| Fibrosis is a leading cause of morbidity and mortality worldwide as it alters the architecture of organs and tissues, thereby disrupting normal function. Fibrosis can affect almost any organ or tissue and is associated with a wide variety of diseases and injuries. Fibrosis, seen in the liver, lung, heart, kidney, and skin, is a significant global disease burden. Burns, which often result in severe fibrotic scarring, rank 15 th in the global disease burden. The hallmarks of fibrosis, including increased presence, differentiation, and persistence of myofibroblasts(myoFBs); increased collagen deposition; decreased collagen degradation; and increased contraction, are conserved in most models of organ fibrosis. Hypertrophic scar is one of the fibrotic diseases, which is a pathologic scarring induced by abnormal wound healing after injuries of skin. After skin injury, there is an intermixed sequence of cellular, extracellular, and biomolecular events in wounds within the framework of the commonly designated phases of inflammation, proliferation and reparation, followed by a lengthy phase of remodeling or maturation that may last several months or years. In these three stages, there are complicated interactions within a complex network of profibrotic and antifibrotic molecules, such as growth factors, proteolytic enzymes and extracellular matrix(ECM) proteins. The delicate balance of deposition and degradation of ECM protein will be disrupted when either excessive production of collagen, proteoglycans and fibronectin by fibroblasts or deficient degradation and remodeling of ECM occurs. Hypertrophic scar occurs when the inflammatory response to injury is prolonged, leading to the pathological characteristics of hypertrophic scar including increased vascularization, hypercellularity and excessive collagen deposition..Renal fibrosis is the common final manifestation of a wide variety of chronic kidney diseases(CKD) that is characterized pathologically by glomerulosclerosis, interstitial fibrosis, tubular atrophy, peritubular capillary rarefaction, and inflammation. The pathogenesis is a progressive process that ultimately leads to the complete destruction of the kidney parenchyma and end-stage renal failure. Recent studies indicate that the myofibroblast is the most important contributor to kidney fibrogenesis because it not only deposits extracellular matrix(ECM) but also produces an array of inflammatory cytokines as well as toxic oxygen radicals. In the interstitial space of the kidney under the condition resulting from the above-mentioned insults, myofibroblasts intensively accumulate and synthesize the components of the matrix and also contribute directly to parenchymal and microvascular destruction. Currently, the identified contributors responsible for the development of renal fibrosis include epithelial injury, inflammation and the subsequent innate and adaptive immune responses. Furthermore, recent studies have implicated components of several signaling pathways in regulating the transition of mesenchymal progenitors to myofibroblasts, such as transforming growth factor-β(TGF-β), platelet-derived growth factor(PDGF), connective tissue growth factor(CTGF), epidermal growth factor(EGF), hedgehog(Hh), Gli, Wnt and Notch.Eukaryotic initiation factor 6(e IF6 also known as Integrin β4 binding protein-ITGB4BP), is a ribosome anti-association factor that regulates translational initiation and ribosome synthesis due to its capacity to modulate ribosome 60 S availability and 80 S subunit formation. Additionally, e IF6 also plays a crucial role in cell cycle progression, cell growth and tumorigenesis. Moreover, eIF6 has been identified as an interacting protein of the hypertrophic scar-related protein P311, which indicates that it may be involved in the regulation of hypertrophic scar formation as well as myofibroblast differentiation. However, the expression of e IF6 and its potential role in kidney fibrosis remain unknown.e IF6 m RNA length is 1108 bp, and its open reading frame consists of 735 nucleotides. Previous study with fluorescence in situ hybridization demonstrated that e IF6 gene locates on the 20q11.2 region of the long arm of 21 st chromosome. eIF6 gene is comprised of 7 exons and 6 introns, and there are no TATA promoter binding domain and Cp G islands. In eukaryote cells, the sequence of eIF6 gene is conservative. e IF6 gene is found not only in Homo sapiens, mice, rats, Xenopus laevis, and other vertebrates, but also in Drosophila, sepia, molluscum, and other invertebrates, and the domains of eIF6 gene are highly conservative between vertebrates and invertebrates.After the process of transcription and post-transcription, genes turn into mature m RNA, and are translated into polypeptide chains on ribosomes in the cytoplasm. During the growth and differentiation of eukaryote cells, most genes in the genome are not transcribed and only a few genes can be expressed. The regulation of mRNA translation is based on many levels, among which, the regulation of initial stage in eukaryotes is very important. In the nucleus, the binding of eIF6 with 60 s subunit inhibits the formation of 80 s subunit. In the initial stage of protein translation, as the eukaryotic initiation factor(eIF), e IF6 is phosphorylated under the action of CK1-PKC, and depolymerized down from 60 s subunit, which enhances the formation of 80 s subunit through the binding of 40 s subunit with 60 s subunit, and at last the protein translation is initiated. On the contrary, if the eIF6 is removed, the inhibition of microRNA towards the target protein expression is