EGFR/MAPK Signaling Regulates the Proliferation of Drosophila Renal and Nephric Stem Cells

Tissue homeostasis,accomplished through the self-renewal and differentiation of resident stem cells,is critical for the maintenance of adult tissues throughout an animal's lifetime.Adult Drosoplula Malpighian tubules(MTs or fly kidney) are maintained by renal and nephric stem cells(RNSCs) via self-renewing divisions,however,it is unclear how RNSC proliferation and differentiation are regulated.Here we show that EGFR/MAPK signaling is dispensable for RNSC maintenance,but required for RNSC proliferation in vivo.Inactivation of the EGFR/MAPK pathway blocks or greatly retards RNSC cell cycle progression:conversely,over-activation of EGFR/MAPK signaling results in RNSC over-proliferation and disrupts the normal differentiation of renablasts(RBs),the immediate daughters of RNSC divisions.Our data further suggest that EGFR/MAPK signaling functions independently of JAK/STAT signaling and that dMyc and CycE partially mediate EGFR/MAPK signaling in MTs.Together,our data suggest a principal role of EGFR/MAPK signaling in regulating RNSC proliferation,which may provide important clues for understanding mammalian kidney repair and regeneration following injury.     

Kidney regeneration and resident stem cells

Given its complexity, high metabolic activity and excretory functions, the kidney is particularly susceptible to acute ischemic and toxin-mediated injury. Current therapies do not facilitate kidney regeneration, and there is an increasing interest in newer therapies that are based on cellular sources of kidney regeneration, such as stem cell therapy. Our understanding of cellular sources for kidney regeneration and stem cells present in the adult kidney has dramatically evolved over the recent years. Herein, we discuss the current understanding of kidney stem cells present in the adult mammalian kidney and their role in kidney regeneration. We have also summarized the best available evidence supporting the role of stem cells in kidney regeneration.     

Kidney regeneration and resident stem cells

Given its complexity, high metabolic activity and excretory functions, the kidney is particularly susceptible to acute ischemic and toxin-mediated injury. Current therapies do not facilitate kidney regeneration, and there is an increasing interest in newer therapies that are based on cellular sources of kidney regeneration, such as stem cell therapy. Our understanding of cellular sources for kidney regeneration and stem cells present in the adult kidney has dramatically evolved over the recent years. Herein, we discuss the current understanding of kidney stem cells present in the adult mammalian kidney and their role in kidney regeneration. We have also summarized the best available evidence supporting the role of stem cells in kidney regeneration.     

Stem cell marker TRA-1-60 is expressed in foetal and adult kidney and upregulated in tubulo-interstitial disease

The kidney has an intrinsic ability to repair itself when injured. Epithelial cells of distal tubules may participate in regeneration. Stem cell marker, TRA-1-60 is linked to pluripotency in human embryonic stem cells and is lost upon differentiation. TRA-1-60 expression was mapped and quantified in serial sections of human foetal, adult and diseased kidneys. In 8- to 10-week human foetal kidney, the epitope was abundantly expressed on ureteric bud and structures derived therefrom including collecting duct epithelium. In adult kidney inner medulla/papilla, comparisons with reactivity to epithelial membrane antigen, aquaporin-2 and Tamm–Horsfall protein, confirmed extensive expression of TRA-1-60 in cells lining collecting ducts and thin limb of the loop of Henle, which may be significant since the papillae were proposed to harbour slow cycling cells involved in kidney homeostasis and repair. In the outer medulla and cortex there was rare, sporadic expression in tubular cells of the collecting ducts and nephron, with positive cells confined to the thin limb and thick ascending limb and distal convoluted tubules. Remarkably, in cortex displaying tubulo-interstitial injury, there was a dramatic increase in number of TRA-1-60 expressing individual cells and in small groups of cells in distal tubules. Dual staining showed that TRA-1-60 positive cells co-expressed Pax-2 and Ki-67, markers of tubular regeneration. Given the localization in foetal kidney and the distribution patterns in adults, it is tempting to speculate that TRA-1-60 may identify a population of cells contributing to repair of distal tubules in adult kidney.     

Genetic control of development of the Malpighian vessels in Drosophila melanogaster

Malpighian tubules of insects are a functional analog of mammalian kidneys and serve as a classical model for studying the structure and functions of transport epithelium. The review contains the data on structural organization, functioning, and formation of the Malpighian tubules during embryogenesis in Drosophila melanogaster. Various systems of genes are described that control the program of development of the renal (Malpighian) tubules in D. melanogaster. A special attention is paid to the ways of signal transduction and factors involved in cell differentiation, proliferation, and morphological transformation during development of the Malpighian tubules. Evolutionarily conservative genetic systems are considered that are involved in the control of development of both the renal epithelium of Drosophila and mammalian kidneys. A relationship was noted between the disturbed balance of genetic material and congenital defects of the human excretory system.     

