Pazopanib attenuated bleomycin-induced pulmonary fibrosis via suppressing TGF-ß1 signaling pathway.

Background: Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive interstitial lung disease with a high mortality rate and limited treatment efficacy. Nintedanib, a tyrosine kinase inhibitor, is clinically used to treat pulmonary fibrosis. At present, only nintedanib is on the market for the treatment of pulmonary fibrosis. Pazopanib is a drug for the treatment of renal cell carcinoma and advanced soft tissue sarcoma. Methods: In this study, we explored whether pazopanib can attenuate bleomycin (BLM)-induced pulmonary fibrosis and explored its antifibrotic mechanism. In vivo and in vitro investigations were carried out to investigate the efficacy and mechanism of action of pazopanib in pulmonary fibrosis. Results: In vivo experiments showed that pazopanib can alleviate pulmonary fibrosis caused by BLM, reduce the degree of collagen deposition and improve lung function. In vitro experiments showed that pazopanib suppressed transforming growth factor-ß1 (TGF-ß1)-induced myofibroblast activation and promoted apoptosis and autophagy in myofibroblasts. Further mechanistic studies demonstrated that pazopanib inhibited the TGF-ß1/Smad and non-Smad signaling pathways during fibroblast activation. Conclusions: In conclusion, pazopanib attenuated BLM-induced pulmonary fibrosis by suppressing the TGF-ß1 signaling pathway. Pazopanib inhibits myofibroblast activation, migration, autophagy, apoptosis, and extracellular matrix (ECM) buildup by downregulating the TGF-ß1/Smad signal route and the TGF-ß1/non-Smad signal pathway. It has the same target as nintedanib and is a tyrosine kinase inhibitor.

Design of Surface Plasmon Resonance-Based D-Type Double Open-Loop Channels PCF for Temperature Sensing.

Here, we document a D-type double open-loop channel floor plasmon resonance (SPR) photonic crystal fiber (PCF) for temperature sensing. The grooves are designed on the polished surfaces of the pinnacle and backside of the PCF and covered with a gold (Au) film, and stomata are distributed around the PCF core in a progressive, periodic arrangement. Two air holes between the Au membrane and the PCF core are designed to shape a leakage window, which no longer solely averts the outward diffusion of Y-polarized (Y-POL) core mode energy, but also sets off its coupling with the Au movie from the leakage window. This SPR-PCF sensor uses the temperature-sensitive property of Polydimethylsiloxane (PDMS) to reap the motive of temperature sensing. Our lookup effects point out that these SPR-PCF sensors have a temperature sensitivity of up to 3757 pm/°C when the temperature varies from 5 °C to 45 °C. In addition, the maximum refractive index sensitivity (RIS) of the SPR-PCF sensor is as excessive as 4847 nm/RIU. These proposed SPR-PCF temperature sensors have an easy nanostructure and proper sensing performance, which now not solely improve the overall sensing performance of small-diameter fiber optic temperature sensors, but also have vast application prospects in geo-logical exploration, biological monitoring, and meteorological prediction due to their remarkable RIS and exclusive nanostructure.

Rho-family small GTPases are involved in forskolin-induced cell-cell contact formation of renal glomerular podocytes in vitro.

Intercellular adhesions between renal glomerular epithelial cells (also called podocytes) are necessary for the proper function of the glomerular filtration barrier. Although our knowledge of the molecular composition of podocyte cell-cell contact sites has greatly progressed, the underlying molecular mechanism regulating the formation of these cell-cell contacts remains largely unknown. We have used forskolin, an activator of adenylyl cyclase that elevates the level of intracellular cAMP, to investigate the effect of cAMP and three Rho-family small GTPases (RhoA, Cdc42, and Rac1) on the regulation of cell-cell contact formation in a murine podocyte cell line. Transmission electron microscopy and the immunostaining of cell adhesion molecules and actin-associated proteins have revealed a structural change at the site of cell-cell contact following forskolin treatment. The activity of the Rho-family small GTPases before and after forskolin treatment has been evaluated with a glutathione-S-transferase pull-down assay. Forskolin reinforces the integrity of cell-cell contacts, resulting in the closure of an intercellular adhesion zipper, accompanied by a redistribution of cell adhesion molecules and actin-associated proteins in a continuous linear pattern at cell-cell contacts. The Rho-family small GTPases Rac1 and Cdc42 are activated during closure of the adhesion zipper, whereas RhoA is suppressed. Thus, cAMP promotes the assembly of cell-cell contacts between podocytes via a mechanism that probably involves Rho-family small GTPases.

