Pax2在肾脏发育和肾疾病中的调控作用

配对盒基因2(Paired box2,Pax2)是肾脏发育中重要的转录因子,在前、中、后肾发育的全过程表达,集中分布在发育的各级小管和间充质成分,具有特定的时空特性。研究表明Pax2与多种调节肾脏发育的因子Gdnf、Ret、SHH、Wnt4及Fgf等相互作用,共同精准诱导生肾索形成,前/中肾管的形成及分化,输尿管芽的发生及分支,肾单位的诱导分化。Pax2的变异导致多种先天性肾脏及输尿管发育畸形,最易发生在肾-视神经盘缺损综合征。在肾细胞癌、Wilms瘤和多种肾小球及肾小管获得性疾病中存在Pax2的异常表达,其诊断和治疗价值将是今后研究的重点。文章主要对Pax2的分子结构、在肾脏发育和肾疾病的表达及调控进行了综述。     

Pax8, a murine paired box gene expressed in the developing excretory system and thyroid gland.

Several mouse genes designated 'Pax genes' contain a highly conserved DNA sequence homologous to the paired box of Drosophila. Here we describe the isolation of Pax8, a novel paired box containing clone from an 8.5 day p.c. mouse embryo cDNA library. An open reading frame of 457 amino acids (aa) contains the 128 aa paired domain near the amino terminus. Another conserved region present in some other paired box genes, the octapeptide Tyr-Ser-Ile-Asn-Gly-Leu-Leu-Gly, is located 43 aa C-terminal to the paired domain. Using an interspecies backcross system, we have mapped the Pax8 gene within the proximal portion of mouse chromosome 2 in a close linkage to the surf locus. Several developmental mutations are located in this region. In situ hybridization was used to determine the pattern of Pax8 expression during mouse embryogenesis. Pax8 is expressed transiently between 11.5 and 12.5 days of gestation along the rostrocaudal axis extending from the myelencephalon throughout the length of the neural tube, predominantly in two parallel regions on either side of the basal plate. We also detected Pax8 expression in the developing thyroid gland beginning at 10.5 days of gestation, during the thyroid evagination. In the mesonephros and metanephros the expression of Pax8 was localized to the mesenchymal condensations, which are induced by the nephric duct and ureter, respectively. These condensations develop to functional units, the nephrons, of the kidney. These data are consistent with a role for Pax8 in the induction of kidney epithelium. The embryonic expression pattern of Pax8 is compared with that of Pax2, another recently described paired box gene expressed in the developing excretory system.     

Parallel waves of inductive signaling and mesenchyme maturation regulate differentiation of the chick mesonephros

The mesonephros is a linear kidney that, in chicken embryos, stretches between the axial levels of the 15th to the 30th somites. Mesonephros differentiation proceeds from anterior to posterior and is dependent on signals from the nephric duct, which migrates from anterior to posterior through the mesonephric region. If migration of the nephric duct is blocked, markers of tubule differentiation, including Lhx1 and Wnt4, are not activated posterior to the blockade. However, activation and maintenance of the early mesonephric mesenchyme markers Osr1, Eya1 and Pax2 proceeds normally in an anterior-to-posterior wave, indicating that these genes are not dependent on inductive signals from the duct. The expression of Lhx1 and Wnt4 can be rescued in duct-blocked embryos by supplying a source of canonical Wnt signaling, although epithelial structures are not obtained, suggesting that the duct may express other tubule-inducing signals in addition to Wnts. In the absence of the nephric duct, anterior mesonephric mesenchyme adjacent to somites exhibits greater competence to initiate tubular differentiation in response to Wnt signaling than more posterior mesonephric mesenchyme adjacent to unsegmented paraxial mesoderm. It is proposed that mesonephric tubule differentiation is regulated by two independent parallel waves, one of inductive signaling from the nephric duct and the other of competence of the mesonephric mesenchyme to undergo tubular differentiation, both of which travel from anterior to posterior in parallel with the formation of new somites.     

Regulation of ureteric bud outgrowth by Pax2-dependent activation of the glial derived neurotrophic factor gene.