Species Diversity Regarding the Presence of Proximal Tubular Progenitor Cells of the Kidney

The cellular source for tubular regeneration following kidney injury is a matter of dispute, with reports suggesting a stem or progenitor cells as the regeneration source while linage tracing studies in mice seemingly favor the classical theory, where regeneration is performed by randomly surviving cells. We, and others have previously described a scattered cell population localized to the tubules of human kidney, which increases in number following injury. Here we have characterized the species distribution of these proximal tubular progenitor cells (PTPCs) in kidney tissue from chimpanzee, pig, rat and mouse using a set of human PTPC markers. We detected PTPCs in chimpanzee and pig kidneys, but not in mouse tissue. Also, subjecting mice to the unilateral urethral obstruction model, caused clear signs of tubular injury, but failed to induce the PTPC phenotype in renal tubules.Key words: Acute tubular necrosis, tubular regeneration, species diversity, proximal tubules     

Intrinsic epithelial cells repair the kidney after injury

Understanding the mechanisms of nephron repair is critical for the design of new therapeutic approaches to treat kidney disease. The kidney can repair after even a severe insult, but whether adult stem or progenitor cells contribute to epithelial renewal after injury and the cellular origin of regenerating cells remain controversial. Using genetic fate-mapping techniques, we generated transgenic mice in which 94%-95% of tubular epithelial cells, but no interstitial cells, were labeled with either beta-galactosidase (lacZ) or red fluorescent protein (RFP). Two days after ischemia-reperfusion injury (IRI), 50.5% of outer medullary epithelial cells coexpress Ki67 and RFP, indicating that differentiated epithelial cells that survived injury undergo proliferative expansion. After repair was complete, 66.9% of epithelial cells had incorporated BrdU, compared to only 3.5% of cells in the uninjured kidney. Despite this extensive cell proliferation, no dilution of either cell-fate marker was observed after repair. These results indicate that regeneration by surviving tubular epithelial cells is the predominant mechanism of repair after ischemic tubular injury in the adult mammalian kidney.     

A Drosophila group E Sox gene is dynamically expressed in the embryonic alimentary canal

We have identified a novel Drosophila Sox-domain gene, Sox100B, related to the vertebrate group E genes Sox9 and Sox10. In vertebrates, group E Sox genes are expressed in the developing gonad, adult kidney and gut as well as other tissues. During embryogenesis in Drosophila, Sox100B is expressed in two rows of large intestinal cells, in midgut basophilic cells, in the Malpighian tubules and at the posterior cap of gonadal mesoderm. Our observations indicate that aspects of tissue-specific expression, as well as sequence, are conserved between vertebrate and invertebrate group E Sox proteins.     

Non-apoptotic function of apoptotic proteins in the development of Malpighian tubules of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Drosophila metamorphosis is characterized by the histolysis of larval structures by programmed cell death, which paves the way for the establishment of adult-specific structures under the influence of the steroid hormone ecdysone. Malpighian tubules function as an excretory system and are one of the larval structures that are not destroyed during metamorphosis and are carried over to adulthood. The pupal Malpighian tubules evade destruction in spite of expressing apoptotic proteins, Reaper, Hid, Grim, Dronc and Drice. Here we show that in the Malpighian tubules expression of apoptotic proteins commences right from embryonic development and continues throughout the larval stages. Overexpression of these proteins in the Malpighian tubules causes larval lethality resulting in malformed tubules. The number and regular organization of principal and stellate cells of Malpighian tubules is disturbed, in turn disrupting the physiological functioning of the tubules as well. Strikingly, the localization of beta-tubulin, F-actin and Disclarge (Dlg) is also disrupted. These results suggest that the apoptotic proteins could be having non-apoptotic function in the development of Malpighian tubules.     

Severely damaged kidneys possess multipotent renoprotective stem cells

Background aimsTissue-specific stem cells are a promising target for kidney regeneration, because it has been shown that they play a primary role in kidney repair. Several methods have been developed for the isolation of stem/progenitor cells from healthy kidneys but the existence of these cells in chronically damaged kidneys has not been noticed so far.MethodsA mouse model of chronic kidney failure was developed by ligation of the left ureter for 5 months, and then isolation of stem cells from this tissue as well as normal kidneys was attempted.ResultsWe found that multipotent stem cells could be isolated from both types of tissue. In addition, the cells isolated from damaged kidneys showed potential for homing to the site of injury and a renoprotective effect in an animal model of cisplatin-induced nephropathy.ConclusionsThese results show that multipotent renoprotective stem cells exist in severely damaged kidneys, which could be a target for designing new therapies.     

