The Use of Atomic Force Microscopy in Comprehensive Assessment of the "Mother-Placenta-Fetus" System in Obstetric and Endocrine Pathology during Pregnancy.

Atomic force microscopy is not very popular in practical health care, therefore, its potential is not studied enough, for example, in obstetrics when studying the "mother-placenta-fetus" system. Our study summarizes the possibilities of using atomic force microscopy for detection of various circulatory disorders and vascular changes at the microscopic level in the uterus (endometrium and myometrium), placenta, and umbilical cord in the main variants of obstetric and endocrine pathology. For instance, in the case of endocrine pathologies, changes in the form of stasis, sludge, diapedesis, ischemia, destruction and separation of endotheliocytes in villous blood vessels were found in the mother. The oxygen content in erythrocytes also naturally decreased in pathologies; poikilo- and anisocytosis were observed.

Umbilical cord plasma-derived exosomes from preeclamptic women induce vascular dysfunction by targeting HMGCS1 in endothelial cells.

Hypertension is one of the major clinical manifestations of preeclampsia. Vascular dysfunction is crucial for the occurrence and progression of hypertension. Exosomes are emerging as mediators of intercellular communication and can participate in angiogenesis. In this study, we hypothesize that umbilical cord plasma-derived exosomes from preeclamptic women (PE-uexo) impair vascular development by regulating endothelial cells. Here, umbilical cord plasma samples from women with normal pregnancies and matched preeclamptic patients were used to isolate circulating exosomes. Proliferation, Transwell and tube formation assays indicated that PE-uexo impaired the angiogenesis of human umbilical vein endothelial cells (HUVECs). On the basis of microarray analysis of HUVECs treated with PE-uexo or exosomes from women with normal pregnancies, we showed that the expression of 3-hydroxy-3-methylglutaryl-CoA synthase 1 (HMGCS1) was decreased in the PE-uexo-treated HUVECs. Furthermore, downregulation of HMGCS1 in HUVECs attenuated the proliferation and migration of these cells. Interestingly, HMGCS1 was decreased in P0 HUVECs from preeclamptic pregnancies compared with normotensive pregnancies. Together, these observations suggest that PE-uexo disrupts normal function in vascular endothelial cells by targeting HMGCS1, which may result in vascular disorders in the offspring.

Towards biobanking technologies for natural and bioengineered multicellular placental constructs.

Clinical application of a large variety of biomaterials is limited by the imperfections in storage technology. Perspective approaches utilizing low-temperature storage are especially challenging for multicellular structures, such as tissues, organs, and bioengineered constructs. Placenta, as a temporary organ, is a widely available unique biological material, being among the most promising sources of various cells and tissues for clinical and experimental use in regenerative medicine and tissue engineering. The aim of this study was to analyse the mechanisms of cryoinjuries in different placental tissues and bioengineered constructs as well as to support the viability after low temperature storage, which would contribute to development of efficient biobanking technologies. This study shows that specificity of cryodamage depends on the structure of the studied object, intercellular bonds, as well as interaction of its components with cryoprotective agents. Remarkably, it was possible to efficiently isolate cells after thawing from all of the studied tissues. While the outcome was lower in comparison to the native non-frozen samples, the phenotype and expression levels of pluripotency genes remained unaffected. Further progress in eliminating of recrystallization processes during thawing would significantly improve biobanking technologies for multicellular constructs and tissues.

Clasificación morfológica de células endoteliales de venas del cordón umbilical humano en imágenes digitales de cultivos in vitro / Morphological classification of endothelial cells of veins from the human umbilical cord in digital images of in vitro cultures

Se realizó un estudio descriptivo y transversal desde julio hasta octubre de 2017, por especialistas de la Universidad de Oriente y de la Universidad de Sao Paulo, Brasil, para analizar desde el punto de vista morfológico células endoteliales de venas del cordón umbilical humano, presentes en imágenes digitales de cultivos in vitro 2D, tratadas con la ß2GPI. . Se propuso la clasificación supervisada celular considerando 3 clases: circulares, deformadas alargadas y deformadas poco alargadas, según los coeficientes de formas elíptico y circular, todo lo cual permitió identificar formas celulares relevantes. Para comparar los resultados de las muestras de control y las tratadas, se calcularon los intervalos de confianza para cada una de las clases, con un nivel de confianza de 95 por ciento. Se concluye que el análisis de las alteraciones morfológicas in vitro puede ser utilizada en cultivos 2D precoces (de 24 y 48 horas) para la cuantificación de la angiogénesis

A descriptive and cross-sectional study was carried out from July to October, 2017, by specialists of the Oriente University and the University of Sao Paulo, Brazil, to analyze from the morphological point of view endothelial cells of the human umbilical cord veins, which were present in digital images of 2D in vitro cultures, treated with the ß2GPI. The cellular supervised classification was proposed considering 3 classes: circular, distorted elongated and distorted not very elongated, according to the coefficients of elliptic and circular shapes, all that allowed to identify outstanding cellular forms. To compare the results of the control and treated samples, the intervals of confidence were calculated for each of the classes, with a 95 percent level of confidence. It was concluded that the analysis of the morphological disorders in vitro can be used in early 2D cultures (24 and 48 hours) for the quantification of the angiogenesis

Multiple intracerebroventricular injections of human umbilical cord mesenchymal stem cells delay motor neurons loss but not disease progression of SOD1G93A mice.

