Krüppel-like factor 6 participates in extravillous trophoblast cell differentiation and its expression is reduced in abnormally invasive placenta.

Trophoblast cell differentiation is of paramount importance for successful pregnancy. Krüppel-like factor 6 (KLF6), a transcription factor with diverse roles in cell physiology and tumor biology, is required for trophoblast differentiation through the syncytial pathway. Herein, we demonstrate that extravillous trophoblast (EVT) cell migration and mesenchymal phenotype are increased upon KLF6 downregulation or the expression of a deletion mutant lacking its transcriptional regulatory domain (KΔac). Raman spectroscopy revealed molecular modifications compatible with increased differentiation in cells stably expressing the KΔac mutant. Moreover, abnormally invasive placenta showed lower KLF6 immunostaining compared with the normal placenta. Thus, impaired KLF6 expression or function stimulates EVT migration and differentiation in vitro and may contribute to the physiopathology of the abnormally invasive placenta.

Tuning the photoluminescence by engineering surface states/size of S, N co-doped carbon dots for cellular imaging applications.

In this research, we have synthesized carbon dots (CDs) co-doped with nitrogen and sulfur by facile hydrothermal method, using citric acid and cysteine as carbon source. The effect of solid-state thermic treatment (STT) at 303-453 K on the size, surface, fluorescence and cellular cytotoxicity of the CDs were systematically investigated. Through a simple STT, it was possible to tune surface states and the average size of the CDs, causing a permanent red shift. Initially, CDs showed a decrease in cell viability with increasing concentration. However, after STT, its viability remained constant with an increase in concentration. Here, we show the possibility to label the cells cytoplasm according to the CDs fluorescence emission before (blue emission) and after STT (red emission). The CDs studied in this paper show selective luminescence properties, which are fundamental for any cell imaging application.

Krüppel-like factor 6 (KLF6) requires its amino terminal domain to promote villous trophoblast cell fusion.

INTRODUCTION: Villous cytotrophoblast (vCTB) cells fuse to generate and maintain the syncytiotrophoblast layer required for placental development and function. Krüppel-like factor 6 (KLF6) is a ubiquitous transcription factor with an N-terminal acidic transactivation domain and a C-terminal zinc finger DNA-binding domain. KLF6 is highly expressed in placenta, and it is required for proper placental development. We have demonstrated that KLF6 is necessary for cell fusion in human primary vCTBs, and in the BeWo cell line. MATERIALS AND METHODS: Full length KLF6 or a mutant lacking its N-terminal domain were expressed in BeWo cells or in primary vCTB cells isolated from human term placentas. Cell fusion, gene and protein expression, and cell proliferation were analyzed. Moreover, Raman spectroscopy and atomic force microscopy (AFM) were used to identify biochemical, topography, and elasticity cellular modifications. RESULTS: The increase in KLF6, but not the expression of its deleted mutant, is sufficient to trigger cell fusion and to raise the expression of ß-hCG, syncytin-1, the chaperone protein 78 regulated by glucose (GRP78), the ATP Binding Cassette Subfamily G Member 2 (ABCG2), and Galectin-1 (Gal-1), all molecules involved in vCTB differentiation. Raman and AFM analysis revealed that KLF6 reduces NADH level and increases cell Young's modulus. KLF6-induced differentiation correlates with p21 upregulation and decreased cell proliferation. Remarkable, p21 silencing reduces cell fusion triggered by KLF6 and the KLF6 mutant impairs syncytialization and decreases syncytin-1 and ß-hCG expression. DISCUSSION: KLF6 induces syncytialization through a mechanism that involves its regulatory transcriptional domain in a p21-dependent manner.

Flavonoids induce cell death in Leishmania amazonensis: in vitro characterization by flow cytometry and Raman spectroscopy.

Leishmaniasis comprises a group of infectious diseases with worldwide distribution, of which both the visceral and cutaneous forms are caused by Leishmania parasites. In the absence of vaccines, efficacious chemotherapy remains the basis for leishmaniasis control. The available drugs are expensive and associated with several secondary adverse effects. Due to these limitations, the development of new antileishmanial compounds is imperative, and plants offer various perspectives in this regard. The present study evaluated the in vitro leishmanicidal activity of flavonoids isolated from Solanum paludosum Moric. and investigated the mechanisms of cell death induced by them. These compounds were evaluated in vitro for their antileishmanial activity against Leishmania amazonensis promastigotes and they showed prominent leishmanicidal activity. The EtOAc fraction, gossypetin 3,7,8,4'-tetra-O-methyl ether (1), and kaempferol 3,7-di-O-methyl ether (3) were selected to be used in an in vitro assay against L. amazonensis amastigotes and cell death assays. The flavonoids (1) and (3) presented significant activity against L. amazonensis amastigotes, exhibiting the IC50 values of 23.3 ± 4.5 µM, 34.0 ± 9.6 µM, and 10.5 ± 2.5 µM for the EtOAc fraction, (1), and (3), respectively, without toxic effects to the host cells. Moreover, (1) and (3) induced blocked cell cycle progression at the G1/S transition, ultimately leading to G1/G0 arrest. Flavonoid (3) also induced autophagy. Using Raman spectroscopy in conjunction with principal component analysis, the biochemical changes in the cellular components induced by flavonoids (1) and (3) were presented. The obtained results indicated that the mechanisms of action of (1) and (3) occurred through different routes. The results support that the flavonoids derived from S. paludosum can become lead molecules for the design of antileishmanial prototypes.

Macrophage adhesion on fibronectin evokes an increase in the elastic property of the cell membrane and cytoskeleton: an atomic force microscopy study.

Interactions between cells and microenvironments are essential to cellular functions such as survival, exocytosis and differentiation. Cell adhesion to the extracellular matrix (ECM) evokes a variety of biophysical changes in cellular organization, including modification of the cytoskeleton and plasma membrane. In fact, the cytoskeleton and plasma membrane are structures that mediate adherent contacts with the ECM; therefore, they are closely correlated. Considering that the mechanical properties of the cell could be affected by cell adhesion-induced changes in the cytoskeleton, the purpose of this study was to investigate the influence of the ECM on the elastic properties of fixed macrophage cells using atomic force microscopy. The results showed that there was an increase (~50%) in the Young's modulus of macrophages adhered to an ECM-coated substrate as compared with an uncoated glass substrate. In addition, cytochalasin D-treated cells had a 1.8-fold reduction of the Young's modulus of the cells, indicating the contribution of the actin cytoskeleton to the elastic properties of the cell. Our findings show that cell adhesion influences the mechanical properties of the plasma membrane, providing new information toward understanding the influence of the ECM on elastic alterations of macrophage cell membranes.