The receptor guanylate cyclase Gyc76C and a peptide ligand, NPLP1-VQQ, modulate the innate immune IMD pathway in response to salt stress

Receptorguanylate cyclases (rGCs) modulate diverse physiological processes including mammalian cardiovascular function and insect eclosion. The Drosophila genome encodes several receptor and receptor-like GCs, but no ligand for any Drosophila rGC has yet been identified. By screening peptide libraries in Drosophila S2 cells, the Drosophila peptide NPLP1-VQQ (NLGALKSSPVHGVQQ) was shown to be a ligand for the rGC, Gyc76C (CG42636, previously CG8742, l(3)76BDl, DrGC-1). In the adult fly, expression of Gyc76C is highest in immune and stress-sensing epithelial tissues, including Malpighian tubules and midgut; and NPLP1-VQQ stimulates fluid transport and increases cGMP content in tubules. cGMP signaling is known to modulate the activity of the IMD innate immune pathway in tubules via activation and nuclear translocation of the NF-kB orthologue, Relish, resulting in increased anti-microbial peptide (AMP) gene expression; and so NPLP1-VQQ might act in immune/stress responses. Indeed, NPLP1-VQQ induces nuclear translocation of Relish in intact tubules and increases expression of the anti-microbial peptide gene, diptericin. Targeted Gyc76C RNAi to tubule principal cells inhibited both NPLP1-VQQ-induced Relish translocation and diptericin expression. Relish translocation and increased AMP gene expression also occurs in tubules in response to dietary salt stress. Gyc76C also modulates organismal survival to salt stress - ablation of Gyc76C expression in only tubule principal cells prevents Relish translocation, reduces diptericin expression, and reduces organismal survival in response to salt stress. Thus, the principal-cell localized NPLP1-VQQ/Gyc76C cGMP pathway acts to signal environmental (salt) stress to the whole organism.     

Genetic Control of Development of Malpighian Tubules in Drosophila melanogaster

Malpighian tubules of insects are a functional analog of mammalian kidneys and serve as a classical model for studying the structure and functions of transport epithelium. The review contains the data on structural organization, functioning, and formation of the Malpighian tubules during embryogenesis in Drosophila melanogaster. Various systems of genes are described that control the program of development of the renal (Malpighian) tubules in D. melanogaster. A special attention is paid to the ways of signal transduction and factors involved in cell differentiation, proliferation, and morphological transformation during development of the Malpighian tubules. Evolutionarily conservative genetic systems are considered that are involved in the control of development of both the renal epithelium ofDrosophila and mammalian kidneys. A relationship was noted between the disturbed balance of genetic material and congenital defects of the human excretory system.     

Ion and solute transport by Prestin in Drosophila and Anopheles

The gut and Malpighian tubules of insects are the primary sites of active solute and water transport for controlling hemolymph and urine composition, pH, and osmolarity. These processes depend on ATPase (pumps), channels and solute carriers (Slc proteins). Maturation of genomic databases enables us to identify the putative molecular players for these processes. Anion transporters of the Slc4 family, AE1 and NDAE1, have been reported as HCO(3)(-) transporters, but are only part of the story. Here we report Dipteran (Drosophila melanogaster (d) and Anopheles gambiae (Ag)) anion exchangers, belonging to the Slc26 family, which are multi-functional anion exchangers. One Drosophila and two Ag homologues of mammalian Slc26a5 (Prestin) and Slc26a6 (aka, PAT1, CFEX) were identified and designated dPrestin, AgPrestinA and AgPrestinB. dPrestin and AgPrestinB show electrogenic anion exchange (Cl(-)/nHCO(3)(-), Cl(-)/SO(4)(2-) and Cl(-)/oxalate(2-)) in an oocyte expression system. Since these transporters are the only Dipteran Slc26 proteins whose transport is similar to mammalian Slc26a6, we submit that Dipteran Prestin are functional and even molecular orthologues of mammalian Slc26a6. OSR1 kinase increases dPrestin ion transport, implying another set of physiological processes controlled by WNK/SPAK signaling in epithelia. All of these mRNAs are highly expressed in the gut and Malpighian tubules. Dipteran Prestin proteins appear suited for central roles in bicarbonate, sulfate and oxalate metabolism including generating the high pH conditions measured in the Dipteran midgut lumen. Finally, we present and discuss Drosophila genetic models that integrate these processes.     

