PAX2 is regulated by estrogen/progesterone through promoter methylation in endometrioid adenocarcinoma and has an important role in carcinogenesis via the AKT/mTOR signaling pathway.

Endometrioid adenocarcinoma (EEC) is one of the most common cancers of the female reproductive system. In recent years, much emphasis has been placed on early diagnosis and treatment. PAX2 (Paired box 2) inactivation is reportedly an important biomarker for endometrioid intraepithelial neoplasia (EIN) and EEC. However, the role of PAX2 in EEC carcinogenesis remains unclear. PAX2 expression and associated clinical characteristics were analyzed via The Cancer Genome Atlas, Gene Expression Omnibus, and Cancer Cell Line Encyclopedia databases and clinical paired EIN/EEC tissue samples. Bioinformatic analysis was conducted to identify the putative molecular function and mechanism of PAX2. Cell proliferation, colony formation, cell migration, and invasion assays in vitro, and mouse xenograft models were utilized to study the biological functions of PAX2 in vivo. Pyrosequencing and the demethylating drug 5-Aza-dc were used to verify promoter methylation in clinical tissues and cell lines, respectively. The mechanism underlying the regulatory effect of estrogen (E2) and progesterone (P4) on PAX2 expression was investigated by receptor block assay and double luciferase reporter assay. PAX2 expression was found to be significantly downregulated in EIN and EEC tissues, its overexpression inhibited EEC cell malignant behaviors in vivo and in vitro and inhibited the AKT/mTOR signaling pathway. PAX2 inactivation in EEC was related to promoter methylation, and its expression was regulated by E2 and P4 through their receptors via promoter methylation. Our findings elucidated the expression and function of PAX2 in EEC and have provided hitherto undocumented evidence of the underlying molecular mechanisms. PAX2 expression is suppressed by estrogen prompting its methylation through estrogen receptor. Furthermore, PAX2 regulates the AKT/mTOR signaling pathway to influence EEC progression. © 2024 The Pathological Society of Great Britain and Ireland.

Mice with a Pax2 missense variant display impaired glomerular repair.

PAX2 regulates kidney development, and its expression persists in parietal epithelial cells (PECs), potentially serving as a podocyte reserve. We hypothesized that mice with a Pax2 pathogenic missense variant (Pax2A220G/+) have impaired PEC-mediated podocyte regeneration. Embryonic wild-type mouse kidneys showed overlapping expression of PAX2/Wilms' tumor-1 (WT-1) until PEC and podocyte differentiation, reflecting a close lineage relationship. Embryonic and adult Pax2A220G/+ mice have reduced nephron number but demonstrated no glomerular disease under baseline conditions. Pax2A220G/+ mice compared with wild-type mice were more susceptible to glomerular disease after adriamycin (ADR)-induced podocyte injury, as demonstrated by worsened glomerular scarring, increased podocyte foot process effacement, and podocyte loss. There was a decrease in PAX2-expressing PECs in wild-type mice after adriamycin injury accompanied by the occurrence of PAX2/WT-1-coexpressing glomerular tuft cells. In contrast, Pax2A220G/+ mice showed no changes in the numbers of PAX2-expressing PECs after adriamycin injury, associated with fewer PAX2/WT-1-coexpressing glomerular tuft cells compared with injured wild-type mice. A subset of PAX2-expressing glomerular tuft cells after adriamycin injury was increased in Pax2A220G/+ mice, suggesting a pathological process given the worse outcomes observed in this group. Finally, Pax2A220G/+ mice have increased numbers of glomerular tuft cells expressing Ki-67 and cleaved caspase-3 compared with wild-type mice after adriamycin injury, consistent with maladaptive responses to podocyte loss. Collectively, our results suggest that decreased glomerular numbers in Pax2A220G/+ mice are likely compounded with the inability of their mutated PECs to regenerate podocyte loss, and together these two mechanisms drive the worsened focal segmental glomerular sclerosis phenotype in these mice.NEW & NOTEWORTHY Congenital anomalies of the kidney and urinary tract comprise some of the leading causes of kidney failure in children, but our previous study showed that one of its genetic causes, PAX2, is also associated with adult-onset focal segmental glomerular sclerosis. Using a clinically relevant model, our present study demonstrated that after podocyte injury, parietal epithelial cells expressing PAX2 are deployed into the glomerular tuft to assist in repair in wild-type mice, but this mechanism is impaired in Pax2A220G/+ mice.

