Not all vascular smooth muscle cell exosomes calcify equally in chronic kidney disease
2018; Elsevier BV; Volume: 93; Issue: 2 Linguagem: Inglês
10.1016/j.kint.2017.08.036
ISSN1523-1755
AutoresAdriana Dusso, María Isabel Colombo, Catherine M. Shanahan,
Tópico(s)Systemic Sclerosis and Related Diseases
ResumoPrevention of medial calcification in patients with chronic kidney disease requires the maintenance of vascular smooth muscle cell fitness. To preserve viability under chronic kidney disease–induced stress, vascular smooth muscle cells increase exosome formation and release, but the result is aggravated pathological calcification. Now Chen et al. report that microvesicles from calcifying vascular smooth muscle cells may propagate procalcifying signals to normal vascular smooth muscle cells. To help design effective strategies to impair procalcifying cell-to-cell communication, this commentary updates the current understanding of the main regulators of microvesicle/exosome biogenesis and secretion. Prevention of medial calcification in patients with chronic kidney disease requires the maintenance of vascular smooth muscle cell fitness. To preserve viability under chronic kidney disease–induced stress, vascular smooth muscle cells increase exosome formation and release, but the result is aggravated pathological calcification. Now Chen et al. report that microvesicles from calcifying vascular smooth muscle cells may propagate procalcifying signals to normal vascular smooth muscle cells. To help design effective strategies to impair procalcifying cell-to-cell communication, this commentary updates the current understanding of the main regulators of microvesicle/exosome biogenesis and secretion. Vascular calcification is the pathological deposition of calcium phosphate salts in the vasculature and is a prevalent and serious complication in patients with chronic kidney disease (CKD), where it associates positively with a disproportionately high risk of cardiovascular mortality. In the course of CKD, vascular calcification develops early, affecting 25% of patients at CKD stages 3 to 4 and increasing to 50%–80% in patients starting hemodialysis (reviewed in Paloian and Giachelli1Paloian N.J. Giachelli C.M. A current understanding of vascular calcification in CKD.Am J Physiol Renal Physiol. 2014; 307: F891-F900Crossref PubMed Scopus (214) Google Scholar). In patients with CKD, calcification affects both the intimal and medial layers of the arterial wall. However, medial calcification, a process that resembles bone mineralization and is carried out by vascular smooth muscle cells (VSMCs), is more pronounced in CKD and is the exclusive form of vascular calcification in pediatric CKD (reviewed in Paloian and Giachelli1Paloian N.J. Giachelli C.M. A current understanding of vascular calcification in CKD.Am J Physiol Renal Physiol. 2014; 307: F891-F900Crossref PubMed Scopus (214) Google Scholar). Important for treatment, despite similar clinical and biochemical risk factors, medial calcification progresses faster in hemodialysis patients with existing calcified lesions compared with patients with noncalcified CKD. In this issue of Kidney International, Chen and collaborators (2018) examine the ability of microvesicles generated by calcifying VSMCs cultured from arteries of rats with CKD to extend procalcifying signals to adjacent VSMCs cultured from normal rats.2Chen N.X. O'Neill K.D. Moe S.M. Matrix vesicles induce calcification of recipient vascular smooth muscle cells through multiple signaling pathways.Kidney Int. 2018; 93: 343-354Abstract Full Text Full Text PDF PubMed Scopus (70) Google Scholar The rationale for the study was the recognized key role of exosomes in cell-to-cell communication and the recent identification of a critical contribution of exosome biosynthesis and release by VSMCs to vascular calcification.3Kapustin A.N. Chatrou M.L. Drozdov I. et al.Vascular smooth muscle cell calcification is mediated by regulated exosome secretion.Circ Res. 2015; 116: 1312-1323Crossref PubMed Scopus (330) Google Scholar The results of the study by Chen et al. have revealed a greater complexity of mechanisms regulating intracellular control and compartmentalization of mineralization by VSMCs. This commentary updates the current understanding and highlights the unanswered questions underlying microvesicle/exosome