prospective, comparative study was to determine if the repair of massive rotator cuff tears using an interposition porcine acellular dermal matrix xenograft improves subjective function, pain, range of motion, and strength at greater than two years follow-up. To our knowledge, this is the largest prospective series reporting outcomes of using porcine acellular dermal matrix xenograft as an interposition graft. Methods: Thirty-seven patients (37 shoulders) with an average age of 66 years (range 51-80 years) were prospectively followed for 33 months (range 23-48) following massive RCR using porcine acellular dermal matrix interposition xenograft. Subjective outcomes were measured using the Visual Analog Scale (VAS) pain score (0-10, 0 = no pain), Modified American Shoulder and Elbow Score (M-ASES), and Short-Form12 (SF-12) scores. Preoperative and postoperative objective outcome measures included active range of motion and supraspinatus and infraspinatus manual muscle strength. Postoperative outcome measures included quantitative muscle strength using a dynamometer and static and dynamic ultrasonography to assess the integrity of the repair. Results: Average VAS pain score decreased from 4.5 to 1.1 (P<0.001). Average postoperative M-ASES was 89.23. Average postoperative SF-12 was 52.6. Mean forward flexion, external and internal rotation significantly

  11. Regulation of intramembranous absorption and amniotic fluid volume by constituents in fetal sheep urine.

    PubMed

    Anderson, Debra F; Jonker, Sonnet S; Louey, Samantha; Cheung, Cecilia Y; Brace, Robert A

    2013-09-01

    Our objective was to test the hypothesis that fetal urine contains a substance(s) that regulates amniotic fluid volume by altering the rate of intramembranous absorption of amniotic fluid. In late gestation ovine fetuses, amniotic fluid volumes, urine, and lung liquid production rates, swallowed volumes and intramembranous volume and solute absorption rates were measured over 2-day periods under control conditions and when urine was removed and continuously replaced at an equal rate with exogenous fluid. Intramembranous volume absorption rate decreased by 40% when urine was replaced with lactated Ringer solution or lactated Ringer solution diluted 50% with water. Amniotic fluid volume doubled under both conditions. Analysis of the intramembranous sodium and chloride fluxes suggests that the active but not passive component of intramembranous volume absorption was altered by urine replacement, whereas both active and passive components of solute fluxes were altered. We conclude that fetal urine contains an unidentified substance(s) that stimulates active intramembranous transport of amniotic fluid across the amnion into the underlying fetal vasculature and thereby functions as a regulator of amniotic fluid volume.

  12. Fetoscopic Amniotic Band Release in a Case of Chorioamniotic Separation: An Innovative New Technique.

    PubMed

    Belfort, Michael A; Whitehead, William E; Ball, Robert; Silver, Robert; Shamshirsaz, Alireza; Ruano, Rodrigo; Espinoza, Jimmy; Becker, Judith; Olutoye, Olutoyin; Hollier, Larry

    2016-04-01

    Introduction Fetoscopic release of amniotic bands has proved its life- and limb-saving potential. Rupture of the amnion and separation of chorion from the amnion and uterine wall can however preclude the standard fetoscopic approach to release the amniotic bands using a single port. Methods and Materials A 28-year-old G1P0 woman was referred to our unit at 19 weeks due to amniotic band syndrome involving the left ankle, the infrapatellar region of the right leg, and the umbilical cord. Of note, part of the fetus was seen outside the amniotic cavity by ultrasonography and the left ankle and foot were severely swollen. The patient underwent a laparotomy and fetoscopic release of the amniotic bands as well as partial amnionectomy using two uterine ports and CO2 as distention. Results The surgery and postoperative recovery course were uneventful. At 341/7 weeks the patient went into labor, which was augmented resulting vaginal delivery of a 2,460-g male infant. The infant was noted to have a shallow skin indentation on the left lower extremity near the ankle. The infant was discharged in excellent condition. Conclusion In those cases where release of an amniotic band is impossible due to membrane separation, surgery in a CO2-filled uterus offers an option. PMID:27298754

  13. Evolution and homology of the astragalus in early amniotes: new fossils, new perspectives.

    PubMed

    O'Keefe, F Robin; Sidor, Christian A; Larsson, Hans C E; Maga, Abdoudaye; Ide, Oumarou

    2006-04-01

    The reorganization of the ankle in basal amniotes has long been considered a key innovation allowing the evolution of more terrestrial and cursorial behavior. Understanding how this key innovation arose is a complex problem that largely concerns the homologizing of the amniote astragalus with the various ossifications in the anamniote tarsus. Over the last century, several hypotheses have been advanced homologizing the amniote astragalus with the many ossifications in the ankle of amphibian-grade tetrapods. There is an emerging consensus that the amniote astragalus is a complex structure emerging via the co-ossification of several originally separate elements, but the identities of these elements remain unclear. Here we present new fossil evidence bearing on this contentious question. A poorly ossified, juvenile astragalus of the large captorhinid Moradisaurus grandis shows clear evidence of four ossification centers, rather than of three centers or one center as posited in previous models of astragalus homology. Comparative material of the captorhinid Captorhinikos chozaensis is also interpretable as demonstrating four ossification centers. A new, four-center model for the homology of the amniote astragalus is advanced, and is discussed in the context of the phylogeny of the Captorhinidae in an attempt to identify the developmental transitions responsible for the observed pattern of ossification within this clade. Lastly, the broader implications for amniote phylogeny are discussed, concluding that the neomorphic pattern of astragalus ossification seen in all extant reptiles (including turtles) arose within the clade Diapsida. PMID:16421891

  14. Intraoperative Fluorescein Staining of Cryopreserved Amniotic Membrane Grafts to Improve Visualization During and After Pterygium Surgery: A Novel Technique

    PubMed Central

    Martinez, J. Alberto; Korchak, Michael; Cremers, Sandra L.

    2016-01-01

    Purpose: To describe a new method of enhancing the visualization of amniotic membrane grafts with fluorescein staining during pterygium surgery. Methods: Pterygium excision surgery using intraoperatively stained cryopreserved amniotic membranes was performed on 346 eyes. A sterile 0.6 mg sodium fluorescein strip was placed directly onto the amniotic membrane in the manufacturer's original packaging, and the stained allograft was then transplanted onto the planned site. Staining intensities, at 3, 5, and 10 minutes of dye immersion, were compared. Immediate postoperative pain rating (scale 0–10), visibility of the fluorescein-stained amniotic membrane graft, and conjunctival autograft and amniotic membrane graft elevation, dehiscence, retraction, or displacement were recorded. The recurrence rate of the study population was compared with that of a previous cohort of 121 patients who underwent pterygium excision with conjunctival autograft without stained amniotic membrane. Results: Direct contact of the fluorescein strip on the amniotic membrane at 3, 5, and 10 minutes showed no differences in subjective staining intensity. Fluorescein-stained amniotic membrane was easily detected on the ocular surface during and 24 hours after pterygium surgery. The average immediate postoperative pain rating was 0.8 ± 1.8. No intraoperative complications or postoperative amniotic membrane graft dehiscence, retraction, or displacement occurred. The recurrence rate using fluorescein-stained amniotic membrane (3 patients, 0.9%, mean follow-up time 31.8 ± 18.6 weeks) did not differ from that of the previous cohort without the stained amniotic membrane (2.5%; χ2(1) = 1.837, P = 0.183). Conclusions: Fluorescein strip staining of the amniotic membrane is a novel and safe intraoperative method to enhance visualization and handling of the graft during and after ocular surgeries. PMID:26751995

  15. The Clinical Significance of Eosinophils in the Amniotic Fluid in Preterm Labor

    PubMed Central

    ROMERO, ROBERTO; KUSANOVIC, JUAN PEDRO; GOMEZ, RICARDO; LAMONT, RONALD; BYTAUTIENE, EGLE; GARFIELD, ROBERT E.; MITTAL, POOJA; HASSAN, SONIA S.; YEO, LAMI

    2012-01-01

    Objective White blood cells are not traditionally considered to be normally present in amniotic fluid. This study was conducted after the observation that a patient with preterm labor and intact membranes had eosinophils as a predominant cell in the amniotic fluid, and had an episode of asthma during the index pregnancy. The goal of this study was to determine whether women presenting with preterm labor with eosinophils in the amniotic fluid had a different outcome than those without eosinophils as the predominant white blood cell in the amniotic cavity. Methods This retrospective case-control study included women who presented with preterm labor and intact membranes between 24 and 34 weeks of gestation. Patients underwent an amniocentesis shortly after admission for the assessment of the microbiologic status of the amniotic cavity and/or fetal lung maturity. Amniotic fluid was cultured for aerobic and anaerobic bacteria as well as genital mycoplasmas. Cytologic studies included amniotic fluid white blood cell count and differential, which was performed on cytocentrifuged specimens. Patients with microbial invasion of the amniotic cavity and/or a white blood cell count >20 cells/mm3 were excluded from the study. Cases were defined as women in whom the differential contained >20% of eosinophils. Controls were selected among women with an amniotic fluid eosinophil count ≤20% and matched for gestational age at amniocentesis. The analysis was conducted with non-parametric statistics. Results The study population consisted of 10 cases and 50 controls. Gestational age and cervical dilatation at admission were similar in both groups. Cases had a lower gestational age at delivery than controls [34.6 weeks, inter-quartile range (IQR) 32–37.3 weeks vs. 38.0 weeks, IQR 35–40 weeks, respectively; p=0.018]. The prevalence of preterm delivery ≤35 weeks was higher among patients who had >20% eosinophils than in the control group [50% (5/10) vs. 18% (9/50), respectively; p

  16. Efficient Direct Reprogramming of Mature Amniotic Cells into Endothelial Cells by ETS Factors and TGFβ Suppression

    PubMed Central

    Ginsberg, Michael; James, Daylon; Ding, Bi-Sen; Nolan, Daniel; Geng, Fuqiang; Butler, Jason M; Schachterle, William; Pulijaal, Venkat R; Mathew, Susan; Chasen, Stephen T; Xiang, Jenny; Rosenwaks, Zev; Shido, Koji; Elemento, Olivier; Rabbany, Sina Y; Rafii, Shahin

    2012-01-01

    ETS transcription factors ETV2, FLI1 and ERG1 specify pluripotent stem cells into endothelial cells (ECs). However, these ECs are unstable and drift towards non-vascular cell fates. We show that human mid-gestation c-Kit− lineage-committed amniotic cells (ACs) can be readily reprogrammed into induced vascular endothelial cells (iVECs). Transient ETV2 expression in ACs generated proliferative but immature iVECs, while co-expression with FLI1/ERG1 endowed iVECs with a vascular repertoire and morphology matching mature stable ECs. Brief TGFβ-inhibition functionalized VEGFR2 signaling, augmenting specification of ACs to iVECs. Genome-wide transcriptional analyses showed that iVECs are similar to adult ECs in which vascular-specific genes are turned on and non-vascular genes are silenced. Functionally, iVECs form long-lasting patent vasculature in Matrigel plugs and regenerating livers. Thus, short-term ETV2 expression and TGFβ-inhibition along with constitutive ERG1/FLI1 co-expression reprogram mature ACs into durable and functional iVECs with clinical-scale expansion potential. Public banking of HLA-typed iVECs would establish a vascular inventory for treatment of genetically diverse disorders. PMID:23084400

  17. Long-term immunomodulatory effect of amniotic stem cells in an Alzheimer's disease model.

    PubMed

    Kim, Kyung-Sul; Kim, Hyun Sook; Park, Ji-Min; Kim, Han Wool; Park, Mi-Kyung; Lee, Hyun-Seob; Lim, Dae Seog; Lee, Tae Hee; Chopp, Michael; Moon, Jisook

    2013-10-01

    Amyloid beta (Aβ) plays a major role in Alzheimer's disease (AD), and neuroinflammatory processes mediated by Aβ plaque-induced microglial cells and astrocytes contribute to AD pathogenesis. The present study examined human placenta amniotic membrane-derived mesenchymal stem cells (AMSCs), which have potent immunomodulatory and paracrine effects in a Tg2576 (APPswe) transgenic mouse model of AD. AMSCs secreted high levels of transforming growth factor-β under in vitro inflammatory environment conditions. Six weeks after the intravenous injection of AMSCs, APPswe mice showed evidence of improved spatial learning, which significantly correlated with the observation of fewer Aβ plaques in brain. The number of ED1-positive phagocytic microglial cells associated with Aβ plaques was higher in AMSC-injected mice than in phosphate-buffered saline-injected mice, and the level of Aβ-degrading enzymes (matrix metallopeptidase-9 and insulin-degrading enzyme) was also significantly higher. Furthermore, the level of proinflammatory cytokines, interleukin-1 and tumor necrosis factor-α, was lower and that of anti-inflammatory cytokines, interleukin-10 and transforming growth factor-β, was higher in AMSC-injected mice than phosphate-buffered saline-injected mice. These effects lasted until 12 weeks after AMSC injection. Taken together, these results collectively suggest that injection of AMSCs might show significant long-lasting improvement in AD pathology and memory function via immunomodulatory and paracrine mechanisms.