Pax protein depletion in proximal tubules triggers conserved mechanisms of resistance to acute ischemic kidney injury preventing transition to chronic kidney disease.

Acute kidney injury (AKI) is a common condition that lacks effective treatments. In part, this shortcoming is due to an incomplete understanding of the genetic mechanisms that control pathogenesis and recovery. Identifying the molecular and genetic regulators unique to nephron segments that dictate vulnerability to injury and regenerative potential could lead to new therapeutic targets to treat ischemic kidney injury. Pax2 and Pax8 are homologous transcription factors with overlapping functions that are critical for kidney development and are re-activated in AKI. Here, we examined the role of Pax2 and Pax8 in recovery from ischemic AKI and found them upregulated after severe AKI and correlated with chronic injury. Surprisingly, proximal-tubule-selective deletion of Pax2 and Pax8 resulted in a less severe chronic injury phenotype. This effect was mediated by protection against the acute insult, similar to pre-conditioning. Prior to injury, Pax2 and Pax8 mutant mice develop a unique subpopulation of proximal tubule cells in the S3 segment that displayed features usually seen only in acute or chronic injury. The expression signature of these cells was strongly enriched with genes associated with other mechanisms of protection against ischemic AKI including caloric restriction, hypoxic pre-conditioning, and female sex. Thus, our results identified a novel role for Pax2 and Pax8 in mature proximal tubules that regulates critical genes and pathways involved in both the injury response and protection from ischemic AKI.

Retained PAX2 expression associated with DNA mismatch repair deficiency in endometrial endometrioid adenocarcinoma.

AIMS: Loss of expression of tumour suppressor PAX2 and MMR deficiency (dMMR) has been frequently seen in endometrial endometrioid adenocarcinoma (EEC). However, the relationship between PAX2 expression and MMR status is unknown. METHODS AND RESULTS: We studied the PAX2 expression and examined its association with MMR status at the protein and genetic levels in 180 cases of EEC. Overall, total loss of PAX2 expression was found in about 70%, while retained PAX2 expression was seen in 30% of EEC. Among 125 cases with loss of PAX2, 68.8% were found in EECs with pMMR, while 31.2% were seen in those with dMMR. Among 55 cases of EECs with retained PAX2 expression, 92.7% were EECs with dMMR and 7.3% were those with pMMR (P < 0.001). While dMMR cases with MLH1 hypermethylation show almost equal retained or loss of PAX2 expression (52% versus 48%), dMMR with genetic alterations had significantly more retained PAX2 expression than loss of PAX2 (92.3% versus 7.7%), regardless of somatic or germline mutations. Loss of PAX2 was observed in 97.3% of dMMR with MLH1 hypermethylation compared to 2.7% of dMMR with genetic alterations (P < 0.001). Aggressive features such as higher tumour grades (FIGO 2-3) and advanced clinical stage (T2-T4) were significantly more frequently seen in dMMR with retained PAX2 expression, compared those to pMMR with loss of PAX2 expression. CONCLUSION: Our study demonstrates a close correlation between retained PAX2 expression and dMMR in EEC. The molecular mechanism and clinical significance linking these two pathways in EEC remains to be unravelled.

Immunohistochemical co-expression of PAX2 and CAIX predicts better prognosis in clear cell renal cell carcinoma after nephrectomy: A retrospective observational study.