biogenesis and secretion from VSMCs. Ultimately, this knowledge will be essential to customize therapeutic strategies aimed at attenuating or delaying both the initiation of calcification and potentially the propagation of calcifying signals between VSMCs. Figure 1 summarizes the mechanisms for pathological calcium deposition. First, VSMCs undergo a process of phenotypic transition that involves the loss of their contractile phenotype, required to maintain vascular tone, together with the upregulation of markers of osteochondrogenesis. Simultaneously, VSMCs release matrix vesicles that colocalize with elastin and collagen fibrils and form the nidus for mineralization.1Paloian N.J. Giachelli C.M. A current understanding of vascular calcification in CKD.Am J Physiol Renal Physiol. 2014; 307: F891-F900Crossref PubMed Scopus (214) Google Scholar Recent characterization of the biogenesis of these calcifying matrix vesicles identified at least a subpopulation as exosomes, because the endosomal pathway and inward budding of the membrane of late endosomes or multivesicular bodies (MVB) participate in their formation,3Kapustin A.N. Chatrou M.L. Drozdov I. et al.Vascular smooth muscle cell calcification is mediated by regulated exosome secretion.Circ Res. 2015; 116: 1312-1323Crossref PubMed Scopus (330) Google Scholar as depicted in Figure 1. In addition, the role of sphingomyelin phosphodiesterase 3, also known as neutral sphingomyelinase 2 (nSM2), in the regulation of exosome release was shown. Furthermore, although elevated extracellular calcium and tumor necrosis factor α, common features in CKD, increased the expression of nSM2 and the secretion of calcifying exosomes, chemical inhibition of nSM2 prevented VSMC calcification.3Kapustin A.N. Chatrou M.L. Drozdov I. et al.Vascular smooth muscle cell calcification is mediated by regulated exosome secretion.Circ Res. 2015; 116: 1312-1323Crossref PubMed Scopus (330) Google Scholar These findings render the inhibition of nSM2 a novel strategy to attenuate calcification initiation. The severe bone mineralization defects in the nSM2 null mice strongly support the contribution of this mechanism to bone and vascular calcification. Intriguingly, Chen and coworkers2Chen N.X. O'Neill K.D. Moe S.M. Matrix vesicles induce calcification of recipient vascular smooth muscle cells through multiple signaling pathways.Kidney Int. 2018; 93: 343-354Abstract Full Text Full Text PDF PubMed Scopus (70) Google Scholar demonstrate multiple procalcifying signals induced exclusively by the so called "cellular-derived exosome-like vesicles" obtained from collagenase digestion of VSMCs cultured from arteries of CKD rats upon their endocytosis by cultured VSMCs from normal rats exposed to high phosphorus. In contrast, there were no calcifying signals from exosome-like vesicles freely released into the incubation media by identical cells in culture. The calcifying signals from these matrix-trapped cellular-derived exosomes included increased intracellular calcium and oxidative stress in recipient VSMCs. However, exposure to high phosphate had no effect on the calcifying potency of the cellular exosome-like vesicles in increasing intracellular calcium and oxidative stress. They also showed that activation of transient calcium rises by these exosome-like vesicles was dependent on mitogen-activated protein kinase signaling; however, this was independent of the activation of oxidative stress and osteogenic gene expression. Thus, the authors show a potential contribution for intercellular signaling in propagating calcifying signals, but the mechanisms driving this calcification potential remain unclear. In addition, a critical unanswered question is how these procalcifying signals from matrix-trapped cellular-derived exosome-like vesicles reach a neighboring normal VSMC. One mechanism could be uptake from the extracellular matrix by phagocytotic processes. Alternatively, nanotubes, bridging CKD-derived VSMCs and normal VSMCs, could contribute, although the authors indicated that nanotube formation was not observed in these studies. It is important to note that in the work by Chen et al. the vesicles were not fully characterized and therefore their composition as well as release and uptake pathways are difficult to determine. Both matrix-trapped and free-floating exosome-like vesicles were