In vitro assays for adhesion and migration of osteoblastic cells (Saos-2) on titanium surfaces.

The first event occurring at the boundary between a metal implant and living tissue is the attachment of cells onto the metal surface of the implant. The attachment characteristics of the metal in this situation are critical in determining its biocompatibility and usefulness as artificial bone and tooth implants. Using the human osteosarcoma cell line Saos-2, we attempted to establish simple and reliable methods for evaluating the attachment of cultured osteoblastic cells onto titanium samples that had been subjected to various surface treatments. Fluorescence actin imaging showed that cells cultured on titanium with hydrofluoric acid etching (HF-Ti) exhibited delayed spreading of their cytoplasm, as compared to cells cultured for the same length of time on nitrided titanium or physically polished titanium. The HF-Ti-cultured cells also exhibited poor assembly of focal contacts, as visualized by vinculin immunofluorescence. Furthermore, in motility assays based on an in vitro wound model, cells cultured on HF-Ti migrated more slowly than cells cultured on other titanium surfaces. These data suggest that Saos-2 cells attach less effectively to the HF-Ti surface. The methods described in this study should be useful for assessing the initial interactions of cultured cells with various materials, including metals.

Phylogenetic investigation of Dogiel's pericellular nests and Cajal's initial glomeruli in the dorsal root ganglion.

Cajal's initial glomeruli (IG) and Dogiel's pericellular nests (PCNs) were first described from methylene blue preparations of healthy animal tissues around the beginning of the last century. Since that time, although many reports have been published concerning these structures, few have focused on their development and phylogeny in healthy animals. The aim of this study was to examine the phylogenetic development of the sensory neurons in Cajal's IG (also called axonal glomeruli) and Dogiel's PCNs in the dorsal root ganglion (DRG) of the healthy adult frog, chick, rat, and rabbit. The three-dimensional architecture of the neurons was observed in ganglia by scanning electron microscopy after removal of the connective tissue. The neurons in the DRG of fish are known to be bipolar, but DRG neurons in the species examined here were found to be pseudounipolar, with single stem processes. The proportion of neurons having IG or PCNs increased with increasing phylogenetic complexity in the species examined here. Cajal's initial glomeruli, the convolution of the stem process near the parent cell body: In frogs, the ganglia were small and the neuronal stem processes were very short and straight. In chicks, the stem processes were longer; sometimes very long, tortuous processes were observed. However, no neurons with typical IG were observed in either species. Typical IG were observed in rats and rabbits; their occurrence was much more frequent in rabbits. Pseudounipolarization, i.e., the transition from bipolar to pseudounipolar neurons, is thought to save space, limit the length of neuronal processes, and reduce conduction time. However, an explanation of the evolutionary advantage of the IG, which is formed by the excessive prolongation of the stem process, remains elusive. The cytological and electrophysiological importance of IG has been discussed. Dogiel's pericellular nests (PCNs), which resemble balls of yarn made of thin unmyelinated nerve fibers around DRG neurons, have been observed in the DRG of rats and rabbits, but not in frogs or chicks. This interesting structure shows not only ontogenetic development in healthy animals but also phylogenetic development among species. The nerve fibers in the PCNs were less than 1.2 mum in diameter and had some varicosities. An immunohistochemical study using anti-tyrosine hydroxylase (TH) antibody revealed that some PCNs contain TH-positive nerve fibers and varicosities. Such TH-positive PCNs disappear after sympathectomy. These results suggest that the PCNs are made up of autonomic nerve fibers.