Clear cell renal cell carcinoma (ccRCC) is a lethal malignancy with high metastatic probability. Paired box 2 gene product (PAX2) carbonic anhydrase IX were biomolecules closely linked with ccRCC development and outcomes of multiple malignancies. We aim to explore the role of immunohistochemical staining of PAX2 and CAIX to predict ccRCC prognosis after nephrectomy. Surgical specimens of patients who were pathologically diagnosed as ccRCC were reviewed. Expression levels of PAX2 and CAIX were assessed via immunohistochemical staining. Recurrence-free survival (RFS) and overall survival were compared among different phenotypes. Inverse probability of treatment weighting (IPTW) was used for adjustment of confounding factors. 56 patients were included. Patients with PAX2 and CAIX high-expression (the two-high group, n=8) had significantly longer RFS and OS than those of simultaneously down-expression (the two-low group, n=31). Median RFS was 38.4 (95% CI: 32.3-NA) for the two-high group and 14.8 (95% CI: 13.4-39.0) months for the two-low group (P=0.043). IPTW confirmed PAX2 and CAIX co-expression is associated with less recurrence risk HR: 0.39, 95% CI: 0.17-0.92, P=0.031). Co-expression of PAX2 and CAIX is associated better prognosis of ccRCC. We are looking for validation by large cohort studies.

Renal coloboma syndrome/dominant optic atrophy with severe retinal atrophy and de novo digenic mutations in PAX2 and OPA1.

Renal coloboma syndrome (RCS) and dominant optic atrophy are mainly caused by heterozygous mutations in PAX2 and OPA1, respectively. We describe a patient with digenic mutations in PAX2 and OPA1. A female infant was born without perinatal abnormalities. Magnetic resonance imaging at 4 months of age showed bilateral microphthalmia and optic nerve hypoplasia. Appropriate body size was present at 2 years of age, and mental development was favorable. Color fundus photography revealed severe retinal atrophy in both eyes. Electroretinography showed slight responses in the right eye, but no responses in the left eye, suggesting a high risk of blindness. Urinalysis results were normal, creatinine-based estimated glomerular filtration rate was 63.5 mL/min/1.73 m2, and ultrasonography showed bilateral hypoplastic kidneys. Whole exome sequencing revealed de novo frameshift mutations in PAX2 and OPA1. Both variants were classified as pathogenic (PVS1, PS2, PM2) based on the guidelines from the American College of Medical Genetics and Genomics (ACMG). Genetic testing for ocular diseases should be considered for patients with suspected RCS and a high risk of total blindness.

The Role of the PAX Genes in Renal Cell Carcinoma.

Renal cell carcinoma (RCC) is a significant oncological challenge due to its heterogeneous nature and limited treatment options. The PAX developmental gene family encodes nine highly conserved transcription factors that play crucial roles in embryonic development and organogenesis, which have been implicated in the occurrence and development of RCC. This review explores the molecular landscape of RCC, with a specific focus on the role of the PAX gene family in RCC tumorigenesis and disease progression. Of the various RCC subtypes, clear cell renal cell carcinoma (ccRCC) is the most prevalent, characterized by the loss of the von Hippel-Lindau (VHL) tumor suppressor gene. Here, we review the published literature on the expression patterns and functional implications of PAX genes, particularly PAX2 and PAX8, in the three most common RCC subtypes, including ccRCC, papillary RCC (PRCC), and chromophobe RCC (ChRCC). Further, we review the interactions and potential biological mechanisms involving PAX genes and VHL loss in driving the pathogenesis of RCC, including the key signaling pathways mediated by VHL in ccRCC and associated mechanisms implicating PAX. Lastly, concurrent with our update regarding PAX gene research in RCC, we review and comment on the targeting of PAX towards the development of novel RCC therapies.

Fgf, Hh, and pax2a differentially regulate expression of pax5 and pou3f3b in vestibular and auditory maculae in the zebrafish otic vesicle.

BACKGROUND: The vertebrate inner ear contains distinct sensory epithelia specialized for auditory or vestibular function. In zebrafish, the first sensory epithelia form at opposite ends of the otic vesicle and are functionally distinct: the anterior utricular macula is essential for vestibular function whereas the posterior saccular macula is critical for hearing. Mechanisms distinguishing these maculae are not clear. Here, we examined the effects of manipulating Fgf or Hh on expression of pax5 and pou3f3b, unique markers of utricular and saccular identity. We also examined the roles of pax2a and atoh1a/b, early regulators of sensory specification. RESULTS: fgf3 and fgf8a were uniquely required for pax5 and pou3f3b, respectively. Elevating Fgf or blocking Hh expanded expression of pax5 but repressed pou3f3b, while blocking Fgf had the opposite effect. Blocking sensory specification did not affect pax5 or pou3f3b, but both markers were lost in pax2a-/- mutants. Maintenance of pax2a expression requires Fgf, Hh and Pax2a itself. CONCLUSION: Specification of utricular identity requires high Fgf and is repressed by Hh, whereas saccular identity requires Hh plus low Fgf. pax2a acts downstream of Fgf and Hh to maintain both fates. Comparison with mouse suggests this may reflect a broadly conserved developmental mechanism.