成体干细胞肺重建研究进展

成体干细胞来源广泛,无伦理争议,成为近几年的关注热点。研究表明以骨髓来源的间充质干细胞为代表的成体干细胞具有较强的多系分化潜能,可以广泛的参与包括肺在内的受损组织的修复与重建。在动物实验中已观察到,供体来源的成体干细胞可以定向分化为受损肺组织的多种功能细胞,并且有抑制纤维化等病变产生的能力。在本文中,回顾了近年来与肺损伤重建和疾病治疗相关的干细胞研究的最新进展,并探讨了成体干细胞治疗肺疾病与损伤的临床应用前景。     

Use of mouse hematopoietic stem and progenitor cells to treat acute kidney injury

New and effective treatment for acute kidney injury remains a challenge. Here, we induced mouse hematopoietic stem and progenitor cells (HSPC) to differentiate into cells that partially resemble a renal cell phenotype and tested their therapeutic potential. We sequentially treated HSPC with a combination of protein factors for 1 wk to generate a large number of cells that expressed renal developmentally regulated genes and protein. Cell fate conversion was associated with increased histone acetylation on promoters of renal-related genes. Further treatment of the cells with a histone deacetylase inhibitor improved the efficiency of cell conversion by sixfold. Treated cells formed tubular structures in three-dimensional cultures and were integrated into tubules of embryonic kidney organ cultures. When injected under the renal capsule, they integrated into renal tubules of postischemic kidneys and expressed the epithelial marker E-cadherin. No teratoma formation was detected 2 and 6 mo after cell injection, supporting the safety of using these cells. Furthermore, intravenous injection of the cells into mice with renal ischemic injury improved kidney function and morphology by increasing endogenous renal repair and decreasing tubular cell death. The cells produced biologically effective concentrations of renotrophic factors including VEGF, IGF-1, and HGF to stimulate epithelial proliferation and tubular repair. Our study indicates that hematopoietic stem and progenitor cells can be converted to a large number of renal-like cells within a short period for potential treatment of acute kidney injury.     

Characterization of the short neuropeptide F receptor from Drosophila melanogaster

A seven transmembrane G-protein coupled receptor has been cloned from Drosophila melanogaster. This receptor shows structural similarities to vertebrate Neuropeptide Y(2) receptors and is activated by endogenous Drosophila peptides, recently designated as short neuropeptide Fs (sNPFs). sNPFs have so far been found in neuroendocrine tissues of four other insect species and of the horseshoe crab. In locusts, they accelerate ovarian maturation, and in mosquitoes, they inhibit host-seeking behavior. Expression analysis by RT-PCR shows that the sNPF receptor (Drm-sNPF-R) is present in several tissues (brain, gut, Malpighian tubules and fat body) from Drosophila larvae as well as in ovaries of adult females. All 4 Drosophila sNPFs clearly elicited a calcium response in receptor expressing mammalian Chinese hamster ovary cells. The response is dose-dependent and appeared to be very specific. The short NPF receptor was not activated by any of the other tested arthropod peptides, not even by FMRFamide-related peptides (also ending in RFamide), indicating that the Arg residue at position 4 from the amidated C-terminus appears to be crucial for the response elicited by the sNPFs.     

Salamander limb regeneration involves the activation of a multipotent skeletal muscle satellite cell population

In contrast to mammals, salamanders can regenerate complex structures after injury, including entire limbs. A central question is whether the generation of progenitor cells during limb regeneration and mammalian tissue repair occur via separate or overlapping mechanisms. Limb regeneration depends on the formation of a blastema, from which the new appendage develops. Dedifferentiation of stump tissues, such as skeletal muscle, precedes blastema formation, but it was not known whether dedifferentiation involves stem cell activation. We describe a multipotent Pax7⁺ satellite cell population located within the skeletal muscle of the salamander limb. We demonstrate that skeletal muscle dedifferentiation involves satellite cell activation and that these cells can contribute to new limb tissues. Activation of salamander satellite cells occurs in an analogous manner to how the mammalian myofiber mobilizes stem cells during skeletal muscle tissue repair. Thus, limb regeneration and mammalian tissue repair share common cellular and molecular programs. Our findings also identify satellite cells as potential targets in promoting mammalian blastema formation.     

Adult stem-like cells in kidney

Human pluripotent cells are promising for treatment for kidney diseases, but the protocols for derivation of kidney cell types are still controversial. Kidney tissue regeneration is well confirmed in several lower vertebrates such as fish, and the repair of nephrons after tubular damages is commonly observed after renal injury. Even in adult mammal kidney, renal progenitorcell or system is reportedly presents suggesting that adult stem-like cells in kidney can be practical clinical targets for kidney diseases. However, it is still unclear if kidney stem cells or stem-like cells exist or not. In general, stemness is defined by several factors such as self-renewal capacity, multi-lineage potency and characteristic gene expression profiles. The definite use of stemness may be obstacle to understand kidney regeneration, and here we describe the recent broad findings of kidney regeneration and the cells that contribute regeneration.