  18. Amniotic fluid iodine concentrations do not vary in pregnant women with varying iodine intake.

    PubMed

    García-Fuentes, Eduardo; Gallo, Manuel; García, Laureano; Prieto, Stephanie; Alcaide-Torres, Javier; Santiago, Piedad; Velasco, Inés; Soriguer, Federico

    2008-06-01

    Iodine deficiency is an important clinical and public health problem. Its prevention begins with an adequate intake of iodine during pregnancy. International agencies recommend at least 200 microg iodine per d for pregnant women. We assessed whether iodine concentrations in the amniotic fluid of healthy pregnant women are independent of iodine intake. This cross-sectional, non-interventional study included 365 consecutive women who underwent amniocentesis to determine the fetal karyotype. The amniocentesis was performed with abdominal antisepsis using chlorhexidine. The iodine concentration was measured in urine and amniotic fluid. The study variables were the intake of iodized salt and multivitamin supplements or the prescription of a KI supplement. The mean level of urinary iodine was 139.0 (SD 94.5) microg/l and of amniotic fluid 15.81 (SD 7.09) microg/l. The women who consumed iodized salt and those who took a KI supplement had significantly higher levels of urinary iodine than those who did not (P = 0.01 and P = 0.004, respectively). The urinary iodine levels were not significantly different in the women who took a multivitamin supplement compared with those who did not take this supplement, independently of iodine concentration or multivitamin supplement. The concentrations of iodine in the amniotic fluid were similar, independent of the dietary iodine intake. Urine and amniotic fluid iodine concentrations were weakly correlated, although the amniotic fluid values were no higher in those women taking a KI supplement. KI prescription at recommended doses increases the iodine levels in the mother without influencing the iodine levels in the amniotic fluid.

  19. Effective management of major lower extremity wounds using an acellular regenerative tissue matrix: a pilot study.

    PubMed

    Brigido, Stephen A; Boc, Steven F; Lopez, Ramon C

    2004-01-01

    Wound healing is a significant problem in orthopedics. Graftjacket tissue matrix (Wright Medical Technology, Inc, Arlington, Tenn), a novel acellular regenerative tissue matrix, has been designed to aid wound closure. A prospective, randomized study was initiated to determine the efficacy of this tissue product in wound repair compared with conventional treatment. Lower extremity wounds are refractile to healing in patients with diabetes mellitus. Therefore, researchers used diabetic foot ulcers to evaluate the efficacy of GraftJacket tissue matrix in wound repair. Only a single administration of the tissue matrix was required. After 1 month of treatment, preliminary results demonstrate that this novel tissue matrix promotes faster healing at a statistically significant rate over conventional treatment. Because wounds in this series of patients are deep and circulation around the wound is poor, the preliminary results suggest that this tissue matrix will be applicable to other types of orthopedic wounds.

  20. Gravity related behavior of the acellular slime mold Physarum polycephalum (7-IML-1)

    NASA Technical Reports Server (NTRS)

    Block, I.

    1992-01-01

    The objective of the experiment is to investigate the effect of near weightlessness on a single cell. The test object is the acellular slime mold Physarum polycephalum. This cell is composed of a network of protoplastic strands which perform rhythmic contractions in the minute range. These contractions of the strands' ectoplastic walls generate the force to drive the vigorous shuttle streaming of fluid protoplasm inside the strands (hydrostatic pressure flow). A net transport of protoplasm in one direction determines the direction of the cell's locomotion itself. In this way, gravity modifies the contraction rhythm of the strands, the streaming velocity of protoplasm in the strands, and the direction of locomotion of the whole slime mold (geotaxis). The other parts of this experiment will address the major question of how this cell, which does not possess any specialized gravireceptors, gets the information about the direction of the gravity vector. Details of the experimental setup are given.

  1. Tissue-engineered acellular small diameter long-bypass grafts with neointima-inducing activity.

    PubMed

    Mahara, Atsushi; Somekawa, Shota; Kobayashi, Naoki; Hirano, Yoshiaki; Kimura, Yoshiharu; Fujisato, Toshiya; Yamaoka, Tetsuji

    2015-07-01

    Researchers have attempted to develop efficient antithrombogenic surfaces, and yet small-caliber artificial vascular grafts are still unavailable. Here, we demonstrate the excellent patency of tissue-engineered small-caliber long-bypass grafts measuring 20-30 cm in length and having a 2-mm inner diameter. The inner surface of an acellular ostrich carotid artery was modified with a novel heterobifunctional peptide composed of a collagen-binding region and the integrin α4β1 ligand, REDV. Six grafts were transplanted in the femoral-femoral artery crossover bypass method. Animals were observed for 20 days and received no anticoagulant medication. No thrombogenesis was observed on the luminal surface and five cases were patent. In contrast, all unmodified grafts became occluded, and severe thrombosis was observed. The vascular grafts reported here are the first successful demonstrations of short-term patency at clinically applicable sizes. PMID:25941782

  2. Tetanus-diphtheria-acellular pertussis vaccination of adults in the USA.

    PubMed

    Gidengil, Courtney A; Sandora, Thomas J; Lee, Grace M

    2008-07-01

    Pertussis is an important cause of morbidity and mortality, and its incidence has been increasing in adolescents and adults over the past two decades. Waning immunity in adolescents and adults may be partially responsible. Adults can suffer significant illness from pertussis and its complications, such as pneumonia, rib fractures and syncope. Moreover, adults serve as a source of disease for infants, who are more vulnerable to severe complications and even death. The economic burden of pertussis is substantial, in terms of both medical and nonmedical costs. Fortunately, the burden of pertussis disease can now be safely and effectively reduced by vaccinating adults with tetanus-diphtheria-acellular pertussis (Tdap) vaccine. Further research is needed to elucidate the role of vaccination in pregnant women and those over 65 years of age, and also to determine whether further booster doses of Tdap are needed.

  3. Rat-derived amniotic epithelial cells differentiate into mature hepatocytes in vivo with no evidence of cell fusion.

    PubMed

    Marongiu, Michela; Serra, Maria Paola; Contini, Antonella; Sini, Marcella; Strom, Stephen C; Laconi, Ezio; Marongiu, Fabio

    2015-06-15

    Amniotic epithelial cells (AEC) derived from human placenta represent a useful and noncontroversial source for liver-based regenerative medicine. Previous studies suggested that human- and rat-derived AEC differentiate into hepatocyte-like cells upon transplantation. In the retrorsine (RS) model of liver repopulation, clusters of donor-derived cells engrafted in the recipient liver and, importantly, showed characteristics of mature hepatocytes. The aim of the current study was to investigate the possible involvement of cell fusion in the emergence of hepatocyte clusters displaying a donor-specific phenotype. To this end, 4-week-old GFP(+)/DPP-IV(-) rats were treated with RS and then transplanted with undifferentiated AEC isolated from the placenta of DPP-IV(+) pregnant rats at 16-19 days of gestational age. Results indicated that clusters of donor-derived cells were dipeptidyl peptidase type IV (DPP-IV) positive, but did not express the green fluorescent protein (GFP), suggesting that rat amniotic epithelial cells (rAEC) did not fuse within the host parenchyma, as no colocalization of the two tags was observed. Moreover, rAEC-derived clusters expressed markers of mature hepatocytes (eg, albumin, cytochrome P450), but were negative for the expression of biliary/progenitor markers (eg, epithelial cell adhesion molecule [EpCAM]) and did not express the marker of preneoplastic hepatic nodules glutathione S-transferase P (GST-P). These results extend our previous findings on the potential of AEC to differentiate into mature hepatocytes and suggest that this process can occur in the absence of cell fusion with host-derived cells. These studies support the hypothesis that amnion-derived epithelial cells can be an effective cell source for the correction of liver disease.

  4. Rat-Derived Amniotic Epithelial Cells Differentiate into Mature Hepatocytes In Vivo with No Evidence of Cell Fusion

    PubMed Central

    Marongiu, Michela; Serra, Maria Paola; Contini, Antonella; Sini, Marcella; Strom, Stephen C.; Laconi, Ezio

    2015-01-01

    Amniotic epithelial cells (AEC) derived from human placenta represent a useful and noncontroversial source for liver-based regenerative medicine. Previous studies suggested that human- and rat-derived AEC differentiate into hepatocyte-like cells upon transplantation. In the retrorsine (RS) model of liver repopulation, clusters of donor-derived cells engrafted in the recipient liver and, importantly, showed characteristics of mature hepatocytes. The aim of the current study was to investigate the possible involvement of cell fusion in the emergence of hepatocyte clusters displaying a donor-specific phenotype. To this end, 4-week-old GFP+/DPP-IV− rats were treated with RS and then transplanted with undifferentiated AEC isolated from the placenta of DPP-IV+ pregnant rats at 16–19 days of gestational age. Results indicated that clusters of donor-derived cells were dipeptidyl peptidase type IV (DPP-IV) positive, but did not express the green fluorescent protein (GFP), suggesting that rat amniotic epithelial cells (rAEC) did not fuse within the host parenchyma, as no colocalization of the two tags was observed. Moreover, rAEC-derived clusters expressed markers of mature hepatocytes (eg, albumin, cytochrome P450), but were negative for the expression of biliary/progenitor markers (eg, epithelial cell adhesion molecule [EpCAM]) and did not express the marker of preneoplastic hepatic nodules glutathione S-transferase P (GST-P). These results extend our previous findings on the potential of AEC to differentiate into mature hepatocytes and suggest that this process can occur in the absence of cell fusion with host-derived cells. These studies support the hypothesis that amnion-derived epithelial cells can be an effective cell source for the correction of liver disease. PMID:25647334

  5. Immune Responses to Pertussis Antigens in Infants and Toddlers after Immunization with Multicomponent Acellular Pertussis Vaccine

    PubMed Central

    Wang, Li; Chen, Qingxia

    2014-01-01

    Given the resurgence of pertussis despite high rates of vaccination with the diphtheria-tetanus-acellular pertussis (DTaP) vaccine, a better understanding of vaccine-induced immune responses to Bordetella pertussis is needed. We investigated the antibody, cell-mediated, and cytokine responses to B. pertussis antigens in children who received the primary vaccination series (at 2, 4, and 6 months) and first booster vaccination (at 15 to 18 months) with 5-component acellular pertussis (aP) vaccine. The majority of subjects demonstrated a 4-fold increase in antibody titer to all four pertussis antigens (pertussis toxin [PT], pertactin [PRN], filamentous hemagglutinin [FHA], and fimbriae [FIM]) following the primary series and booster vaccination. Following the primary vaccine series, the majority of subjects (52 to 67%) mounted a positive T cell proliferative response (stimulation index of ≥3) to the PT and PRN antigens, while few subjects (7 to 12%) mounted positive proliferative responses to FHA and FIM. One month after booster vaccination (age 16 to 19 months), our study revealed significant increase in gamma interferon (IFN-γ) production in response to the PT and FIM antigens, a significant increase in IL-2 production with the PT, FHA, and PRN antigens, and a lack of significant interleukin-4 (IL-4) secretion with any of the antigens. While previous reports documented a mixed Th1/Th2 or Th2-skewed response to DTaP vaccine in children, our data suggest that following the first DTaP booster, children aged 16 to 19 months have a cytokine profile consistent with a Th1 response, which is known to be essential for clearance of pertussis infection. To better define aP-induced immune responses following the booster vaccine, further studies are needed to assess cytokine responses pre- and postbooster in DTaP recipients. PMID:25253666

  6. Is Sterile Better Than Aseptic? Comparing the Microbiology of Acellular Dermal Matrices

    PubMed Central

    Klein, Gabriel M.; Nasser, Ahmed E.; Phillips, Brett T.; Gersch, Robert P.; Fourman, Mitchell S.; Lilo, Sarit E.; Fritz, Jason R.; Khan, Sami U.; Dagum, Alexander B.