Pax2a, Sp5a and Sp5l act downstream of Fgf and Wnt to coordinate sensory-neural patterning in the inner ear.

In zebrafish, sensory epithelia and neuroblasts of the inner ear form simultaneously in abutting medial and lateral domains, respectively, in the floor of the otic vesicle. Previous studies support regulatory roles for Fgf and Wnt, but how signaling is coordinated is poorly understood. We investigated this problem using pharmacological and transgenic methods to alter Fgf or Wnt signaling from early placodal stages to evaluate later changes in growth and patterning. Blocking Fgf at any stage reduces proliferation of otic tissue and terminates both sensory and neural specification. Wnt promotes proliferation in the otic vesicle but is not required for sensory or neural development. However, sustained overactivation of Wnt laterally expands sensory epithelia and blocks neurogenesis. pax2a, sp5a and sp5l are coregulated by Fgf and Wnt and show overlapping expression in the otic placode and vesicle. Gain- and loss-of-function studies show that these genes are together required for Wnt's suppression of neurogenesis, as well as some aspects of sensory development. Thus, pax2a, sp5a and sp5l are critical for mediating Fgf and Wnt signaling to promote spatially localized sensory and neural development.

Dynamics of muscle growth and regeneration: Lessons from the teleost.

The processes of myogenesis during both development and regeneration share a number of similarities across both amniotes and teleosts. In amniotes, the process of muscle formation is considered largely biphasic, with developmental myogenesis occurring through hyperplastic fibre deposition and postnatal muscle growth driven through hypertrophy of existing fibres. In contrast, teleosts continue generating new muscle fibres during adult myogenesis through a process of eternal hyperplasia using a dedicated stem cell system termed the external cell layer. During developmental and regenerative myogenesis alike, muscle progenitors interact with their niche to receive cues guiding their transition into myoblasts and ultimately mature myofibres. During development, muscle precursors receive input from neighbouring embryological tissues; however, during repair, this role is fulfilled by other injury resident cell types, such as those of the innate immune response. Recent work has focused on the role of macrophages as a pro-regenerative cell type which provides input to muscle satellite cells during regenerative myogenesis. As zebrafish harbour a satellite cell system analogous to that of mammals, the processes of regeneration can be interrogated in vivo with the imaging intensive approaches afforded in the zebrafish system. This review discusses the strengths of zebrafish with a focus on both the similarities and differences to amniote myogenesis during both development and repair.

PAX2 Gene Mutation in Pediatric Renal Disorders-A Narrative Review.

The PAX2 gene is a transcription factor that is essential for the development of the urinary system among other transcription factors. The role of PAX2 is highlighted from the seventh week of gestation, when it is involved in development processes and the emergence of nephrons and collecting tubes. Being an important factor in renal development, mutations of this gene can produce severe alterations in the development of the urinary tract, namely congenital anomalies of the kidneys and urinary tract. The first reported cases described with the PAX2 mutation included both renal anomalies and the involvement of other organs, such as the eyes, producing renal coloboma syndrome. Over the years, numerous cases have been reported, including those with only renal and urinary tract anomalies. The aim of this review is to present a summary of pediatric patients described to have mutations in the PAX2 gene to contribute to a better understanding of the genetic mechanism causing anomalies of the kidneys and urinary tract. In this review, we have included only pediatric cases with renal and urinary tract disorders, without the involvement of other organs. From what we know so far from the literature, this is the first review gathering pediatric patients presenting the PAX2 mutation who have been diagnosed exclusively with renal and urinary tract disorders.