Stem cell therapy is considered a promising approach in the treatment of amyotrophic lateral sclerosis (ALS) and mesenchymal stem cells (MSCs) seem to be the most effective in ALS animal models. The umbilical cord (UC) is a source of highly proliferating fetal MSCs, more easily collectable than other MSCs. Recently we demonstrated that human (h) UC-MSCs, double labeled with fluorescent nanoparticles and Hoechst-33258 and transplanted intracerebroventricularly (ICV) into SOD1G93A transgenic mice, partially migrated into the spinal cord after a single injection. This prompted us to assess the effect of repeated ICV injections of hUC-MSCs on disease progression in SOD1G93A mice. Although no transplanted cells migrated to the spinal cord, a partial but significant protection of motor neurons (MNs) was found in the lumbar spinal cord of hUC-MSCs-treated SOD1G93A mice, accompanied by a shift from a pro-inflammatory (IL-6, IL-1ß) to anti-inflammatory (IL-4, IL-10) and neuroprotective (IGF-1) environment in the lumbar spinal cord, probably linked to the activation of p-Akt survival pathway in both motor neurons and reactive astrocytes. However, this treatment neither prevented the muscle denervation nor delayed the disease progression of mice, emphasizing the growing evidence that protecting the motor neuron perikarya is not sufficient to delay the ALS progression.

Lactation-Related MicroRNA Expression in Microvesicles of Human Umbilical Cord Blood.

BACKGROUND The complex process by which lactation is initiated upon neonate delivery remains incompletely understood. Microvesicles (MVs) can transmit microRNAs (miRNAs) into recipient cells to influence cell function, and recent studies have identified miRNAs essential for mammary gland development and lactation. This study aimed to investigate the expression of lactation-related miRNAs in MVs isolated from human umbilical cord blood immediately after delivery. MATERIAL AND METHODS Umbilical cord blood samples were collected from 70 healthy pregnant women, and MVs were isolated through differential centrifugation and characterized by transmission electron microscopy, Western blotting, and nanoparticle tracking analysis. Lactation-related miRNAs were screened using bioinformatics tools for miRNA target prediction, gene ontology, and signaling pathway analyses. miRNA PCR arrays were used for miRNA expression analysis, and the results were validated by real-time PCR. Upon exposure of HBL-100 human mammary epithelial cells to MVs, MV uptake was examined by fluorescence confocal microscopy and b-casein secretion was detected by ELISA. RESULTS Spherical MVs extracted from umbilical cord blood expressed CD63 and had an average diameter of 167.0±77.1 nm. We profiled 337 miRNAs in human umbilical cord blood MVs and found that 85 were related to lactation by bioinformatics analysis. The 25 most differentially expressed lactation-related miRNAs were validated by real-time PCR. MV uptake by HBL-100 cells was after 4 h in culture, and significantly increased secretion of ß-casein was observed after 96 h from cells exposed to MVs (P<0.05). CONCLUSIONS Umbilical cord blood MVs contain many lactation-related miRNAs and can induce ß-casein production by HBL-100 cells in vitro. Thus, umbilical cord blood MVs may mediate secretion of ß-casein through miRNAs, thereby playing an important role in fetal-maternal crosstalk.

Explants-isolated human placenta and umbilical cord cells share characteristics of both epithelial and mesenchymal stem cells.

In recent years, identification of new sources of adult stem cells developed rapidly, pursuing to find easily available tissues, which will give rise to homogenous stem cells populations. Up to present, bone marrow-derived mesenchymal stem cells (BM-MSCs) are unanimously considered to fulfill the criteria for being used in clinical settings, but adipose stem cells, placental and umbilical cord stem cells, and other tissue-derived stem cells are making their way to being used at least in autologous transplantation. We isolated cellular populations from placental tissue and umbilical cord using the explants method. The placental (PL) and umbilical cord (UC)-derived cells were cultured and expanded in appropriate conditions for generation of stem cells. We assessed the stemness characteristics of the tissue-isolated cells and compared them to an established MSCs line. For this purpose, we determined the immunophenotype, morphological and ultrastructural characteristics, as well as functional abilities of PL- and UC-derived cells. Flow cytometric evaluation of cells revealed presence of CD90, CD73, and CD105 stem cells markers, while the cells were negative for CD34, CD45 and HLA-DR. Immunocytochemical staining showed that 100% of PL- and UC-derived cells are positive for vimentin and CD105 expression, while cytokeratin was revealed in less than 10% in both tissue-isolated cells. Morphological and ultrastructural characteristics of cells exposed analogous cellular size and intracellular organization, similar to MSCs, but detailed view of UC-derived cells by transmission electron microscopy (TEM) demonstrated presence of intercellular junctions-desmosomes, similar to epithelial cells. Both PL- and UC-derived cells confirmed their trilineage potential, being able to differentiate into adipocytes, osteoblasts, and chondrocytes in different proportions. Flow chamber in vitro assay was used to determine to what extent PL- and UC-derived cells are able to adhere to substrates (VCAM and ICAM) and we showed progressively decreased adhesion of both cellular types, inversely proportional to the generated shear stress. We may conclude that explants-isolated placental and umbilical cord cells are endowed with characteristics of both epithelial and mesenchymal stem cells, and purification procedures are additionally required for safe use of these cells in diverse clinical applications.