    2016-01-01

    Introduction: Postoperative infections are a major complication associated with tissue-expander-based breast reconstruction. The use of acellular dermal matrix (ADM) in this surgery has been identified as a potential reservoir of infection, prompting the development of sterile ADM. Although aseptic and sterile ADMs have been investigated, no study has focused on the occurrence and clinical outcome of bacterial colonization before implantation. Methods: Samples of aseptic AlloDerm, sterile Ready-To-Use AlloDerm, and AlloMax were taken before implantation. These samples were incubated in Tryptic soy broth overnight before being streaked on Trypticase soy agar, MacConkey agar, and 5% blood agar plates for culture and incubated for 48 hours. Culture results were cross-referenced with patient outcomes for 1 year postoperatively. Results: A total of 92 samples of ADM were collected from 63 patients. There were 15 cases of postoperative surgical site infection (16.3%). Only 1 sample of ADM (AlloMax) showed growth of Escherichia coli, which was likely a result of contamination. That patient did not develop any infectious sequelae. Patient outcomes showed no difference in the incidence of seroma or infection between sterile and aseptic ADMs. Conclusions: This study evaluates the microbiology of acellular dermal matrices before use in breast reconstruction. No difference was found in the preoperative bacterial load of either aseptic or sterile ADM. No significant difference was noted in infection or seroma formation. Given these results, we believe aseptic processing used on ADMs is equivalent to sterile processing in our patient cohort in terms of clinical infection and seroma occurrence postoperatively. PMID:27482500

  7. The use of dry amniotic membrane in pterygium surgery

    PubMed Central

    Noureddin, Gelareh S; Yeung, Sonia N

    2016-01-01

    Pterygium is a fibrovascular growth of the bulbar conjunctiva that crosses the limbus and extends over the peripheral cornea, in some cases resulting in significant visual morbidity. When treatment is indicated, surgery is necessary, and several management options exist. These include excision, conjunctival autografting, and the use of adjuvant therapies. This paper reviews the incidence and prevalence of pterygia and also describes the various techniques currently used to treat this condition. These management options are compared to the use of dry amniotic membrane grafting (AMG), specifically with regard to recurrence rates, time to recurrence, safety and tolerability, as well as patient factors including cosmesis and quality of life. AMG has been used in the treatment of ocular surface disease due to a variety of benefits, including its anti-inflammatory properties, as well as its ability to promote epithelial growth and suppress transforming growth factor-β signaling and fibroblast proliferation. However, rates of recurrence for AMG following pterygium excision still surpass other commonly used techniques, including conjunctival and limbal autografting. Nevertheless, there are circumstances in which AMG may be most beneficial to the patient, such as when preexisting conjunctival scarring is present, when the conjunctiva must be spared for future glaucoma filtering surgery, or in cases of large or double-headed pterygia. Therefore, surgeons should be prepared to offer this procedure as an option to their patients for the treatment of pterygia. PMID:27143848

  8. Satellited 4q identified in amniotic fluid cells

    SciTech Connect

    Miller, I.; Hsieh, C.L.; Songster, G.

    1995-01-16

    Extra material was identified on the distal long arm of a chromosome 4 in an amniotic fluid specimen sampled at 16.6 weeks of gestational age. There was no visible loss of material from chromosome 4, and no evidence for a balanced rearrangement. The primary counseling issue in this case was advanced maternal age. Ultrasound findings were normal, and family history was unremarkable. The identical 4qs chromosome was observed in cells from a paternal peripheral blood specimen and appeared to be an unbalanced rearrangement. This extra material was NOR positive in lymphocytes from the father, but was negative in the fetal amniocytes. Father`s relatives were studied to verify the familial origin of this anomaly. In situ hybridization with both exon and intron sequences of ribosomal DNA demonstrated that ribosomal DNA is present at the terminus of the 4qs chromosome in the fetus, father, and paternal grandmother. This satellited 4q might have been derived from a translocation event that resulted in very little or no loss from the 4q and no specific phenotype. This derivative chromosome 4 has been inherited through at least 3 generations of phenotypically normal individuals. 8 refs., 3 figs.

  9. [Complicated but successful resuscitation after amniotic fluid embolism].

    PubMed

    Bouman, E A; Gutiérrez y Leon, J A; van der Salm, P C; Christiaens, G C; Bruinse, H W; Broeders, I A

    2001-04-14

    A 33-year-old woman, gravida IV, para III with unexplained polyhydramnios was admitted to give birth at 29 weeks of pregnancy. Directly after the spontaneous breaking of the membranes, asystolia occurred. Following emergency resuscitation the sinus rhythm returned. Upon the relaparotomy due to a large filling requirement and increasing abdomen size, 'crush' lesions to the spleen and liver were visible; following this a splenectomy was carried out and tampons applied to the liver. After seven months the patient had slight residual symptoms; three weeks after his birth her son was transferred in good condition to another hospital. Amniotic fluid embolism is a rare complication of pregnancy with often serious complications for mother and child. The diagnosis is based on the clinical symptoms of cardiac arrest or sudden profound shock, acute respiratory failure, and/or disseminated intravascular coagulation, occurring in most cases during or soon after delivery, in the absence of an alternative cause (in particular primary cardiopulmonary causes). If the clinical picture deviates from the expected post-resuscitation course alternative diagnoses or resuscitation injuries must be considered.

  10. The Anolis Lizard Genome: An Amniote Genome without Isochores?

    PubMed Central

    Costantini, Maria; Greif, Gonzalo; Alvarez-Valin, Fernando; Bernardi, Giorgio

    2016-01-01

    Two articles published 5 years ago concluded that the genome of the lizard Anolis carolinensis is an amniote genome without isochores. This claim was apparently contradicting previous results on the general presence of an isochore organization in all vertebrate genomes tested (including Anolis). In this investigation, we demonstrate that the Anolis genome is indeed heterogeneous in base composition, since its macrochromosomes comprise isochores mainly from the L2 and H1 families (a moderately GC-poor and a moderately GC-rich family, respectively), and since the majority of the sequenced microchromosomes consists of H1 isochores. These families are associated with different features of genome structure, including gene density and compositional correlations (e.g., GC3 vs flanking sequence GC and intron GC), as in the case of mammalian and avian genomes. Moreover, the assembled Anolis chromosomes have an enormous number of gaps, which could be due to sequencing problems in GC-rich regions of the genome. In conclusion, the Anolis genome is no exception to the general rule of an isochore organization in the genomes of vertebrates (and other eukaryotes). PMID:26992416

  11. Acellularization-Induced Changes in Tensile Properties Are Organ Specific - An In-Vitro Mechanical and Structural Analysis of Porcine Soft Tissues

    PubMed Central

    Aust, Gabriela; Boldt, Andreas; Fritsch, Sebastian; Keil, Isabel; Koch, Holger; Möbius, Robert; Scheidt, Holger A.; Wagner, Martin F. X.; Hammer, Niels

    2016-01-01

    Introduction Though xenogeneic acellular scaffolds are frequently used for surgical reconstruction, knowledge of their mechanical properties is lacking. This study compared the mechanical, histological and ultrastructural properties of various native and acellular specimens. Materials and Methods Porcine esophagi, ureters and skin were tested mechanically in a native or acellular condition, focusing on the elastic modulus, ultimate tensile stress and maximum strain. The testing protocol for soft tissues was standardized, including the adaption of the tissue’s water content and partial plastination to minimize material slippage as well as templates for normed sample dimensions and precise cross-section measurements. The native and acellular tissues were compared at the microscopic and ultrastructural level with a focus on type I collagens. Results Increased elastic modulus and ultimate tensile stress values were quantified in acellular esophagi and ureters compared to the native condition. In contrast, these values were strongly decreased in the skin after acellularization. Acellularization-related decreases in maximum strain were found in all tissues. Type I collagens were well-preserved in these samples; however, clotting and a loss of cross-linking type I collagens was observed ultrastructurally. Elastins and fibronectins were preserved in the esophagi and ureters. A loss of the epidermal layer and decreased fibronectin content was present in the skin. Discussion Acellularization induces changes in the tensile properties of soft tissues. Some of these changes appear to be organ specific. Loss of cross-linking type I collagen may indicate increased mechanical strength due to decreasing transverse forces acting upon the scaffolds, whereas fibronectin loss may be related to decreased load-bearing capacity. Potentially, the alterations in tissue mechanics are linked to organ function and to the interplay of cells and the extracellular matrix, which is different in

  12. [Prenatal evaluation of fetal lung maturity by determination of phosphatidylglycerol (PG) concentration in amniotic fluid].

    PubMed

    Krasomski, G; Sałacińska, B; Broniarczyk, D; Swiatkowska, E

    2001-09-01

    Phosphatidylglycerol (PG) is considered to be the most important phospholipid of lung surfactant, responsible for its biological activity. The aim of the study was to investigate the predictive value of phosphatidylglycerol (PG) concentration in amniotic fluid in prediction of neonatal lung maturity. The study was carried out on 180 pregnant women, chosen by random selection, hospitalized in Polish Mother's Health Centre Hospital in the period from 15.06.1994 to 31.12.1995. 223 samples of amniotic fluid were tested--phosphatidylglycerol (PG) concentration was assayed by immunological test Amnio Stat FLM. In the study value PG < or = 0.2 microgram/ml was found in 110 (49.3%) samples of amniotic fluid, PG = 0.5 microgram/ml in 57 (25.6%) and PG > or = 2.0 micrograms/ml in 56 (25.1%). In our research predictive value of PG in relation to clinical lung maturity of neonates was verified. It turned out that PG concentration in amniotic fluid < or = 0.2 microgram/ml indicates a possibility of RDS occurring in neonates born before 72 hours of performed determination. Concentration PG in amniotic fluid PG > or = 2.0 micrograms/ml corresponds to complete clinical lung maturity of neonates with predictive value 98%. PMID:11757479

  13. Matrix metalloproteinase-2 is elevated in midtrimester amniotic fluid prior to the development of preeclampsia

    PubMed Central

    Lavee, Michal; Goldman, Shlomit; Daniel-Spiegel, Etty; Shalev, Eliezer

    2009-01-01

    Objective To evaluate levels of matrix metalloproteinases (MMP) and their inhibitors (TIMP) in second trimester amniotic fluid of women with hypertensive disorders compared to normotensive women. Study Design Amniotic fluid was obtained from 133 women undergoing genetic second trimester amniocentesis. Zymography was performed for MMP characterization and an MMP-2 ELISA kit was used to determine MMP-2 levels. TIMP-2 expression was evaluated using western blot. Results Mean amniotic fluid MMP-2 and TIMP-2 levels were significantly higher in women who developed a hypertensive disorder compared to normotensive women (P < 0.0004 and P < 0.01, respectively). When subdivided into subgroups, amniotic fluid from women who eventually developed preeclampsia or superimposed preeclampsia showed significantly higher MMP-2 levels than normotensive women (P < 0.05). However, no statistical difference in MMP-2 levels was found between patients with gestational hypertension and normotensive patients. Conclusion Higher amniotic fluid MMP-2 and TIMP-2 levels are found in women who eventually develop preeclampsia. PMID:19698156

  14. Lamotrigine in pregnancy - therapeutic drug monitoring in maternal blood, amniotic fluid, and cord blood.

    PubMed

    Paulzen, Michael; Lammertz, Sarah E; Veselinovic, Tanja; Goecke, Tamme W; Hiemke, Christoph; Gründer, Gerhard

    2015-09-01

    This study is the first to measure and correlate lamotrigine concentrations in maternal blood, amniotic fluid, and umbilical cord blood and account for distribution of the drug between these three compartments. Concentrations of lamotrigine were measured in six mother-infant pairs at the time of delivery. Daily doses of lamotrigine ranged between 200 and 650 mg. Daily doses were correlated with maternal serum and umbilical cord blood concentrations, and serum levels were correlated with levels in amniotic fluid. Lamotrigine levels in serum correlated strongly with the lamotrigine levels in amniotic fluid (r=+0.986, P<0.001) and cord blood (r=+0.928, P=0.008). The penetration ratio into amniotic fluid was in a range between 0.31 and 0.75 (mean 0.58, SD 0.17); the penetration ratio into the fetal circulation, calculated on the basis of umbilical cord blood levels, was found to be in a range between 0.48 and 1.27 (mean 0.81, SD 0.28). Lamotrigine concentrations in amniotic fluid provided evidence that maternally administered lamotrigine is accessible to the fetus in a manner not previously appreciated. Furthermore, the penetration ratio into umbilical cord blood calculated here is in line with the largest study carried out so far to explore transplacental transfer.