PAX2 and CAKUT Phenotypes: Report on Two New Variants and a Review of Mutations from the Leiden Open Variation Database.

PAX2 is a transcription factor expressed during embryogenesis in the eye, ear, CNS, and genitourinary tract, and is one of the major regulators of kidney development. Mutations in this gene are associated with papillorenal syndrome (PAPRS), a genetic condition characterized by optic nerve dysplasia and renal hypo/dysplasia. In the last 28 years, many cohort studies and case reports highlighted PAX2's involvement in a large spectrum of kidney malformations and diseases, with or without eye abnormalities, defining the phenotypes associated with PAX2 variants as "PAX2-related disorders". Here, we reported two new sequence variations and reviewed PAX2 mutations annotated on the Leiden Open Variation Database 3.0. DNA was extracted from the peripheral blood of 53 pediatric patients with congenital abnormalities of the kidney and urinary tract (CAKUT). PAX2 gene-coding exonic and flanking intronic regions were sequenced with Sanger technology. Two unrelated patients and two twins carrying one known and two unknown PAX2 variations were observed. The frequency of PAX2-related disorders in this cohort was 5.8%, considering all CAKUT phenotypes (16.7% in the PAPRS phenotype and 2.5% in non-syndromic CAKUT). Although PAX2 mutations have a higher frequency in patients with PAPRS or non-syndromic renal hypoplasia, from the review of variants reported to date in LOVD3, PAX2-related disorders are detected in pediatric patients with other CAKUT phenotypes. In our study, only one patient had a CAKUT without an ocular phenotype, but his twin had both renal and ocular involvement, confirming the extreme inter- and intrafamilial phenotypic variability.

Co-Expression of Multiple PAX Genes in Renal Cell Carcinoma (RCC) and Correlation of High PAX Expression with Favorable Clinical Outcome in RCC Patients.

Renal cell carcinoma (RCC) is the most common form of kidney cancer, consisting of multiple distinct subtypes. RCC has the highest mortality rate amongst the urogenital cancers, with kidney renal clear cell carcinoma (KIRC), kidney renal papillary cell carcinoma (KIRP), and kidney chromophobe carcinoma (KICH) being the most common subtypes. The Paired-box (PAX) gene family encodes transcription factors, which orchestrate multiple processes in cell lineage determination during embryonic development and organogenesis. Several PAX genes have been shown to be expressed in RCC following its onset and progression. Here, we performed real-time quantitative polymerase chain reaction (RT-qPCR) analysis on a series of human RCC cell lines, revealing significant co-expression of PAX2, PAX6, and PAX8. Knockdown of PAX2 or PAX8 mRNA expression using RNA interference (RNAi) in the A498 RCC cell line resulted in inhibition of cell proliferation, which aligns with our previous research, although no reduction in cell proliferation was observed using a PAX2 small interfering RNA (siRNA). We downloaded publicly available RNA-sequencing data and clinical histories of RCC patients from The Cancer Genome Atlas (TCGA) database. Based on the expression levels of PAX2, PAX6, and PAX8, RCC patients were categorized into two PAX expression subtypes, PAXClusterA and PAXClusterB, exhibiting significant differences in clinical characteristics. We found that the PAXClusterA expression subgroup was associated with favorable clinical outcomes and better overall survival. These findings provide novel insights into the association between PAX gene expression levels and clinical outcomes in RCC patients, potentially contributing to improved treatment strategies for RCC.

Pax2a, but not pax2b, influences cell survival and periocular mesenchyme localization to facilitate zebrafish optic fissure closure.