Transient increase of TUNEL-positive cells on postnatal day 20 in the developing rat olfactory bulb.

In the developing central nervous system, apoptosis plays an important role in the normal organization of the neuronal circuit. The timing of neurogenesis, proliferation, and migration of the neurons in the developing olfactory bulb (OB) is well studied; however, the involvement of apoptosis in this process is not fully understood. In this study, we examined the changes in the distribution and the number of apoptotic cells in the rat OB during embryonic and postnatal periods, by using terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end-labeling (TUNEL) staining. Although the number of TUNEL-positive cells was relatively small during the embryonic period, it gradually increased after birth, and peaked on postnatal day 20 with statistical significance, especially in the granule cell layer of the main OB. This transient increase of TUNEL-positive cells on postnatal day 20 may be involved in a critical event during maturation of the OB.

Rho-ROCK signal pathway regulates microtubule-based process formation of cultured podocytes--inhibition of ROCK promoted process elongation.

BACKGROUND: Podocytes, renal glomerular visceral epithelial cells, have two kinds of processes, namely major processes containing microtubules (MTs) and foot processes with actin filaments (AFs). The present study investigated how MTs are organized by the Rho-ROCK signal transduction pathway during process formation of podocytes. METHOD: After induction of differentiation, podocytes of the conditionally immortalized mouse cell line were treated with Y-27632, a specific inhibitor of ROCK, and exoenzyme C3, an inhibitor of RhoA, as well as with forskolin whose effects include inhibition of RhoA, in order to inhibit the Rho-ROCK pathway. RESULTS: Inhibition of ROCK significantly enhanced the formation of thick processes containing MT bundles. Y-27632 promoted process formation even in the presence of latrunculin A which disrupts AFs, strongly suggesting that ROCK directly regulates MT assembly. Treatment with Y-27632 increased MT stability, and stabilized MTs preferentially localized in podocyte processes. Moreover, when treated with a combination of Y-27632 and forskolin, and with Y-27632 and C3 as well, podocytes developed not only MT-based thick processes but also AF-based thin projections. CONCLUSIONS: These data indicate a contribution of ROCK in MT organization to promote podocyte process formation, although it was originally thought to regulate AF assembly. AF-based thin projections seem to be induced mainly by inhibition of RhoA and ROCK. The present study reveals a significant role of the Rho-ROCK signal pathway in the reorganization of both MTs and AFs during process formation of podocytes.

Process formation of the renal glomerular podocyte: is there common molecular machinery for processes of podocytes and neurons?

The renal glomerular podocyte exhibits a highly arborized morphology. In comparison with the neuron, which is the best studied process-bearing cell, the podocyte major processes share many cell biological characteristics with neuronal dendrites. Both podocytes and neurons develop microtubule-based thick processes with branching morphology and both have thin actin-based projections (i.e. podocyte foot processes and dendritic spines). Formation of podocyte processes and neuronal dendrites depends on the assembly of microtubules. Because the assembly of microtubules is regulated by phosphorylation of microtubule-associated proteins, inhibition of protein phosphatases abolishes and inhibition of protein kinases promotes process formation. Podocytes and dendrites also share the machinery of intracellular traffic of membranous vesicles, as well as cytoskeletal elements, which is indispensable for the elongation of these processes. Furthermore, these two cell types share expression of various molecules working for signal transduction, transmembranous transport and intercellular contacts. Such common gene expression implies a similar transcriptional regulation in these cells. Concerning the formation of podocyte foot processes and dendritic branches, actin filaments are thought to play a central role in orchestrating the function of various molecules and the regulation of actin assembly is necessary to establish and maintain such sophisticated cellular architecture. The molecular mechanism of foot process formation seems to include Rho family small GTP-binding proteins, which are known to be responsible for the establishment of dendritic branching morphology.