  15. Complement Split Products in Amniotic Fluid in Pregnancies Subsequently Developing Early-Onset Preeclampsia

    PubMed Central

    Banadakoppa, Manu; Vidaeff, Alex C.; Yallampalli, Uma; Ramin, Susan M.; Belfort, Michael A.; Yallampalli, Chandra

    2015-01-01

    Objective. To determine the second-trimester amniotic fluid concentrations of complement split products in pregnancies subsequently affected by early-onset preeclampsia. Study Design. Cohort of 731 women with singleton pregnancies undergoing second-trimester genetic amniocentesis followed up to delivery and analyzed as a nested case-control study. Cases of preeclampsia developing before 34 weeks' gestation (n = 15) were compared with 47 uncomplicated term controls. Amniotic fluid collected at amniocentesis was tested for complement split products Bb, C4a, C3a, and C5a. Results. Women who developed early-onset preeclampsia as compared with the term pregnant controls had significantly higher (P = 0.04) median amniotic fluid C3a levels (318.7 ng/mL versus 254.5 ng/mL). Median amniotic fluid Bb levels were also significantly higher (P = 0.03) in preeclamptic women than in normal pregnant women (1127 ng/mL versus 749 ng/mL). Median levels of C4a and C5a were not significantly different between the groups. Conclusion. Our data suggest that complement activation in early pregnancy is associated with early-onset preeclampsia. We believe this to be the first prospective study to link complement activation in amniotic fluid in early pregnancy and later development of preeclampsia. Our findings provide evidence that immune dysregulation may precede the clinical manifestations of preeclampsia and that the alternative complement pathway is principally involved. PMID:26556948

  16. Amniotic membrane graft for primary pterygium: comparison with conjunctival autograft and topical mitomycin C treatment

    PubMed Central

    Ma, D. H.; See, L.; Liau, S.; Tsai, R. J.

    2000-01-01

    AIM—To study the efficacy and safety of amniotic membrane graft as an adjunctive therapy after removal of primary pterygium, and to compare the clinical outcome with conjunctival autograft and topical mitomycin C.
METHODS—80 eyes of 71 patients with primary pterygia were treated with excision followed by amniotic membrane graft. The result was compared retrospectively with 56 eyes of 50 patients receiving conjunctival autograft, and 54 eyes of 46 patients receiving topical mitomycin C. Patients were followed for at least 6 months, and the averaged follow up periods for the three groups were 13.8, 22.8, and 18.4 months, respectively.
RESULTS—There were three recurrences (3.8%) in the amniotic membrane graft group, three recurrences (5.4%) in the conjunctival autograft group, and two recurrences (3.7%) in the topical mitomycin C group. There was no significant difference in recurrence rate among the three groups (p = 0.879). No major complications occurred in the amniotic membrane graft group or the conjunctival autograft group. One case of infectious scleritis due to scleral ischaemia occurred in the topical mitomycin C group.
CONCLUSION—This study showed that amniotic membrane graft was as effective as conjunctival autograft and mitomycin C in preventing pterygium recurrence, and can be considered as a preferred grafting procedure for primary pterygium.

 PMID:10966947

  17. Amniotic fluid 'sludge' detected in patients with subchorionic hematoma: a report of two cases.

    PubMed

    Tskitishvili, E; Tomimatsu, T; Kanagawa, T; Sawada, K; Kinugasa, Y; Mimura, K; Kimura, T

    2009-04-01

    Amniotic fluid 'sludge' is defined as the presence of dense aggregates of particulate matter in close proximity to the internal cervical os. It is of clinical significance in asymptomatic patients at high risk for spontaneous delivery, and in patients with preterm labor and intact membranes. Subchorionic hematoma is another ultrasound finding that is associated with a higher incidence of threatened miscarriage and preterm delivery. We report two cases of occurrence of amniotic fluid sludge in patients with previously detected large subchorionic hematoma. In the first case subchorionic hematoma and amniotic fluid sludge were detected by ultrasonography at 13 + 1 and 18 + 6 weeks' gestation, respectively, followed by preterm premature rupture of membranes, placental abruption and emergency Cesarean section. In the second case subchorionic hematoma and amniotic fluid sludge were detected by ultrasound at 11 + 3 and 15 + 5 weeks' gestation, respectively, followed by miscarriage with histological chorioamnionitis. The coincidence of subchorionic hematoma and amniotic fluid sludge in these cases points to a possible connection between these two significant ultrasound findings. PMID:19308930

  18. Amniotic fluid 'sludge' detected in patients with subchorionic hematoma: a report of two cases.

    PubMed

    Tskitishvili, E; Tomimatsu, T; Kanagawa, T; Sawada, K; Kinugasa, Y; Mimura, K; Kimura, T

    2009-04-01

    Amniotic fluid 'sludge' is defined as the presence of dense aggregates of particulate matter in close proximity to the internal cervical os. It is of clinical significance in asymptomatic patients at high risk for spontaneous delivery, and in patients with preterm labor and intact membranes. Subchorionic hematoma is another ultrasound finding that is associated with a higher incidence of threatened miscarriage and preterm delivery. We report two cases of occurrence of amniotic fluid sludge in patients with previously detected large subchorionic hematoma. In the first case subchorionic hematoma and amniotic fluid sludge were detected by ultrasonography at 13 + 1 and 18 + 6 weeks' gestation, respectively, followed by preterm premature rupture of membranes, placental abruption and emergency Cesarean section. In the second case subchorionic hematoma and amniotic fluid sludge were detected by ultrasound at 11 + 3 and 15 + 5 weeks' gestation, respectively, followed by miscarriage with histological chorioamnionitis. The coincidence of subchorionic hematoma and amniotic fluid sludge in these cases points to a possible connection between these two significant ultrasound findings.

  19. Large, central acellular zones indicating myoepithelial tumor differentiation in high-grade invasive ductal carcinomas as markers of predisposition to lung and brain metastases.

    PubMed

    Tsuda, H; Takarabe, T; Hasegawa, F; Fukutomi, T; Hirohashi, S

    2000-02-01

    High-grade invasive ductal carcinomas (IDCs) of the breast with large, central acellular zones on their cut surfaces are usually associated with the myoepithelial immunophenotype of carcinoma cells, which includes the expression of S-100 protein, alpha-smooth muscle actin, and keratin 14. To clarify the clinical significance of these features of IDCs, the authors compared the incidence of the myoepithelial immunophenotype immunohistochemically, patient prognosis, and metastatic sites of the tumor between 20 high-grade IDCs with large, central acellular zones and 40 control high-grade IDCs without these zones. The myoepithelial immunophenotype was detected in 16 IDCs (80%) with large, central acellular zones but in only seven IDCs (18%) without. The risk ratio of metastasis, especially in the brain and lung, and death from cancer were significantly higher (p = 0.0096 and p = 0.030) for the 20 IDCs with large, central acellular zones than for those without by Cox's univariate analysis. Using Cox's multivariate analysis, large, central acellular zones in IDCs were an indicator of high risk of brain and lung metastases and of death by cancer independent of nodal status and tumor size. Examination of large, central acellular zones and myoepithelial immunophenotype in high-grade IDCs appears helpful in predicting patient prognosis and preferential metastatic sites of the tumors.

  20. Development of a Sterile Amniotic Membrane Tissue Graft Using Supercritical Carbon Dioxide.

    PubMed

    Wehmeyer, Jennifer L; Natesan, Shanmugasundaram; Christy, Robert J

    2015-07-01

    Numerous techniques have been reported for preparing and sterilizing amniotic membrane (AM) for use in clinical applications. However, these preparations either do not produce completely sterile tissue or are detrimental to molecules unique to the tissue matrix, thus compromising beneficial wound-healing properties of the AM graft. The objective of this work was to produce a sterile human AM tissue graft using a novel preparation technique involving supercritical carbon dioxide (SCCO2). AM tissue was subjected to various sterilization treatment groups that optimized the duration of exposure to SCCO2 and the amount of peracetic acid (PAA) required to achieve a sterility assurance level of 10(-6) log reduction in bacterial load. Effects of sterilization treatment on the histological, biophysical, and biochemical properties of the sterile AM were evaluated and compared with those of native AM tissue. Exposure of the AM tissue to combined SCCO2 and PAA sterilization treatment did not significantly alter tissue architecture, the amounts of pertinent extracellular matrix proteins (type IV collagen, glycosaminoglycans, elastin) present in the tissue, or the biophysical properties of the tissue. AMs treated with SCCO2 were also found to be excellent substrates for adipose-derived stem cell (ASC) attachment and proliferation in vitro. Human ASCs, attached to all treatment groups after 24 h of culture and continued to proliferate over the next few days. The current study's results indicate that SCCO2 can be used to sterilize AM tissue grafts while simultaneously preserving their biological attributes. The preservation of these features make AM appealing for use in numerous clinical and tissue engineering applications.