BACKGROUND: Pax2 is required for optic fissure development in many organisms, including humans and zebrafish. Zebrafish loss-of-function mutations in pax2a display coloboma, yet the etiology of the morphogenetic defects is unclear. Further, pax2 is duplicated in zebrafish, and a role for pax2b in optic fissure development has not been examined. RESULTS: Using a combination of imaging and molecular genetics, we interrogated a potential role for pax2b and examined how loss of pax2 affects optic fissure development. Although optic fissure formation appears normal in pax2 mutants, an endothelial-specific subset of periocular mesenchyme (POM) fails to initially localize within the optic fissure, yet both neural crest and endothelial-derived POM ectopically accumulate at later stages in pax2a and pax2a; pax2b mutants. Apoptosis is not up-regulated within the optic fissure in pax2 mutants, yet cell death is increased in tissues outside of the optic fissure, and when apoptosis is inhibited, coloboma is partially rescued. In contrast to pax2a, loss of pax2b does not appear to affect optic fissure morphogenesis. CONCLUSIONS: Our results suggest that pax2a, but not pax2b, supports cell survival outside of the optic fissure and POM abundance within it to facilitate optic fissure closure.

[Genetic analysis of a Chinese pedigree with chronic kidney disease due to variant of PAX2 gene].

OBJECTIVE: To explore the genetic basis of a Chinese pedigree affected with chronic kidney disease (CKD). METHODS: A Chinese pedigree comprised of 10 individuals from four generation who had visited the First Affiliated Hospital of Dali University from August 15, 2018 to July 5, 2021 was selected as the study subject. Clinical data of the proband were collected, and a pedigree survey was conducted. The proband was subjected to whole exome sequencing (WES). Candidate variant was verified by Sanger sequencing and bioinformatic analysis. RESULTS: The proband, a 41-year-old female, has been diagnosed with chronic nephritis for more than 4 years. Routine urinary examination showed proteinuria and blood creatinine of 1 130 µmol/L. Renal biopsy has revealed hyperplastic glomerulonephritis, moderate tubulointerstitial disease and renal arteriosclerosis. Her elder sister, younger brother, younger sister and mother were all diagnosed with CKD stage 5. Except for her elder sister, all of them had deceased, whilst no abnormality was found in the remainders. Genetic testing revealed that the proband and four family members had harbored a c.467G>A missense variant of the PAX2 gene. The variant has been associated with focal segmental glomerulosclerosis and classified as likely pathogenic (PS1+PP3+PP4) based on the guidelines from the American College of Medical Genetics and Genomics (ACMG). CONCLUSION: The c.167G＞A variant of the PAX2 gene probably underlay the CKD in this Chinese pedigree.

Multiple roles for Pax2 in the embryonic mouse eye.

The vertebrate eye anlage grows out of the brain and folds into bilayered optic cups. The eye is patterned along multiple axes, precisely controlled by genetic programs, to delineate neural retina, pigment epithelium, and optic stalk tissues. Pax genes encode developmental regulators of key morphogenetic events, with Pax2 being essential for interpreting inductive signals, including in the eye. PAX2 mutations cause ocular coloboma, when the ventral optic fissure fails to close. Previous studies established that Pax2 is necessary for fissure closure and to maintain the neural retina -- glial optic stalk boundary. Using a Pax2GFP/+ knock-in allele we discovered that the mutant optic nerve head (ONH) lacks molecular boundaries with the retina and RPE, rendering the ONH larger than normal. This was preceded by ventronasal cup mispatterning, a burst of overproliferation and followed by optic cup apoptosis. Our findings support the hypothesis that ONH cells are tripotential, requiring Pax2 to remain committed to glial fates. This work extends current models of ocular development, contributes to broader understanding of tissue boundary formation and informs the underlying mechanisms of human coloboma.

The proliferative and the antifibrotic side of PAX2 in tubular repair.

Regenerative repair following injury to proximal tubular epithelial cells (PTECs) is essential to restore the kidney to normal function in acute kidney injury. Failure to accomplish this leads to chronic kidney disease. Expression of the paired-box transcription factor Pax2 in PTECs is required for their regenerative proliferation and repair. However, a loss-of-function study now shows that the absence of Pax2 not only impacts PTEC proliferation but also causes myofibroblast recruitment leading to excessive tubulointerstitial fibrosis.

Single cell dynamics of embryonic muscle progenitor cells in zebrafish.

