American Journal of Pathology最新文献

Epithelial barrier dysfunction is a hallmark of inflammatory bowel diseases; however, the mechanisms underlying such impairment remain incompletely understood. In the present study, a dextran sulfate sodium-induced colitis model was used to investigate how the inflammatory environment damages the intestinal mucosa. The results demonstrated that colitogenic ambiance enhances intestinal epithelial cell death, delays epithelial cell proliferation, and exacerbates mucosal erosion. Unexpectedly, this work identified a previously unrecognized role for Notch signaling in mediating these effects. Specifically, the colitogenic milieu reduces Notch/mechanistic target of rapamycin complex 1 (mTORC1)-mediated intestinal epithelial cell proliferation to promote goblet cell differentiation. Chemical activation of Notch signaling stimulated intestinal epithelial cell proliferation and reduced goblet cell differentiation in the colitic mucosa, further aggravating mucosal damage. Conversely, inhibition of Notch or mTORC1 signaling during mucosal repair reduced intestinal epithelial cell proliferation and enhanced goblet cell differentiation, corroborating the implication of Notch and mTORC1 signaling in both processes. Collectively, these findings uncover a context-dependent role for the Notch-mechanistic target of rapamycin axis in regulating intestinal epithelial cell proliferation and differentiation in the colitic mucosa and suggest that its targeted modulation may hold therapeutic potential in inflammatory bowel diseases.

Insights into how normal epithelial cells adapt to microenvironmental perturbations may reveal molecular vulnerabilities that become obscured later in carcinogenesis, and hypoxia is common in colorectal cancer (CRC). Although colon mucosa exists in a state of physiological hypoxia and is susceptible to ischemic injury, normal colon epithelial adaptive responses to changes in oxygenation are largely uncharacterized. In this study, human colon organoids (colonoids) were subjected to sustained hypoxia in vitro with characterization of consequent phenotypes and transcriptional changes. Hypoxia tolerance in human colonoids resulted in robust down-regulation of alcohol dehydrogenase 1C (ADH1C), which was also validated in archival tissue from patients with ischemic colitis. ADH1C transcripts revealed a nonuniform expression pattern in normal colon epithelium, with enrichment in transit-amplifying and progenitor epithelial cells. Ectopic expression of ADH1C in colonoids subjected to hypoxia increased reactive oxygen species and reduced NADPH compared with those in normoxia, suggesting that hypoxia-induced ADH1C down-regulation facilitates neutralization of reactive oxygen species. Hypoxia-induced ADH1C down-regulation also showed reduced transit-amplifying cell signatures and increased expression of regeneration-associated stem cell marker FGFBP1. Finally, ADH1C-low CRC showed significant enrichment for hypoxia-associated colon epithelial signatures compared with ADH1C-high CRC. Taken together, these results establish ADH1C as a mediator of colon epithelial hypoxia responses and epithelial identity with relevance to human CRC.

Corneal ectasias are a significant cause of vision morbidity worldwide. In humans, corneal ectasias are characterized by tissue mechanical weakening, stromal thinning, and bulging. Previous histopathology studies showed a high rate of keratocyte apoptosis in corneas with ectasia. A mouse model expressing a keratocyte lineage-specific reporter KeraRT/tetO-Cre/mTmG/DTR was created to elucidate the roles of keratocyte death in the development of corneal ectasias. This mouse model allows selective death of keratocytes at chosen times during stromal development and in mature stromas. Slit-lamp examination as well as histopathology and advanced imaging techniques were used to assess stromal structure after keratocyte genetic ablation. It was found that genetic ablation of keratocytes in the first 20 days after birth induces corneal thinning and ectasia. A corneal hydrops-like phenotype (severe ectasia) occurred more frequently if keratocyte death was induced in the first week after birth. Inducing keratocyte death at an age where some degree of corneal maturation has occurred, >3 weeks of age, did not create changes in corneal thickness, transparency, or curvature, or any noticeable abnormalities in microstructure.