  1. In vitro and in vivo properties of distinct populations of amniotic fluid mesenchymal progenitor cells

    PubMed Central

    Roubelakis, Maria G; Bitsika, Vasiliki; Zagoura, Dimitra; Trohatou, Ourania; Pappa, Kalliopi I; Makridakis, Manousos; Antsaklis, Aristidis; Vlahou, Antonia; Anagnou, Nicholas P

    2011-01-01

    Abstract Human mesenchymal progenitor cells (MPCs) are considered to be of great promise for use in tissue repair and regenerative medicine. MPCs represent multipotent adherent cells, able to give rise to multiple mesenchymal lineages such as osteoblasts, adipocytes or chondrocytes. Recently, we identified and characterized human second trimester amniotic fluid (AF) as a novel source of MPCs. Herein, we found that early colonies of AF-MPCs consisted of two morphologically distinct adherent cell types, termed as spindle-shaped (SS) and round-shaped (RS). A detailed analysis of these two populations showed that SS-AF-MPCs expressed CD90 antigen in a higher level and exhibited a greater proliferation and differentiation potential. To characterize better the molecular identity of these two populations, we have generated a comparative proteomic map of SS-AF-MPCs and RS-AF-MPCs, identifying 25 differentially expressed proteins and 10 proteins uniquely expressed in RS-AF-MPCs. Furthermore, SS-AF-MPCs exhibited significantly higher migration ability on extracellular matrices, such as fibronectin and laminin in vitro, compared to RS-AF-MPCs and thus we further evaluated SS-AF-MPCs for potential use as therapeutic tools in vivo. Therefore, we tested whether GFP-lentiviral transduced SS-AF-MPCs retained their stem cell identity, proliferation and differentiation potential. GFP-SS-AF-MPCs were then successfully delivered into immunosuppressed mice, distributed in different tissues and survived longterm in vivo. In summary, these results demonstrated that AF-MPCs consisted of at least two different MPC populations. In addition, SS-AF-MPCs, isolated based on their colony morphology and CD90 expression, represented the only MPC population that can be expanded easily in culture and used as an efficient tool for future in vivo therapeutic applications. PMID:21166769

  2. Development of a Sterile Amniotic Membrane Tissue Graft Using Supercritical Carbon Dioxide.

    PubMed

    Wehmeyer, Jennifer L; Natesan, Shanmugasundaram; Christy, Robert J

    2015-07-01

    Numerous techniques have been reported for preparing and sterilizing amniotic membrane (AM) for use in clinical applications. However, these preparations either do not produce completely sterile tissue or are detrimental to molecules unique to the tissue matrix, thus compromising beneficial wound-healing properties of the AM graft. The objective of this work was to produce a sterile human AM tissue graft using a novel preparation technique involving supercritical carbon dioxide (SCCO2). AM tissue was subjected to various sterilization treatment groups that optimized the duration of exposure to SCCO2 and the amount of peracetic acid (PAA) required to achieve a sterility assurance level of 10(-6) log reduction in bacterial load. Effects of sterilization treatment on the histological, biophysical, and biochemical properties of the sterile AM were evaluated and compared with those of native AM tissue. Exposure of the AM tissue to combined SCCO2 and PAA sterilization treatment did not significantly alter tissue architecture, the amounts of pertinent extracellular matrix proteins (type IV collagen, glycosaminoglycans, elastin) present in the tissue, or the biophysical properties of the tissue. AMs treated with SCCO2 were also found to be excellent substrates for adipose-derived stem cell (ASC) attachment and proliferation in vitro. Human ASCs, attached to all treatment groups after 24 h of culture and continued to proliferate over the next few days. The current study's results indicate that SCCO2 can be used to sterilize AM tissue grafts while simultaneously preserving their biological attributes. The preservation of these features make AM appealing for use in numerous clinical and tissue engineering applications. PMID:25471248

  3. Amniotic Fluid-Derived Mesenchymal Stem Cells Prevent Fibrosis and Preserve Renal Function in a Preclinical Porcine Model of Kidney Transplantation

    PubMed Central

    Baulier, Edouard; Favreau, Frederic; Le Corf, Amélie; Jayle, Christophe; Schneider, Fabrice; Goujon, Jean-Michel; Feraud, Olivier; Bennaceur-Griscelli, Annelise; Turhan, Ali G.

    2014-01-01

    It is well known that ischemia/reperfusion injuries strongly affect the success of human organ transplantation. Development of interstitial fibrosis and tubular atrophy is the main deleterious phenomenon involved. Stem cells are a promising therapeutic tool already validated in various ischemic diseases. Amniotic fluid-derived mesenchymal stem cells (af-MSCs), a subpopulation of multipotent cells identified in amniotic fluid, are known to secrete growth factors and anti-inflammatory cytokines. In addition, these cells are easy to collect, present higher proliferation and self-renewal rates compared with other adult stem cells (ASCs), and are suitable for banking. Consequently, af-MSCs represent a promising source of stem cells for regenerative therapies in humans. To determine the efficiency and the safety of af-MSC infusion in a preclinical porcine model of renal autotransplantation, we injected autologous af-MSCs in the renal artery 6 days after transplantation. The af-MSC injection improved glomerular and tubular functions, leading to full renal function recovery and abrogated fibrosis development at 3 months. The strong proof of concept generated by this translational porcine model is a first step toward evaluation of af-MSC-based therapies in human kidney transplantation. PMID:24797827

  4. Egg shape changes at the theropod-bird transition, and a morphometric study of amniote eggs.

    PubMed

    Deeming, D Charles; Ruta, Marcello

    2014-11-01

    The eggs of amniotes exhibit a remarkable variety of shapes, from spherical to elongate and from symmetrical to asymmetrical. We examine eggshell geometry in a diverse sample of fossil and living amniotes using geometric morphometrics and linear measurements. Our goal is to quantify patterns of morphospace occupation and shape variation in the eggs of recent through to Mesozoic birds (neornithe plus non-neornithe avialans), as well as in eggs attributed to non-avialan theropods. In most amniotes, eggs show significant deviation from sphericity, but departure from symmetry around the equatorial axis is mostly confined to theropods and birds. Mesozoic bird eggs differ significantly from extant bird eggs, but extinct Cenozoic bird eggs do not. This suggests that the range of egg shapes in extant birds had already been attained in the Cenozoic. We conclude with a discussion of possible biological factors imparting variation to egg shapes during their formation in the oviduct.

  5. LIF analysis of cervical mucus and amniotic fluid for maturity monitoring in pregnancy

    NASA Astrophysics Data System (ADS)

    Vaitkuviene, Aurelija; Auksorius, Egidijus; Ramasauskaite, Diana; Smilgeviciute, Ale; Tamasauskas, Oldas; Vanseviciute, Rasa; Veleckas, Doras

    2004-09-01

    The aim of this study was to assess the feasibility of autoflorescence spectroscopy in the diagnosis of cervix maturity through cervical mucus florescence and foetal lung maturity through amniotic fluid fluorescence. LED and broadband Mercury light were used to induce fluorescence in cervical mucus and amniotic fluid respectively. Mature specimens compared to immature ones showed a significant decrease in cervical mucus fluorescence values measured at 420 nm (p = 0.0004) and in measured amniotic fluid fluorescence values at 410 nm (p = 0.0686). Probability-based classification algorithm was developed to identify samples 'maturity' through analysis of the fluorescence spectra. Employing fluorescence intensity at 420 nm for cervix maturity diagnosis rendered optimal sensitivity of 92.9%, specificity of 83.3% and area under the ROC curve of 91.1%.

  6. Pathogenesis and management of peripartum coagulopathic calamities (disseminated intravascular coagulation and amniotic fluid embolism).

    PubMed

    Levi, Marcel

    2013-01-01

    Acute coagulopathic peripartum calamities are relatively rare but contribute importantly to maternal morbidity and mortality in the Western world. Abruptio placenta, amniotic fluid embolism, and retained fetal or placental material may lead to fulminant intravascular activation of coagulation which results in thromboembolic complications and consumption coagulopathy causing severe hemorrhage. The central underlying pathophysiological pathway in the coagulopathy associated with these syndromes is the occurrence of tissue factor, released from the placenta and amniotic fluid, in the circulation, in combination with low levels of physiological anticoagulant factors during pregnancy. The diagnosis of DIC may be made trough conventional composite scoring systems employing routine coagulation tests, whereas for the diagnosis of amniotic fluid embolism measurement of insulin like growth factor binding protein-1 seems promising. Therapy is aimed at removing the precipitating factor combined with supportive adjunctive treatment options.

  7. The epithelial mesenchymal transition process may contribute to the pathogenesis of amniotic band syndrome.

    PubMed

    Romero-Valdovinos, M; Bobadilla-Sandoval, N; Flisser, A; Vadillo-Ortega, F

    2014-09-01

    The etiology of the amniotic band syndrome is unknown, and has been subject of debate since the time of Hippocrates. The most accepted theories fail to cover all the abnomalities found in affected children. During organogenesis the epithelial-mesenchymal transition process (EMTP) participates in adequate formation of different organs from three embryo layers. Altered activation of EMTP occurs when the epithelial homeostasis is disturbed, the resulting myofibroblasts are able to secrete extracellular matrix proteins and deposit them on the tissues contributing to a fibrotic phenotype. If injury occurs during organogenesis, wound healing could be exaggerated and fibrotic response could be triggered. The molecule that regulates both of these processes (EMTP and fibrosis) is the transforming growth factor β (TGFβ); indeed null animals for TGFβ isoforms show similar defects than those seen in the amniotic band syndrome. Based on documented evidence this review intends to explain how the epithelial mesenchymal transition process may contribute to the pathogenesis of amniotic band syndrome. PMID:24998668

  8. Amniote yolk sacs: diversity in reptiles and a hypothesis on their origin.

    PubMed

    Elinson, Richard P; Stewart, James R; Bonneau, Laurie J; Blackburn, Daniel G

    2014-01-01

    Oviparous amniotes produce a large yolky egg that gives rise to a free-living hatchling. Structural characteristics and functional attributes of the egg are best known for birds, which have a large mass of fluid yolk surrounded by an extraembryonic yolk sac. Yolk nutrients are delivered to the embryo via the vascular yolk sac. This developmental pattern and nutrient transport mechanism is thought to be representative of all other lineages of amniotes. Recent discovery of a snake with cellularized yolk organized around a meshwork of blood vessels reveals an additional pattern for yolk mobilization, which may also occur in other squamate reptiles (lizards and snakes). This complex yolk sac raises interesting questions about developmental mechanisms and suggests a possible model for the transition between the egg of anamniotes and that of amniotes.

  9. Use of amniotic membrane graft in the surgical management of cicatricial ectropion associated with cetuximab therapy.

    PubMed

    Vinod, Kateki; Diaz, Vicente

    2015-01-01

    Cetuximab (Erbitux) is an antiepidermal growth factor receptor (EGFR) monoclonal antibody that has been shown to delay the progression of metastatic colorectal cancer. The cutaneous side effects of cetuximab resulting from its effects on normal epidermal cells are well established. Periocular side effects, including blepharitis, trichomegaly, dry eye and conjunctivitis, have also been reported. We present a case of cicatricial ectropion associated with cetuximab therapy successfully managed with surgical repair using amniotic membrane graft. A 60-year-old man presented with bilateral lower eyelid cicatricial ectropion developing 3 days after the addition of cetuximab therapy to his baseline chemotherapeutic regimen. This was successfully managed with surgical repair using an amniotic membrane graft. Surgical repair with the amniotic membrane graft is a viable treatment option for cicatricial ectropion associated with EGFR inhibitor therapy. PMID:25576167

  10. Comparative Transcriptome Analysis of Cell-Free Fetal RNA from Amniotic Fluid and RNA from Amniocytes in Uncomplicated Pregnancies

    PubMed Central

    Jung, Y. W.; Shim, S. H.; Sung, S. R.; Park, J. E.; Cha, D. H.; Ahn, E. H

    2015-01-01

    Objectives We aimed to compare tissue-specific expression profiles and biological pathways of RNA from amniocytes and amniotic fluid supernatant (AFS) from second-trimester pregnancies by using transcriptome analysis. Additionally, we wanted to explore whether cell-free RNA from AFS exhibits a unique gene expression signature that more adequately reflects the fetal developmental process than amniocyte RNA. Methods Amniotic fluid samples were prospectively collected in the second trimester of pregnancy from euploid fetuses. Total RNA was extracted from amniocytes and AFS and hybridized to Affymetrix GeneChip Human Arrays. Significantly differentially expressed transcripts between amniocytes and AFS were obtained by using Welch’s t-test. Unsupervised hierarchical clustering was used to visualize overall expression characteristics and differences in transcripts between AFS and amniocytes. The biological functions of selected genes were analyzed using various online Gene Ontology databases. Results A total of 3,072 and 15,633 transcripts were detected in the second-trimester AFS and amniocytes, respectively. Hierarchical clustering revealed differential transcript expression between AFS and amniocytes. We found 353 genes that were specifically enriched in the AFS only, and tissue expression analysis showed enrichment of brain-specific genes in the AFS. Biological pathway analysis revealed that AFS-specific transcripts were mainly involved in embryonic development, cardiovascular development, and cellular morphology pathways. Conclusion This study demonstrated differential tissue-specific gene expression profiles and biological pathways between AFS and amniocytes. The results suggested that AFS is the preferred RNA source to investigate potential biomarkers of fetal neurodevelopment. PMID:26181329

  11. Preliminary Results of Bioactive Amniotic Suspension with Allograft for Achieving One and Two-Level Lumbar Interbody Fusion

    PubMed Central

    Kerr, Eubulus J.; Utter, Philip A.; Cavanaugh, David A.; Frank, Kelly A.; Moody, Devan; McManus, Brian; Stone, Marcus B.