Muscle stem cells hold a great therapeutic potential in regenerating damaged muscles. However, the in vivo behavior of muscle stem cells during muscle growth and regeneration is still poorly understood. Using zebrafish as a model, we describe the in vivo dynamics and function of embryonic muscle progenitor cells (MPCs) in the dermomyotome. These cells are located in a superficial layer external to muscle fibers and express many extracellular matrix (ECM) genes, including collagen type 1 α2 (col1a2). Utilizing a new col1a2 transgenic line, we show that col1a2+ MPCs display a ramified morphology with dynamic cellular processes. Cell lineage tracing demonstrates that col1a2+ MPCs contribute to new myofibers in normal muscle growth and also during muscle regeneration. A combination of live imaging and single cell clonal analysis reveals a highly choreographed process of muscle regeneration. Activated col1a2+ MPCs change from the quiescent ramified morphology to a polarized and elongated morphology, generating daughter cells that fuse with existing myofibers. Partial depletion of col1a2+ MPCs severely compromises muscle regeneration. Our work provides a dynamic view of embryonic muscle progenitor cells during zebrafish muscle growth and regeneration.

Large-scale death of retinal astrocytes during normal development is non-apoptotic and implemented by microglia.

Naturally occurring cell death is a fundamental developmental mechanism for regulating cell numbers and sculpting developing organs. This is particularly true in the nervous system, where large numbers of neurons and oligodendrocytes are eliminated via apoptosis during normal development. Given the profound impact of death upon these two major cell populations, it is surprising that developmental death of another major cell type-the astrocyte-has rarely been studied. It is presently unclear whether astrocytes are subject to significant developmental death, and if so, how it occurs. Here, we address these questions using mouse retinal astrocytes as our model system. We show that the total number of retinal astrocytes declines by over 3-fold during a death period spanning postnatal days 5-14. Surprisingly, these astrocytes do not die by apoptosis, the canonical mechanism underlying the vast majority of developmental cell death. Instead, we find that microglia engulf astrocytes during the death period to promote their developmental removal. Genetic ablation of microglia inhibits astrocyte death, leading to a larger astrocyte population size at the end of the death period. However, astrocyte death is not completely blocked in the absence of microglia, apparently due to the ability of astrocytes to engulf each other. Nevertheless, mice lacking microglia showed significant anatomical changes to the retinal astrocyte network, with functional consequences for the astrocyte-associated vasculature leading to retinal hemorrhage. These results establish a novel modality for naturally occurring cell death and demonstrate its importance for the formation and integrity of the retinal gliovascular network.

PAX2 promoter methylation and AIB1 overexpression promote tamoxifen resistance in breast carcinoma patients.

INTRODUCTION: Disease recurrence is an important obstacle in estrogen receptor positive (ER+) tamoxifen treated breast carcinoma patients. Tamoxifen resistance-related molecular mechanisms are not fully understood. Alteration in DNA methylation which contributes to transcriptional regulation of cancer-related genes plays a crucial role in tamoxifen response. In the present study, the contribution of promoter methylation and mRNA expression of PAX2 and AIB1 in the development of breast carcinoma and tamoxifen refractory was assessed. METHODS: Methylation specific-high resolution melting (MS-HRM) analysis and Real-time quantitative PCR (RT-qPCR) experiment were performed to analyze the promoter methylation and mRNA expression levels of PAX2 and AIB1 genes in 102 breast tumors and adjacent normal breast specimens. RESULTS: We indicated that PAX2 expression is decreased in breast tissues due to hypermethylation in its promoter region. Compared to the adjacent normal tissues, the tumors exhibited significantly lower relative mRNA levels of PAX2 and increased expression of AIB1. Aberrant promoter methylation of PAX2 and overexpression of AIB1 was observed in tamoxifen resistance patients compared to the sensitive ones. Cox regression analysis exhibited that the increased promoter methylation status of PAX2 and overexpression of AIB1 remained as unfavorable identifiers which influence patients' survival independently. CONCLUSIONS: Our results revealed that the aberration in PAX2 promoter methylation and AIB1 overexpression are associated with the tamoxifen response in breast carcinoma patients. Further research is needed to demonstrate the potential of using PAX2 and AIB1 expression and their methylation-mediated regulation as predictive or prognostic biomarkers or as a new target therapy for better disease management.