positive for CD63, but this is not necessarily indicative of an endosomal origin for both. Indeed, the main difference between these 2 vesicle populations was that the matrix-trapped population lacked fetuin-A, whereas the media-derived exosomes had a high fetuin content. These findings raise 2 important questions regarding the biogenesis of the 2 vesicle populations: (i) What is the origin of the fetuin in media-derived exosomes because VSMCs do not express fetuin? and (ii) Why do matrix-derived exosomes, generated under identical stimulation, lack fetuin? Most likely the presence of serum and exosomes in the incubation media could account for the fetuin content, but it fails to explain why extracellular fetuin was not incorporated into matrix-derived vesicles. Alternatively, the media-derived exosomes could represent a heterogenous population of exosomes, with some coisolated from serum exosomes during ultracentrifugation. An important message from these controversial findings is that the existing enthusiasm for the use of exosome content as an early estimate of procalcifying activity awaits the development of better methods for exosome purification and characterization (reviewed in Hessvik and Llorente4Hessvik NP, Llorente A. Current knowledge on exosome biogenesis and release [e-pub ahead of print]. Cell Mol Life Sci. https://doi.org/10.1007/s00018-017-2595-9. Accessed October 22, 2017.Google Scholar). For example, high levels of extracellular phosphate were shown to be inefficient in inducing the generation of calcifying exosome-like vesicles, yet in vivo high phosphate is likely to be important because it induces systemic inflammation, favoring the induction of nSM2 expression and calcifying exosome release.3Kapustin A.N. Chatrou M.L. Drozdov I. et al.Vascular smooth muscle cell calcification is mediated by regulated exosome secretion.Circ Res. 2015; 116: 1312-1323Crossref PubMed Scopus (330) Google Scholar Furthermore, high phosphate causes reductions in micro RNA 145 in VSMCs.5Rangrez A.Y. M'Baya-Moutoula E. Metzinger-Le Meuth V. et al.Inorganic phosphate accelerates the migration of vascular smooth muscle cells: evidence for the involvement of miR-223.PLoS One. 2012; 7: e47807Crossref PubMed Scopus (95) Google Scholar Micro RNA 145 is6Cordes K.R. Sheehy N.T. White M.P. et al.miR-145 and miR-143 regulate smooth muscle cell fate and plasticity.Nature. 2009; 460: 705-710Crossref PubMed Scopus (1291) Google Scholar essential for the maintenance of VSMC contractile phenotype, and significantly, micro RNA 145 is a suppressor of the expression of Rab27a, an essential component of the endosomal sorting machinery required for exosome release.7Tang L. Wei D. Yan F. MicroRNA-145 functions as a tumor suppressor by targeting matrix metalloproteinase 11 and Rab GTPase family 27a in triple-negative breast cancer.Cancer Gene Ther. 2016; 23: 258-265Crossref PubMed Scopus (28) Google Scholar Clearly, the degree of micro RNA 145 reduction induced by high serum phosphate and the resulting osteogenic differentiation of VSMCs could enhance Rab27a levels and exosome secretion in an nSM2-independent manner. An additional consideration from the work by Chen et al.2Chen N.X. O'Neill K.D. Moe S.M. Matrix vesicles induce calcification of recipient vascular smooth muscle cells through multiple signaling pathways.Kidney Int. 2018; 93: 343-354Abstract Full Text Full Text PDF PubMed Scopus (70) Google Scholar is the colocalization with lysosome markers of the "cellular-derived" exosome-like vesicles endocytozed by normal VSMCs cultured in high phosphorus conditions. This points to a potential contribution of the balance between autophagy and exosome release according to physiological or pathological stimuli. High phosphate induction of autophagy was shown to attenuate the secretion of calcifying microvesicles and overall calcification. Also, under conditions that stimulate autophagy such as starvation, rapamycin treatment, or LC3 overexpression, MVBs are directed to the autophagy pathway, reducing exosome release. However, recent studies suggest that autophagy can also stimulate the release of MVB contents, including adenosine triphosphate,8Fader C.M. Aguilera M.O. Colombo M.I. ATP is released from autophagic vesicles to the extracellular space in a VAMP7-dependent manner.Autophagy. 