    2016-01-01

    Background Bone graft material for lumbar fusion was historically autologous bone graft (ABG). In recent years alternatives such as allograft, demineralized bone matrix (DBM), ceramics, and bone morphogenetic protein (BMP) have gained favor, although the complications of these are not fully understood. Bioactive amniotic suspension (BAS) with allograft is a new class of material derived from human amniotic tissue. Methods Eligible patients receiving a one or two level lumbar interbody fusion with Nucel, a BAS with allograft, were contacted and scheduled for a mininmim 12 month follow-up visit. Patients were evaluated for fusion using CT's and plain radiographs. Clincal outcomes, including ODI, VAS back and leg were collected, as well as comorbidities including BMI, smoking status, diabetes and previous lumbar surgery. Results One-level patients (N=38) were 71.1% female with mean age of 58.4 ± 12.7 and mean BMI of 30.6 ± 6.08. Two-level patients (N=34) were 58.8% female with mean age of 49.3 ±10.9 and mean BMI of 30.1 ± 5.82. Kinematic fusion was achieved in 97.4% of one-level patients and 100% of two-level patients. Baseline comorbidities were present in 89.5% of one-level patients and 88.2% of two-level patients. No adverse events related to BAS were reported in this study. Conclusion Fusion status is evaluated with many different biologics and varying methods in the literature. BAS with allograft in this study demonstrated high fusion rates with no complications within a largely comorbid population. Although a small population, BAS with allograft results were encouraging for one and two-level lumbar interbody fusion in this study. Further prospective studies should be conducted to investigate safety and efficacy in a larger population. PMID:27162714

  12. The Influence of Fasting in Summer on Amniotic Fluid During Pregnancy

    PubMed Central

    Altunkeser, Ayşegül; Körez, Muslu Kazım

    2016-01-01

    Objectives: Ramadan is a holy month in which eating and drinking are forbidden from dawn to sunset. In this study, we investigated using ultrasonography (USG) whether fasting in summer (as occurred in Ramadan 2014) had an influence on the volume of amniotic fluid during pregnancy. Materials and Methods: The study included 119 pregnant women in total who were admitted to our department with a request of obstetric USG between June 28, 2014, and July 27, 2014. The fasting group included 61 pregnant women and the control group of 58 pregnant women. In our study, all the fasting pregnant women had Sahur (predawn meal eaten before starting fasting) and Iftar (the evening meal for fast-breaking) every day, regularly. The women in the control group did not fast. In addition to amniotic fluid index and fetal biometric measurements during Ramadan, amniotic fluid volume was measured ultrasonographically throughout pregnancy. All ultrasound examinations were performed at least 8 h after Sahur during Ramadan. Chi-square test was utilized to compare the measurements of amniotic fluid volume, and Mann–Whitney U-test was utilized to analyze the differences in fetal growth data. Moreover, difference was considered statistically significant when the P value was <0.05. Results: The mean age was 25.7 years in the fasting group and 25.8 years in the control group. Other characteristics and mean gestational weeks of the two groups were similar. Ultrasonographically, there was no significant difference between two groups in respect to amniotic fluid amount during pregnancy (P = 0.7). There was no significant difference with regard to fetal growth parameters either (P > 0.05). Conclusion: In pregnant women who had regular predawn and fast-breaking meals, fasting in summer did not elicit alteration in the amount of amniotic fluid throughout pregnancy. PMID:27313975

  13. Antioxidant Vitamin Status in the Serum and Amniotic Fluid of Women with Premature Rupture of the Fetal Membranes.

    NASA Astrophysics Data System (ADS)

    Barrett, Bridget M.

    The purpose of this study was to examine the status of antioxidant vitamins in women with premature rupture of the fetal membranes. Specimens of blood and amniotic fluid were obtained from 80 pregnant subjects included both smokers and non-smokers during the third trimester. The concentrations of ascorbic acid (ASA), beta -carotene, retinol and alpha -tocopherol in serum and amniotic fluid were determined. The experimental group consisted of those subjects with PROM while the control subjects were those with normal pregnancy. No statistical differences were found between the PROM and control groups in retinol and vitamin E concentrations in amniotic fluid and serum. Serum ASA concentrations of PROM subjects were not different from controls, but the PROM subjects had significantly lower amniotic fluid ASA concentrations. However, in a study with fewer subjects a lower serum ASA concentration in the PROM subjects was observed. The ratio of amniotic fluid ASA concentration to ASA serum concentration was significantly lower in PROM patients than in controls in both studies. This suggests that low levels of ASA in the amniotic fluid, but not in serum is better associated with PROM. A low amniotic fluid concentration of ASA may reflect an inefficient transfer and/or increased fetal utilization. Alterations in ASA concentration in the amniotic fluid may affect the integrity of the chorioamnion leading to PROM. beta -Carotene was not found in the amniotic fluid. Serum beta-carotene levels were significantly lower in the PROM group compared to the control group. Low concentrations of beta-carotene in maternal serum in smokers not only associated with poor maternal outcome (PROM) but also compromised the fetal outcome (decreased birth weight). Maintenance of adequate serum beta-carotene concentration and amniotic fluid ASA in smokers may result in better maternal and fetal outcome. This study demonstrated that nutrition is an important factor in the prevention of PROM.

  14. Association of postpartum maternal tetanus toxoid, reduced diphtheria toxoid and acellular pertussis vaccine administration and timeliness of infant immunization.

    PubMed

    Kaur, Ishminder; George, Krissa J; Pena-Ricardo, Carolina; Kelly, Barbara A; Watson, Barbara

    2013-11-01

    A retrospective cohort study was conducted on infants of mothers delivering at an inner-city hospital in October 2009 where postpartum maternal tetanus toxoid, reduced diptheria toxoid and acellular pertussis (Tdap) vaccination had been initiated in May 2008. We compared mothers and infants in a Tdap intervention group discharged July 2008 (n=250) with a pre-Tdap control group discharged July 2007 (n=238). Postpartum maternal Tdap impacted positively timeliness of early infant immunization.

  15. Possible role of dentin matrix in region-specific deposition of cellular and acellular extrinsic fibre cementum.

    PubMed

    Takano, Yoshiro; Sakai, Hideo; Watanabe, Eiko; Ideguchi-Ohma, Noriko; Jayawardena, Chantha K; Arai, Kazumi; Asawa, Yukiyo; Nakano, Yukiko; Shuda, Yoko; Sakamoto, Yujiro; Terashima, Tatsuo

    2003-01-01

    The mechanism whereby a region-specific deposition of the two types of cementum (cellular cementum and acellular extrinsic fibre cementum) is regulated on the growing root surface was tested using bisphosphonate-affected teeth of young rats and guinea pigs. The animals were injected subcutaneously with 8 or 10 mg P x kg body weight(-1) x day(-1) of 1-hydroxyethylidene-1,1-bisphosphonate (HEBP) for 1 or 2 weeks. In rat molars, HEBP prevented mineralization of newly formed root dentin matrix and totally inhibited de novo deposition of acellular extrinsic fibre cementum. Instead, thick cellular cementum was induced on the non-mineralized root dentin surface, irrespective of the position of the root. In both animals, cellular cementum was also induced on the non-mineralized surface of root analogue dentin in HEBP-affected incisors, where only acellular extrinsic fibre cementum is deposited under normal conditions. In normal rat molars, dentin sialoprotein (DSP) was concentrated along the dentin-cellular cementum border, but not that of dentin and acellular extrinsic fibre cementum. In HEBP-affected rat incisors, DSP was shown to penetrate through the non-mineralized dentin into the surrounding tissues, but not through the mineralized portions. These data suggest that, at the site of cellular cementum formation, putative inducing factors for cellular cementum might diffuse into the periodontal space through the newly deposited mantle dentin matrix before it is mineralized. At earlier stages of root formation, mantle dentin might mineralize more promptly not to allow such diffusion. The timing of mineralization of mantle dentin matrix might be the key determinant of the types of the cementum deposited on the growing root surface. PMID:14756246

  16. Evo-Devo of Amniote Integuments and Appendages

    PubMed Central

    Wu, Ping; Hou, Lianhai; Plikus, Maksim; Hughes, Michael; Scehnet, Jeffrey; Suksaweang, Sanong; Widelitz, Randall B.; Jiang, Ting-Xin; Chuong, Cheng-Ming

    2015-01-01

    Integuments form the boundary between an organism and the environment. The evolution of novel developmental mechanisms in integuments and appendages allows animals to live in diverse ecological environments. Here we focus on amniotes. The major achievement for reptile skin is an adaptation to the land with the formation of a successful barrier. The stratum corneum enables this barrier to prevent water loss from the skin and allowed amphibian/reptile ancestors to go onto the land. Overlapping scales and production of β-keratins provide strong protection. Epidermal invagination led to the formation of avian feather and mammalian hair follicles in the dermis. Both adopted a proximal - distal growth mode that maintains endothermy. Feathers form hierarchical branches which produce the vane that makes flight possible. Recent discoveries of feathered dinosaurs in China inspire new thinking on the origin of feathers. In the laboratory, epithelial - mesenchymal recombinations and molecular mis-expressions were carried out to test the plasticity of epithelial organ formation. We review the work on the transformation of scales into feathers, conversion between barbs and rachis, and the production of “chicken teeth”. In mammals, tilting the balance of the BMP pathway in K14 noggin transgenic mice alters the number, size and phenotypes of different ectodermal organs, making investigators rethink the distinction between morpho-regulation and pathological changes. Models on the evolution of feathers and hairs from reptile integuments are discussed. A hypothetical Evo-Devo space where diverse integument appendages can be placed according to complex phenotypes and novel developmental mechanisms is presented. PMID:15272390

  17. Amniotic Fluid Metabolomic Analysis in Spontaneous Preterm Birth

    PubMed Central

    Jones, Janice; Gunst, Phillip R.; Kacerovsky, Marian; Fortunato, Stephen J.; Saade, George R.; Basraon, Sanmaan

    2014-01-01

    Objective: To identify metabolic changes associated with early spontaneous preterm birth (PTB; <34 weeks) and term births, using high-throughput metabolomics of amniotic fluid (AF) in African American population. Method: In this study, AF samples retrieved from spontaneous PTB (<34 weeks [n = 25]) and normal term birth (n = 25) by transvaginal amniocentesis at the time of labor prior to delivery were subjected to metabolomics analysis. Equal volumes of samples were subjected to a standard solvent extraction method and analyzed using gas chromatography/mass spectrometry (MS) and liquid chromatography/MS/MS. Biochemicals were identified through matching of ion features to a library of biochemical standards. After log transformation and imputation of minimum observed values for each compound, t test, correlation tests, and false discovery rate corrections were used to identify differentially regulated metabolites. Data were controlled for clinical/demographic variables and medication during pregnancy. Results: Of 348 metabolites measured in AF samples, 121 metabolites had a gestational age effect and 116 differed significantly between PTB and term births. A majority of significantly altered metabolites could be classified into 3 categories, namely, (1) liver function, (2) fatty acid and coenzyme A (CoA) metabolism, and (3) histidine metabolism. The signature of altered liver function was apparent in many cytochrome P450-related pathways including bile acids, steroids, xanthines, heme, and phase II detoxification of xenobiotics with the largest fold change seen with pantothenol, a CoA synthesis inhibitor that was 8-fold more abundant in PTB. Conclusion: Global metabolic profiling of AF revealed alteration in hepatic metabolites involving xenobiotic detoxification and CoA metabolism in PTB. Maternal and/or fetal hepatic function differences may be developmentally related and its contribution PTB as a cause or effect of PTB is still unclear. PMID:24440995

  18. Simultaneous multiresponse optimization of an HPLC method to separate seven cephalosporins in plasma and amniotic fluid: application to validation and quantification of cefepime, cefixime and cefoperazone.