The outgrowth of the ureteric bud from the posterior nephric duct epithelium and the subsequent invasion of the bud into the metanephric mesenchyme initiate the process of metanephric, or adult kidney, development. The receptor tyrosine kinase RET and glial cell-derived neurotrophic factor (GDNF) form a signaling complex that is essential for ureteric bud growth and branching morphogenesis of the ureteric bud epithelium. We demonstrate that Pax2 expression in the metanephric mesenchyme is independent of induction by the ureteric bud. Pax2 mutants are deficient in ureteric bud outgrowth and do not express GDNF in the uninduced metanephric mesenchyme. Furthermore, Pax2 mutant mesenchyme is unresponsive to induction by wild-type heterologous inducers. In normal embryos, GDNF is sufficient to induce ectopic ureter buds in the posterior nephric duct, a process inhibited by bone morphogenetic protein 4. However, GDNF replacement in organ culture is not sufficient to stimulate ureteric bud outgrowth from Pax2 mutant nephric ducts, indicating additional defects in the nephric duct epithelium of Pax2 mutants. Pax2 can activate expression of GDNF in cell lines derived from embryonic metanephroi. Furthermore, Pax2 protein can bind to upstream regulatory elements within the GDNF promoter region and can transactivate expression of reporter genes. Thus, activation of GDNF by Pax2 coordinates the position and outgrowth of the ureteric bud such that kidney development can begin.     

Lim 1 is required for nephric duct extension and ureteric bud morphogenesis

The nephric duct plays a central role in orchestrating the development of the mammalian urogenital system. Lim 1 is a homeobox gene required for head and urogenital development in the mouse but most Lim 1-deficient embryos die by embryonic day 10. To determine the role of Lim 1 in the development of the nephric duct, we conditionally removed Lim 1 in the nephric epithelium just after the nephric duct begins to form using a floxed allele of Lim 1 and Pax2-cre transgenic mice. We report that Lim 1 conditional knockout mice have renal hypoplasia and hydronephrosis. Developmental studies revealed that the caudal portion of the nephric duct did not reach the urogenital sinus at embryonic day 10.5, formation of the ureteric bud was delayed, the ureteric bud was smaller and branching of the ureteric bud reduced. We also found that the nephric duct was generally not maintained and extension of the Müllerian duct inhibited. Molecular analysis indicated that Pax2 was expressed normally but the expression of Wnt9b and E-cadherin in the nephric duct was markedly altered. These results suggest that Lim 1 influences nephric duct extension and ureteric bud outgrowth by regulating and or maintaining the differentiation of the nephric epithelium.     

Morphology of the kidney in larvae of Bufo viridis (Amphibia, Anura, Bufonidae)

This study deals primarily with the morphology and ultrastructure of the pronephros in the green toad Bufo viridis during prometamorphosis when the pronephros and the developing mesonephros function simultaneously. Furthermore, the mesonephros was studied during pro- and postmetamorphosis with emphasis on the distal segments of the nephron. The paired kidneys consist of two cranial pronephroi immediately behind the gill region and two more caudal elongated mesonephroi. Each pronephros consists of a single convoluted tubule which opens into the coelom via three nephrostomes. This tubule is divided into three ciliated tubules, three proximal tubule branches, a common proximal tubule and a distal tubule, which in turn continues into the nephric duct. No intermediate segment is present. The length of the pronephric tubule is 12 mm, including the three branches of the ciliated tubules and proximal tubules. Primary urine is formed upon filtration from an external glomerulus, which is a convoluted capillary lined by podocytes, a specialization of the coelomic epithelium. From the coelom the filtrate is swept into the ciliated tubules. In the collecting duct system of the developing mesonephric nephron epithelial cells with conspicuous, apical osmiophilic granules appear in larvae of 9-10 mm. Heterocellularity of mixed intercalated (mitochondria rich) cells and principal cells is observed in the collecting duct system and nephric duct from a larval body length of 14 mm. As the proliferation of mitochondria-rich cells proceeds, the osmiophilic granules disappear and are completely absent from the adult amphibian mesonephros.     