Expression of PECAM-1 and E-Cadherin in the Umblical Cords of Gestational Diabetic Mothers / Expresión de PECAM-1 y E-cadherina en los Cordones Umbilicales de Madres con Diabetes Gestacional

The purpose of this study is to examine the changes in the umbilical cord in women diagnosed with gestational diabetes mellitus In this study, as a control group human placental tissues from normotensive pregnancies was collected from diabetic women at 28­35 weeks of gestation. Gestational diabetes (n= 20) and normal umbilical cord (n= 20) for a total of 40 units were received.GDM groups compared to the control group was significantly higher values was detected (p<0.01). In GDM group, light microscopy showed erosion of the endothelium and complete rupture of theumbilicalvessels resulting in extravasation of blood within Wharton's jelly. it was observed that the cytoplasmic fragments and cell infiltration of the spill to the subepithelial layer of apoptotic cell PECAM-1 positive reaction showed. E-Cadherin in endothelial side surface of diabetes group showed weak expression in the nucleus and showed positive reaction in smooth muscle.

El objetivo fue examinar los cambios que presenta el cordón umbilical de mujeres con diagnóstico de diabetes mellitus gestacional (DMG). Se incluyeron en el grupo control muestras de tejidos placentarios humanos de embarazos normotensos y de mujeres diabéticas de entre 28­35 semanas de gestación. Las muestras se divieron en cordones umbilicales con cambios de DMG (n= 20) y cordones umbilicales normales (n= 20), constituyendo un total de 40 muestras. El grupo de DMG, en comparación con el grupo control, presentó valores significativamente más elevados (p<0,01). En el grupo de DMG, la microscopía óptica demostró la erosión del endotelio y la ruptura completa de los vasos umbilicales, resultando en la extravasación de sangre dentro de la gelatina . Se observaron fragmentos citoplasmáticos e infiltración celular de la capa subepitelial de células apoptóticas mostró una reacción positiva a PECAM-1. En el grupo de DMG, la E-cadherina de la superficie lateral endotelial mostró una expresión débil en el núcleo y una reacción positiva en el músculo liso.

Transdifferentiation of Umbilical Cord-Derived Mesenchymal Stem Cells Into Epidermal-Like Cells by the Mimicking Skin Microenvironment.

Human umbilical cord-derived mesenchymal stem cells (UC-MSCs) are multipotent, primitive, and have been widely used for skin tissue engineering. Their transdifferentiation is determined by the local microenvironment. In this study, we investigated the potential epidermal differentiation of UC-MSCs and the formation of epidermis substitutes in a 3-dimensional (3D) microenvironment, which was fabricated by UC-MSCs embedded into collagen-chitosan scaffolds (CCSs) combined with an air-liquid interface (ALI) culture system. Using fluorescence microscope, we observed that UC-MSCs were spindle-shaped and evenly distributed in the scaffold. Methyl thiazolyl blue tetrazolium bromide assay and Live/Dead assay indicated that the CCSs have good biocompatibility with UC-MSCs. Immunohistochemistry and western blotting assay showed that UC-MSCs on the surface of the CCSs were positive for the epidermal markers cytokeratin 19 and involucrin at 14 days. In addition, hematoxylin-eosin staining indicated that multilayered epidermis substitutes were established. The constructed epidermis substitutes were applied to treat full-thickness wounds in rats and proved to promote wound healing. In conclusion, manipulating the 3D microenvironment is a novel method for inducing the epidermal differentiation of MSCs to engineer epidermal substitutes, which provides an alternative strategy for skin tissue engineering.

Allantoic, vitelline, and "hybrid" embryonic vestiges of the umbilical cord: an immunohistochemical study.

The embryonic origin of umbilical cord vestiges is well documented; however, their immunophenotype is unknown. This study was conducted to determine whether vitelline and allantoic remnants can be differentiated using immunohistochemical markers. All allantoic remnants were stained with p63 and were negative for CDX2, whereas the vitelline remnants stained with CDX2 and were negative for p63. An unexpected finding was a small number of morphologically ambiguous cases that stained with both markers in a complimentary manner. The term "hybrid" remnant is proposed for these remnants.