2012; 8: 1741-1756Crossref PubMed Scopus (62) Google Scholar which underscores the need for a better understanding of factors determining the balance between autophagy and exosome release, which may be regulated by the cellular metabolic state. Indeed, there is increasing evidence that both autophagy dysregulation and abnormal exosome secretion contribute to human disease (reviewed in Hessvik and Llorente4Hessvik NP, Llorente A. Current knowledge on exosome biogenesis and release [e-pub ahead of print]. Cell Mol Life Sci. https://doi.org/10.1007/s00018-017-2595-9. Accessed October 22, 2017.Google Scholar). Similar to VSMCs, in chondrocytes incubated in osteogenic conditions, elevations in cytosolic calcium were followed by accumulation of annexins A2, A5, and A6 in calcifying matrix vesicles. Interestingly, a very recent publication has shown that the release of annexin 2 is stimulated by interferon gamma–induced autophagy. Moreover, inhibition of annexin 2 release was hampered under conditions that prevented fusion of autophagosomes with MVBs and of MVBs with the cell membrane,9Chen Y.D. Fang Y.T. Cheng Y.L. et al.Exophagy of annexin A2 via RAB11, RAB8A and RAB27A in IFN-gamma-stimulated lung epithelial cells.Sci Rep. 2017; 7: 5676Crossref PubMed Scopus (61) Google Scholar such as Rab11, Rab8, and Rab27a knockdown. These recent results may also link autophagy with the release of calcifying matrix vesicles, although this hypothesis needs to be experimentally addressed. There may be additional mechanisms linking exosome release and authophagy.4Hessvik NP, Llorente A. Current knowledge on exosome biogenesis and release [e-pub ahead of print]. Cell Mol Life Sci. https://doi.org/10.1007/s00018-017-2595-9. Accessed October 22, 2017.Google Scholar For example, cells lacking 2 autophagic proteins Atg12-Atg3 show hampered exosome release, probably because of alterations in late endosomal function, whereas alterations in the function of lysosomes, using the proton pump inhibitor Bafilomycin A1, cause a change in exosome cargos. Additionally, cellular stress conditions, such as a dysfunctional lysosomal pathway, endoplasmic reticulum stress by tunicamycin treatment, hypoxia, or irradiation, lead to an increase in exosome secretion. These increases in the release of exosomes may be a way of eliminating unwanted products accumulated in the cells. However, it is also likely that cells subjected to stress communicate with adjacent cells via the release of microvesicles. Thus, it is expected that VSMCs from CKD respond to different environmental and intracellular stressors, releasing exosomes with distinct cargos and procalcifying activities. Understanding the complexity of these processes is the first step to optimize the control of exosome secretion and impair exosome-mediated cell-to-cell communication to safely minimize the risk of vascular calcification without an adverse impact on bone. All the authors declared no competing interests. AD's research is supported by grants from Plan Estatal de I+D+I 2013-2017, Instituto de Salud Carlos III (ISCIII) Fondo Europeo de Desarrollo Regional (FEDER)(PI14/01452), Plan de Ciencia, Tecnología e Innovación 2013-2017 del Principado de Asturias (GRUPIN14-028), Fundación para el Fomento en Asturias de la Investigación Científica Aplicada a la Tecnología (FICYT), Red de Investigación Renal-REDinREN from ISCIII (RD06/0016/1013, RD12/0021/1023), and Sociedad Asturiana Fomento Investigaciones Metabólicas (SAFIM). MIC's work in related areas is supported by grants PICT 2013-0305 (Agencia) and Sectyp (U.N.CUYO). Matrix vesicles induce calcification of recipient vascular smooth muscle cells through multiple signaling pathwaysKidney InternationalVol. 93Issue 2PreviewIn patients with chronic kidney and end-stage renal diseases, the major risk factor for progression of arterial calcification is the presence of existing (baseline) calcification. Here, we tested whether calcification of arteries is extended from calcified vascular smooth muscle cells (VSMCs) to adjacent normal cells by matrix vesicle–induced alteration of cell signaling. Matrix vesicles isolated from VSMC of rats with chronic kidney disease were co-cultured with VSMCs from normal littermates. Endocytosis of vesicles by recipient cells was confirmed by confocal microscopy. Full-Text PDF Open Archive
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