Eya 1 acts as a critical regulator for specifying the metanephric mesenchyme

Although it is well established that the Gdnf-Ret signal transduction pathway initiates metanephric induction, no single regulator has yet been identified to specify the metanephric mesenchyme or blastema within the intermediate mesoderm, the earliest step of metanephric kidney development and the molecular mechanisms controlling Gdnf expression are essentially unknown. Previous studies have shown that a loss of Eya 1 function leads to renal agenesis that is a likely result of failure of metanephric induction. The studies presented here demonstrate that Eya 1 specifies the metanephric blastema within the intermediate mesoderm at the caudal end of the nephrogenic cord. In contrast to its specific roles in metanephric development, Eya 1 appears dispensable for the formation of nephric duct and mesonephric tubules. Using a combination of null and hypomorphic Eya 1 mutants, we now demonstrated that approximately 20% of normal Eya 1 protein level is sufficient for establishing the metanephric blastema and inducing the ureteric bud formation but not for its normal branching. Using Eya 1, Gdnf, Six 1 and Pax 2 mutant mice, we show that Eya 1 probably functions at the top of the genetic hierarchy controlling kidney organogenesis and it acts in combination with Six 1 and Pax 2 to regulate Gdnf expression during UB outgrowth and branching. These findings uncover an essential function for Eya 1 as a critical determination factor in acquiring metanephric fate within the intermediate mesoderm and as a key regulator of Gdnf expression during ureteric induction and branching morphogenesis.     

Six1 and Six4 are essential for Gdnf expression in the metanephric mesenchyme and ureteric bud formation, while Six1 deficiency alone causes mesonephric-tubule defects

Interaction between the ureteric-bud epithelium and the metanephric mesenchyme is important for kidney development. Six1 and Six4 are the mammalian homologs of Drosophila sine oculis, and they are coexpressed in the nephrogenic mesenchyme. Six1-deficient mice show varying kidney defects, while Six4-deficient mice have no apparent abnormalities. Here, we report Six1/Six4-deficient mice that we generated in order to elucidate the functions of Six4 in Six1-deficient kidney development. The Six1/Six4-deficient mice exhibited more severe kidney phenotypes than the Six1-deficient mice; kidney and ureter agenesis was observed in all the neonates examined. The Six1/Six4-deficient metanephric mesenchyme cells were directed toward kidney lineage but failed to express Pax2, Pax8, or Gdnf, whereas the expression of these genes was partially reduced or unchanged in the case of Six1 deficiency. Thus, Six4 cooperates with Six1 in the metanephric mesenchyme to regulate the level of Gdnf expression; this could explain the absence of the ureteric bud in the Six1/Six4-deficient mice. In contrast, Six1 deficiency alone caused defects in mesonephric-tubule formation, and these defects were not exacerbated in the Six1/Six4-deficient mesonephros. These results highlight the fact that Six1 and Six4 have collaborative functions in the metanephros but not in the mesonephros.     

Mesonephric kidney--a stem cell factory?

Mesonephros is a vestige, transient renal organ that functions only during embryonic development. The anatomy, position and even cellular fate of the mesonephric kidney varies drastically among mammalian species. The origin of mesonephros from intermediate mesoderm and the dependence of its differentiation on the nephric or Wolffian duct have been well established. Commonly accepted is also the mesonephric origin of epididymal ducts of the male reproductive tract. Recently, upon the more profound understanding of the molecular mechanisms involved in the development of the permanent mammalian kidney, some light has been shed over the molecular events taking place during the mesonephric development as well. Because of the functional and structural similarities between the mesonephric and metanephric kidneys, it is not surprising that many molecules regulating metanephric development are also activated during mesonephric development. However, the multifunctional nature of mesonephros has been unexpected. First, it serves as an embryonic secretory organ, in some mammalian species more so than in others. It is thereafter removed by programmed cell death. Second, it is a source of multiple stem cells including somatic cells in the male gonad, vascular endothelial cells, and hematopoietic stem cells. Thus, mesonephros is a challenging model for studies on epithelial differentiation and organogenesis, regulation of apoptosis, sex determination and stem cell differentiation. In this review, we focus in the molecular and stem cell aspects in the differentiation of the mammalian mesonephros.