    PubMed

    Nemutlu, Emirhan; Kir, Sedef; Katlan, Doruk; Beksaç, M Sinan

    2009-11-15

    An HPLC method for the separation of seven cephalosporins [Cefepime (CEP), ceftazidime (CTA), ceftizaxime (CTI), ceftriaxone (CTR), cefotaxime (COT), cefixime (CIX) and cefoperazone (COP)] in human plasma and amniotic fluid has been developed. Optimization of the chromatographic method was performed in three steps: a series of initial experiments followed by two sets of experiments based on different experimental designs. The initial experiments were performed to decide the basic analytical requirements of the method. Then screening experiment fractional factorial design was used in order to decrease the number of parameters by eliminating parameters which having insignificant effect on responses. The parameters having significant effect were further optimized through a full factorial design. Having studied two responses (retention times and resolutions), a desirability function that assess the responses together, was used to find experimental conditions where the system generated desirable results. The desirable results were obtained with XTerra C18 (250 mm x 4.6mm, 5 microm i.d.) column, 40 mM phosphate buffer, pH 3.2, 18% MeOH, 0.85 mL min(-1) flow rate and 32 degrees C column temperature. Gradient elution with MeOH was applied. A simple and efficient solid-phase extraction was applied for the preparation of plasma and amniotic fluid samples. The validation parameters of the method were evaluated in accordance with ICH guideline. The method validated was applied to the analysis of CEP and COP in maternal venous, fetal venous and fetal arterial plasma, and to the analysis of CIX in maternal venous plasma and amniotic fluid.

  19. Amniotic Fluid or Its Fatty Acids Produce Actions Similar to Diazepam on Lateral Septal Neurons Firing Rate

    PubMed Central

    Gutiérrez-García, Ana G.; Vásquez-Hernández, Diana Idania

    2013-01-01

    Human amniotic fluid (AF) contains eight fatty acids (FATs), and both produce anxiolytic-like effects in adult rats and appetitive responses in human newborns. The medial amygdala and lateral septal nucleus function are related to social behavior, but the action of AF or its FATs in this circuit is known. We obtained 267 single-unit extracellular recordings in Wistar rats treated with vehicle (1 mL, s.c.; n = 12), human AF (1 mL, s.c.; n = 12), a FAT mixture (1 mL, s.c.; n = 13), diazepam (1 mg/kg, i.p.; n = 11), and fluoxetine (1 mg/kg, p.o.; n = 12). Compared with the vehicle group, the spontaneous septal firing rate in the AF, FAT mixture, and diazepam groups was the lowest and in the fluoxetine group the highest. Cumulative peristimulus histograms indicated that the significant change in septal firing occurred only in the AF and FAT mixture groups and exclusively in those neurons that increased their firing rate during amygdala stimulation. We conclude that human AF and its FATs produce actions comparable to anxiolytic drugs and are able to modify the responsivity of a circuit involved in social behavior, suggesting facilitation of social recognition processes by maternal-fetal fluids. PMID:23864826

  20. Augmented Dried versus Cryopreserved Amniotic Membrane as an Ocular Surface Dressing

    PubMed Central

    Allen, Claire L.; Clare, Gerry; Stewart, Elizabeth A.; Branch, Matthew J.; McIntosh, Owen D.; Dadhwal, Megha; Dua, Harminder S.; Hopkinson, Andrew

    2013-01-01

    Purpose Dried amniotic membrane (AM) can be a useful therapeutic adjunct in ophthalmic surgery and possesses logistical advantages over cryopreserved AM. Differences in preservation techniques can significantly influence the biochemical composition and physical properties of AM, potentially affecting clinical efficacy. This study was established to investigate the biochemical and structural effects of drying AM in the absence and presence of saccharide lyoprotectants and its biocompatibility compared to cryopreserved material. Methods AM was cryopreserved or dried with and without pre-treatment with trehalose or raffinose and the antioxidant epigallocatechin (EGCG). Structural and visual comparisons were assessed using electron microscopy. Localisation, expression and release of AM biological factors were determined using immunoassays and immunofluorescence. The biocompatibility of the AM preparations co-cultured with corneal epithelial cell (CEC) or keratocyte monolayers were assessed using cell proliferation, cytotoxicity, apoptosis and migration assays. Results Drying devitalised AM epithelium, but less than cryopreservation and cellular damage was reduced in dried AM pre-treated with trehalose or raffinose. Dried AM alone, and with trehalose or raffinose showed greater factor retention efficiencies and bioavailability compared to cryopreserved AM and demonstrated a more sustained biochemical factor time release in vitro. Cellular health assays showed that dried AM with trehalose or raffinose are compatible and superior substrates compared to cryopreserved AM for primary CEC expansion, with increased proliferation and reduced LDH and caspase-3 levels. This concept was supported by improved wound healing in an immortalised human CEC line (hiCEC) co-cultured with dried and trehalose or raffinose membranes, compared to cryopreserved and fresh AM. Conclusions Our modified preservation process and our resultant optimised dried AM has enhanced structural properties

  1. Mesenchymal stem cells from amnion and amniotic fluid in the bovine.

    PubMed

    Corradetti, B; Meucci, A; Bizzaro, D; Cremonesi, F; Lange Consiglio, A

    2013-04-01

    Amnion and amniotic fluid (AF) are noncontroversial and inexhaustible sources of mesenchymal stem cells (MSCs) that can be harvested noninvasively at low cost. As in humans, also in veterinary field, presumptive stem cells derived from these tissues reveal as promising candidates for disease treatment, specifically for their plasticity, their reduced immunogenicity, and high anti-inflammatory potential. The aim of this work is to obtain and characterize, for the first time in bovine species, presumptive MSCs from the epithelial portion of the amnion (AECs) and from the AF (AF-MSCs) to be used for clinical applications. AECs display a polygonal morphology, whereas AF-MSCs exhibit a fibroblastic-like morphology only starting from the second passage, being heterogeneous during the primary culture. For both lines, the proliferative ability has been found constant over the ten passages studied and AECs show a statistically lower (P<0.05) doubling time with respect to AF-MSCs. AECs express MSC-specific markers (ITGB1 (CD29), CD44, ALCAM (CD166), ENG (CD105), and NT5E (CD73)) from P1 to P3; in AF-MSCs, only ITGB1, CD44, and ALCAM mRNAs are detected; NT5E is expressed from P2 and ENG has not been found at any passage. AF-MSCs and AECs are positive for the pluripotent markers (POU5F1 (OCT4) and MYC (c-Myc)) and lack of the hematopoietic markers. When appropriately induced, both cell lines are capable of differentiating into ectodermal and mesodermal lineages. This study contributes to reinforce the emerging importance of these cells as ideal tools in veterinary medicine. A deeper evaluation of the immunological properties needs to be performed in order to better understand their role in cellular therapy.

  2. Epigenetic analysis and suitability of amniotic fluid stem cells for research and therapeutic purposes.

    PubMed

    Phermthai, Tatsanee; Suksompong, Singpetch; Tirawanchai, Nednapis; Issaragrisil, Surapol; Julavijitphong, Suphakde; Wichitwiengrat, Suparat; Silpsorn, Decha; Pokathikorn, Puttachart

    2013-05-01

    Amniotic fluid stem cells (AFSs) are interesting mesenchymal stem cells (MSCs) that are characterized by their great potential for cell proliferation and differentiation compared with other types of MSCs identified to date. However, MSCs in prolonged culture have been found to exhibit defects in genetic stability and differentiation capacity. Epigenetic anomalies have been hypothesized to be a cause of these defects. Here, we investigated the genomic methylation and genetic imprinting in AFSs during prolonged in vitro culture. Four human imprinted genes, insulin-like growth factor 2 (IGF2), H19, small nuclear ribonucleoprotein polypeptide N gene (SNRPN), and mesoderm-specific transcript (MEST), were evaluated for their expression levels and methylation statuses in AFS lines. The data revealed epigenetic instability in high passage number AFS cultures. The real-time polymerase chain reaction analysis showed that the expression levels of the imprinted genes gradually increased with increased time in culture. The loss of parental allele-specific imprinting for at least 1 gene among IGF2, H19, and SNRPN was observed in every AFS line after passage 8 using allelic expression analysis. The imprinting control regions (ICRs) of the IGF2 and H19 genes were assayed for site-specific methylation using bisulfite sequencing. This assay revealed a variable level of methylated CpG sites in the ICRs of IGF2 and H19. This variable level of CpG methylation is related to the aberrant expression of the IGF2 and H19 genes in late-passage AFSs. Our results did not reveal any irregularity in the epigenetic control system in the early-passage AFSs, indicating that the standard in vitro culturing of AFSs used in medical treatments should be limited to 8 passages.

  3. T-Cell Immune Response Assessment as a Complement to Serology and Intranasal Protection Assays in Determining the Protective Immunity Induced by Acellular Pertussis Vaccines in Mice

    PubMed Central

    Ausiello, C. M.; Lande, R.; Stefanelli, P.; Fazio, C.; Fedele, G.; Palazzo, R.; Urbani, F.; Mastrantonio, P.

    2003-01-01

    The relative value of antibodies and/or T-cell immune responses to Bordetella pertussis antigens in the immunity induced by acellular pertussis (aP) vaccines is still an open issue, probably due to the incomplete knowledge on the mechanisms of protective immunity to pertussis. The relevance of T-cell immune responses in protection from pertussis has been demonstrated in murine and human models of infection; thus, in this study, the ability of different vaccine preparations of three component (pertussis toxin, filamentous hemagglutinin, and pertactin) aP vaccines to induce T-cell responses was investigated in mice. All vaccine preparations examined passed the immunogenicity control test, based on antibody titer assessment, according to European Pharmacopoeia standards, and protected mice from B. pertussis intranasal challenge, but not all preparations were able to prime T cells to pertussis toxin, the specific B. pertussis antigen. In particular, one vaccine preparation was unable to induce proliferation and gamma interferon (IFN-γ) production while the other two gave borderline results. The evaluation of T-cell responses to pertussis toxin antigen may provide information on the protective immunity induced by aP vaccines in animal models. Considering the critical role of the axis interleukin-12-IFN-γ for protection from pertussis, our results suggest that testing the induction of a key protective cytokine such as IFN-γ could be an additional tool for the evaluation of the immune response induced by aP vaccines. PMID:12853397

  4. The use of an acellular dermal regenerative tissue matrix in the treatment of lower extremity wounds: a prospective 16-week pilot study.

    PubMed

    Brigido, Stephen A

    2006-09-01

    A prospective, single-centre, randomized controlled study was performed to evaluate the effectiveness of Graftjacket, a human acellular regenerative tissue matrix as a treatment option for chronic non healing lower extremity wounds. Twenty-eight diabetic patients with full-thickness wounds that had been present for at least 6 weeks were treated with sharp debridement and randomized to a single application of Graftjacket tissue matrix plus mineral oil-soaked fluff compression dressing or to a control treatment of wound gel with gauze dressings. All patients were seen weekly. By week 16, 12 of 14 patients treated with Graftjacket tissue matrix demonstrated complete wound closure compared with 4 of 14 patients in the control group. Patients treated with Graftjacket tissue matrix showed a statistically significant higher percentage of wound healing with respect to wound area, and clinically significant differences in wound depth and wound volume. This comparison is not performed to demonstrate that the application of the Grafjacket is more effective than sharp debridement. This study is done to help assign a role to the use of Graftjacket matrix in lower extremity wound care.

  5. Comparison of Achilles tendon repair techniques in a sheep model using a cross-linked acellular porcine dermal patch and platelet-rich plasma fibrin matrix for augmentation.