relieved, and as a result, the expression of the target protein is enhanced. The synthesis of eIF6 protein is relevant to the regulation on microRNA. By binding e IF6 and RNA-induced silencing complex(RISC), the function of the corresponding micro RNA is promoted, the mRNA formation is interfered from the transcriptional level, and the expression of the corresponding target protein is inhibited.Our previous pilot study found that the eukaryotic initial factor 6(eIF6) expression was significant decreased in fibroblasts derived from hypertrophic scar compared with that from normal skin in vitro. Moreover, e IF6 deficiency promoted α-SMA and TGF-β1 expression in fibroblasts, cutaneous fibrosis and renal fibrosis in an animal model. These data implied the involvement of e IF6 in hypertrophic scar formation and renal fibrosis in human being. However, the spatial and temporal expression of e IF6 in the process of hypertrophic scar formation and renal fibrosis remain unknown. The current study was designed to study the e IF6 protein expression pattern during the process of hypertrophic scar formation and renal fibrosis.This research is designed to divide into following three aspects, to further illustrate e IF6 in burns patients of different period hypertrophic scar and other fibrotic disease of chromic glomerulonephritis(Tubulointerstitial fibrosis): 1) the difference of eIF6 expression and distribution of different period hypertrophic scar and normal skin; 2) the differences of e IF6 expression content of different period hypertrophic scar and normal skin of expression content differences; 3) eIF6 expression pattern in Tubulointerstitial fibrosis and relatively normal kidney tissue.1. eIF6 expression pattern of different period of hypertrophic scar and normal skin tissueFirstly, we collect postburn Hypertrophic Scar in different periods, as well as with normal skin tissue samples to detect eIF6 distribution. Results showed that in normal skin tissues, eIF6 was widely distributed in the keratinocytes of epidernis, mainly including the basal layer, spinouts cell layer, the granular layer, transparent layer and stratum corneum. e IF6 expressed in the cell cytoplasm.In contrast, e IF6 expressed in hypertrophic scar differentially. In Proliferative Phase, eIF6 also mainly distributed in the epidermal layer and was significantly lower than that of normal skin tissue. In the basal layer of the epidermis, e IF6 expression is negative. In Mature Phase ofhypertrophic scar, e IF6 expression increased than of Proliferative Phase, but still lower than the normal skin tissue. And the basement layer has a small amount of e IF6-positive cells. Despite the expressions of eIF6 was lower than that of normal skin tissue, significantly higher than the Proliferative Phase, and the basal layer of epidermis dustributed more of eIF6-positive cells.2. eIF6 expression level of different period of hypertrophic scar and normal skin tissueSecondly, we analyzed the expression level of different period of hypertrophic scar and normal skin tissue by Western Blot Analysis. We found that, comparing with normal skin, e IF6 expression level in hypertrophic scar tissue was significantly decreased, and the reduction of Proliferative Phase is the most significant. Although e e IF6 expression volume remains reduced in Regression Phase compared to normal skin, was obviously increased compared to early scars. This clued that e IF6 expression level in scar formation showed a gradually increased trend. In combination of the clinical characteristics that the hypertrophic scar can be regressed gradually. This tips e IF6 can be a negative regulatory molecular in scar formation.3. eIF6 expression pattern of tubular/interstitial fibrosis of kidney diseasesWe examined the expression pattern of the eIF6 protein in human kidney samples using immunohistochemical analysis. In the kidneys with no significant tubulointerstitial fibrosis/glomerulosclerosis, eIF6 was found in the cells of the visceral layer of the glomerulus capsule. Proximal convoluted tubular cells and distal convoluted tubular cells mainly expressed e IF6 in the cytoplasm. Moreover, the interstitial cells were also positive for eIF6. In the kidneys with moderate to severe tubulointerstitial fibrosis/ glomerulosclerosis, histological examination confirmed local fibrosis with excessive collagen fiber deposition surrounding the sclerotic glomerulus or damaged tubules. Sclerotic glomeruli were not positive for eIF6 expression, particularly in the cells of the visceral layer of the glomerulus capsule, but the tubular cells were still positive for eIF6 expression. Most of the fibroblasts in the widened interstitial space were eIF6 negative, but the affected proximal and distal tubular epithelial cells showed a strong e IF6-positive expression pattern.Conclusions: we find that eIF6 expression deceased in hypertrophic scars comparing with normal skin. Following the formation time of hypertrophic scar, eIF6 expression level has increased. Although there was an increase of eIF6 expression in hypertrophic scars of Regression Phase, its expression was still relatively lower than in the normal skin. In renal fibrosis, e IF6 distributed differently than in relatvely normal kidney. In Unilateral Ureteral Obstruction(UUO) eIF6+/- mcie model, we found that the expression of collagen I, TGF-beta 1 and Alpha-SMA increased. It proves that e IF6 may affect TGF-beta 1 signal way to participate renal fibrosis formation. And in the future, e IF6 may become one of the fibrotic disease therapeutic targets. |
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