    PubMed

    Sarrafian, Tiffany L; Wang, Hali; Hackett, Eileen S; Yao, Jian Q; Shih, Mei-Shu; Ramsay, Heather L; Turner, A Simon

    2010-01-01

    The primary goal of this study was to evaluate a cross-linked acellular porcine dermal patch (APD), as well as platelet-rich plasma fibrin matrix (PRPFM), for repair of acute Achilles tendon rupture in a sheep model. The 2 surgically transected tendon ends were reapproximated in groups 1 and 2, whereas a gap was left between the tendon ends in group 3. APD was used to reinforce the repair in group 2, and autologous PRPFM was used to fill the gap, which was also reinforced with APD, in group 3. All sheep were humanely euthanized at 24 weeks after the repair, and biomechanical and histological testing were performed. Tensile strength testing showed a statistically significant difference in elongation between the operated limb and the unoperated contralateral limb in groups 1 and 3, but not in group 2. All operated tendons appeared healed with no apparent fibrosis under light and polarized microscopy. In group 1, all surgical separation sites were identifiable, and healing occurred via increasing tendon thickness. In group 2, healing occurred with new tendon fibers across the separation, without increasing tendon thickness in 2 out of 6 animals. Group 3 showed complete bridging of the gap, with no change in tendon thickness in 2 out of 6 animals. In groups 2 and 3, peripheral integration of the APD to tendon fibers was observed. These findings support the use of APD, alone or with PRPFM, to augment Achilles tendon repair in a sheep model.

  6. Amniotic membrane can be a valid source for wound healing

    PubMed Central

    ElHeneidy, Hossam; Omran, Eman; Halwagy, Ahmed; Al-Inany, Hesham; Al-Ansary, Mirvat; Gad, Amr

    2016-01-01

    Amniotic membrane (AM) can promote proper epithelialization with suppression of excessive fibrosis by creating a supportive milieu for regeneration of chronic ulcer bed. Objective The objective of this study is to investigate whether AM scaffold can modulate the healing of a wound by promoting tissue reconstruction rather than promoting scar tissue formation. Subjects and methods AM was obtained and prepared and then applied to patients with chronic leg ulcers who were randomly divided into two different groups. Group I (control group) included eleven patients in whom ulcers were treated with conventional wound dressings that were changed daily for 8 weeks. Group II (study group) included 14 patients in whom the AM was placed in contact with the ulcer and held in place with a secondary dressing, which was changed daily. Follow-up was done to detect healing rate and detection of ulcer size, assessment of pain, and to take ulcer images (days 0, 7, 14, 21, 30, 45, and 60). Results In group I, all ulcers showed no reduction in their size, and ulcer floor remained the same. Healthy granulations were present in two ulcers (18.2%) and absent in nine ulcers (81.8%). There was no improvement of pain level in the eleven ulcers. In group II, complete healing of 14 ulcers occurred in 14–60 days with a mean of 33.3±14.7; healing rate range was 0.064–2.22 and the mean 0.896±0.646 cm2/day. Healthy granulations were present in 13 ulcers (92.9%) and absent in one ulcer (7.1%). Three ulcers (21.4%) were of mild severity (grade 1 ulcers) while eleven ulcers (78.6%) were of moderate severity (grade 2 ulcers). The healing rate was faster in ulcers of mild severity (1.7±0.438 cm2/day) in comparison to ulcers of moderate severity (0.673±0.498 cm2/day). Eleven cases (78.6%) showed improvement in their pain level on a scale from 0 to 10. Conclusion AM graft can be of value in wound healing. Further studies are needed to confirm these findings. PMID:27390533

  7. Osteogenic differentiation of amniotic fluid mesenchymal stromal cells and their bone regeneration potential.

    PubMed

    Pipino, Caterina; Pandolfi, Assunta

    2015-05-26

    In orthopedics, tissue engineering approach using stem cells is a valid line of treatment for patients with bone defects. In this context, mesenchymal stromal cells of various origins have been extensively studied and continue to be a matter of debate. Although mesenchymal stromal cells from bone marrow are already clinically applied, recent evidence suggests that one may use mesenchymal stromal cells from extra-embryonic tissues, such as amniotic fluid, as an innovative and advantageous resource for bone regeneration. The use of cells from amniotic fluid does not raise ethical problems and provides a sufficient number of cells without invasive procedures. Furthermore, they do not develop into teratomas when transplanted, a consequence observed with pluripotent stem cells. In addition, their multipotent differentiation ability, low immunogenicity, and anti-inflammatory properties make them ideal candidates for bone regenerative medicine. We here present an overview of the features of amniotic fluid mesenchymal stromal cells and their potential in the osteogenic differentiation process. We have examined the papers actually available on this regard, with particular interest in the strategies applied to improve in vitro osteogenesis. Importantly, a detailed understanding of the behavior of amniotic fluid mesenchymal stromal cells and their osteogenic ability is desirable considering a feasible application in bone regenerative medicine.

  8. Amniotic fluid stem cells and their application in cell-based tissue regeneration.

    PubMed

    Baghaban Eslaminejad, Mohamadreza; Jahangir, Shahrbanoo

    2012-10-01

    Advances in stem cell biotechnology hold great promise in the field of tissue engineering and regenerative medicine. Of interest are marrow mesenchymal stem cells (MSCs), embryonic stem cells (ESCs), and induced pluripotent stem cells (iPSCs). In addition, amniotic fluid stem cells (AFSCs) have attracted attention as a viable choice following the search for an alternative stem cell source. Investigators are interested in these cells because they come from the amniotic fluid that is routinely discarded after birth. There have been multiple investigations conducted worldwide in an attempt to better understand AF-SCs in terms of their potential use in regenerative medicine. In this review we give a brief introduction of amniotic fluid followed by a description of the cells present within this fluid. Their history related to stem cell discovery in the amniotic fluid as well as the main characteristics of AF-SCs are discussed. Finally, we elaborate on the potential for these cells to promote regeneration of various tissue defects, including fetal tissue, the nervous system, heart, lungs, kidneys, bones, and cartilage.

  9. Use of quantitative amniotic fluid phosphatidylglycerol as a criterion for fetal lung maturation.

    PubMed

    Tsao, F H; Zachman, R D

    1992-01-01

    Phosphatidylglycerol (PG) in amniotic fluid was quantitatively measured by thin-layer chromatography (TLC) in 941 amniotic fluid samples and related to the presence or absence of respiratory distress syndrome (RDS) in neonates born within 7 days of the amniocentesis. In 639 case tests with PG 1 nmol/ml or more of amniotic fluid, there was no RDS. However, there were six cases of RDS associated with 109 tests of PG from 0 to less than 1 nmol/ml. These 109 tests would all have been reported as PG positive if PG were only qualitatively assessed on the TLC plate by the naked eye. The highest incidence of RDS occurred when PG was absent (23 RDS of 193 patients). Hence, this study suggests that quantitative analysis of PG determines a cutoff point of PG that eliminates false-positive PG assessments (1 nmol/ml in our laboratory). There was no difference in the levels of PG between males and females at equal gestational ages, but the incidence of RDS among male neonates was 2.4 times higher (p less than 0.05) than female neonates in the group with an immature amniotic fluid lecithin to sphingomyelin ratio and PG less than 1.0 nmol/ml. PMID:1550630

  10. Bacterial aetiological agents of intra-amniotic infections and preterm birth in pregnant women

    PubMed Central

    Mendz, George L.; Kaakoush, Nadeem O.; Quinlivan, Julie A.

    2013-01-01

    Infection-related preterm birth is a leading cause of infant mortality and morbidity; knowledge of bacterial populations invading the amniotic cavity and the routes of invasion is required to make progress in the prevention of preterm birth. Significant advances have been made in understanding bacterial communities in the vagina, but much less studied are intra-uterine bacterial populations during pregnancy. A systematic review of data published on the intra-uterine microbiome was performed; molecular information and summaries of species found in healthy individuals and in women with diagnosed infections served to construct a database and to analyse results to date. Thirteen studies fulfilled the review's inclusion criteria. The data of various investigations were collated, organized, and re-analyzed to achieve a more comprehensive understanding of microbial populations in the intra-amniotic space. The most common intra-amniotic bacterial taxa were species that can colonies the vagina in health and disease; there were others associated with the habitats of the mouth, gastrointestinal tract, and respiratory tract. The results suggest a central role for the ascending route of infections during pregnancy, and point to a possible secondary contribution via haematogenous invasion of the intra-amniotic space. The complete census of the intra-uterine microbiome awaits completion. PMID:24137568

  11. Intra-Amniotic Hemorrhage Imitating Gastroschisis: A Case Report and Review of the Literature

    PubMed Central

    Magann, Everett F.; Dinnel, Kinsey I.; Rabie, Nader Z.; Shoemaker, Amanda L.; Manning, Nirvana A.

    2016-01-01

    Patient: Female, 33 Final Diagnosis: Intramamniotic hemorrhage Symptoms: Abdominal pain • uterine contractions • vaginal bleeding Medication: — Clinical Procedure: Cesarean delivery Specialty: Obstetrics and Gynecology Objective: Unusual clinical course Background: A spontaneous intra-amniotic hemorrhage is rarely encountered during pregnancy. The correct diagnosis and management are problematic because of the infrequency of this condition and the high likelihood of a misdiagnosis. Case Report: A primigravida with an uncomplicated pregnancy and a normal targeted ultrasound presented late in the second trimester of pregnancy with antepartum bleeding of unknown origin. A repeat ultrasound was suggestive of an abdominal wall defect (gastroschisis). The patient continued to have antepartum bleeding and developed uterine contractions and abdominal pain necessitating frequent visits to labor and delivery. An MRI ruled out gastroschisis and diagnosed intra-amniotic hematoma. The patient presented with acute abdominal pain and was clinically considered to be having an abruption, and was delivered by cesarean. Old blood was noted in the abdominal cavity and within the uterine cavity. At the time of the cesarean, an area of intra-amniotic hematoma was identified, as well as a retroplacental blood clot. Conclusions: An intra-amniotic hematoma is unusual and may be misdiagnosed. MRI may be helpful in determining the correct diagnosis and subsequent management. PMID:27760979

  12. Osteogenic differentiation of amniotic fluid mesenchymal stromal cells and their bone regeneration potential

    PubMed Central

    Pipino, Caterina; Pandolfi, Assunta

    2015-01-01

    In orthopedics, tissue engineering approach using stem cells is a valid line of treatment for patients with bone defects. In this context, mesenchymal stromal cells of various origins have been extensively studied and continue to be a matter of debate. Although mesenchymal stromal cells from bone marrow are already clinically applied, recent evidence suggests that one may use mesenchymal stromal cells from extra-embryonic tissues, such as amniotic fluid, as an innovative and advantageous resource for bone regeneration. The use of cells from amniotic fluid does not raise ethical problems and provides a sufficient number of cells without invasive procedures. Furthermore, they do not develop into teratomas when transplanted, a consequence observed with pluripotent stem cells. In addition, their multipotent differentiation ability, low immunogenicity, and anti-inflammatory properties make them ideal candidates for bone regenerative medicine. We here present an overview of the features of amniotic fluid mesenchymal stromal cells and their potential in the osteogenic differentiation process. We have examined the papers actually available on this regard, with particular interest in the strategies applied to improve in vitro osteogenesis. Importantly, a detailed understanding of the behavior of amniotic fluid mesenchymal stromal cells and their osteogenic ability is desirable considering a feasible application in bone regenerative medicine. PMID:26029340

  13. Forensic aspects of post-mortem histological detection of amniotic fluid embolism.

    PubMed

    Sinicina, I; Pankratz, H; Bise, K; Matevossian, E

    2010-01-01

    Amniotic fluid embolism (AFE) continues to be one of the most feared and devastating complications of pregnancy. A reliable diagnosis can be made only upon histological examination. A detection of AFE every now and then has a relevant implication on medico-legal aspects of intrapartum or post-partum maternal death. However, there are only isolated reports in the literature concerning the detection interval of amniotic fluid elements after their transfer into the lungs. The objective of this study was to determine how long after the onset of clinical symptoms the elements of amniotic fluid may be detectable in the pulmonary circulation. An autopsy, as well as a histological and toxicological examination of 29 women, who died intrapartum or post-partum were performed. AFE was diagnosed in seven women (25%). The maximum survival time of the women with AFE and also the detection interval of AF in the pulmonary vasculature was 36 h. In the lungs of the women who did not die of AFE, amniotic fluid components were not found. Thus, there is no evidence for a physiologic occurrence of AFE. In women who die some days or even weeks after delivery as a consequence of a haemorrhagic shock following post-partum genital bleeding ensuing from uterine atony, A