Neuroblastoma (NB) with metastasis to bone marrow (BM) usually results in dismal survival. The mechanisms underlying BM metastasis remain largely unclear. In this study, single-cell transcriptome of NB with and without BM metastasis were analyzed, and 146 marker genes of NB cells were identified. Using two-sample Mendelian randomization, N-cadherin (CDH2) and tubulin α 1A (TUBA1A) were identified as key genes possibly affecting BM metastasis. Functional investigations suggest that CDH2 regulates cell cycle and DNA replication, whereas TUBA1A impacts cell adhesion molecules and the cAMP signaling pathway. Dysregulated expression of CDH2 or TUBA1A also alters the immune microenvironment by immune cells and chemokines. Subsequent experiments validated that both CDH2 and TUBA1A were up-regulated in NB cell lines and high-risk patient samples, compared with low-risk cases. Knockdown of CDH2 or TUBA1A led to cell cycle arrest and significantly inhibited NB cell proliferation and migration. Moreover, an oxidation-reduction cycling agent, 2,3-dimethoxy-1,4-naphthoquinone, was identified as a candidate compound with inhibitory activity on NB cells by diminishing CDH2 or TUBA1A expression levels. Together, these results highlight CDH2 and TUBA1A as novel therapeutic targets for NB with BM metastases. The efficacy of 2,3-dimethoxy-1,4-naphthoquinone in suppressing NB cell growth and CDH2/TUBA1A expression suggests its potential for clinical application.

This study evaluated the function of peroxisome proliferator-activated receptor α (PPARα) in vascular endothelial cells (ECs) under physiological and disease conditions. Vascular density and avascular area were evaluated in Griffonia simplicifolia isolectin B4-stained retinas. Endothelial progenitor cells were quantified using flow cytometry. Vascular leakage was evaluated by Evans Blue. The mitochondrial function and morphology were evaluated by a Seahorse analyzer and immunofluorescence staining. Cell senescence was assessed by a senescence-associated β-galactosidase activity assay and Western blot analysis. A significant reduction in the retinal vessel length and vascular mesh density was found in EC-specific PPARα conditional knockout (PPARα^ECKO) mice. Relative to PPARa^flox-KO mice with oxygen-induced retinopathy (OIR), PPARα^ECKO OIR retina showed enlarged avascular areas and decreased endothelial progenitor cell number, whereas EC-specific PPARα conditional transgenic mice showed reduced avascular areas in the OIR retina. Compared with diabetic PPARa^flox-KO mice, diabetic PPARα^ECKO mice showed declined electroretinographic amplitudes, decreased retinal thickness, and increased retinal vascular leakage. PPARα deficiency exacerbated, whereas PPARα activation alleviated, EC mitochondrial dysfunction induced by diabetic stressors. PPARα^-/- ECs developed senescence, prominent oxidant-induced mitochondria fragmentation, and down-regulation of translocase of outer mitochondrial membrane 20 and peroxisome proliferator-activated receptor γ coactivator 1α, relative to wild-type ECs. These results suggest that PPARα in microvascular ECs regulates retinal vascular development and protects ECs against diabetes/hypoxia-induced vascular dysfunction through mitochondrial protective and anti-senescence activities.

Neonatal thymectomy (TX) has been known to induce experimental autoimmune disease models in mice for over half a century. The thymic microenvironment, including thymic epithelial cells (TECs), plays a crucial role in establishing self-tolerance in T cells. However, the extent to which the dynamic changes in the neonatal thymic environment contribute to the onset of autoimmunity remains incompletely understood. In this study, the detailed alterations in the neonatal thymus and peripheral lymphoid organs were analyzed using a mouse model of primary Sjögren disease. Mice treated with TX at 3 days after birth (day 3-TX) exhibited significantly more severe autoimmune pathology than those treated with TX at 7 days after birth. Around day 3, T-cell differentiation and the expansion of TECs, particularly medullary TECs, were markedly accelerated in the neonatal thymus. Furthermore, in day 3-TX mice, the expansion of peripherally induced regulatory T (Treg) cells was impaired, along with the loss of thymic-derived Treg cell output that typically undergoes robust expansion around day 3 after birth. The suppressive activity of Treg cells from day 3-TX mice was significantly impaired compared with that of control Treg cells. These findings suggest that the neonatal thymic environment plays a critical role in regulating peripheral immune tolerance and may influence the pathogenesis of autoimmune diseases.

Ameloblastoma (AM), a locally aggressive odontogenic tumor, exhibits elusive pathogenesis. Here, optic atrophy 1 (OPA1)-mediated mitochondrial hyperfusion was identified as a driver of tumor stemness and progression. Single-cell transcriptomics of primary AM specimens revealed mitochondrial fusion^High epithelial subpopulations exhibiting enriched stemness pathways. A striking up-regulation of OPA1 was observed in AM tissues, establishing a robust correlation between elevated OPA1 expression and up-regulated stemness markers, whereas functional experiments demonstrated that OPA1 overexpression amplifies self-renewal capacity and invasive aggression in hTERT⁺-AM cells. Mechanistically, mitochondrial hyperfusion suppresses Hippo signaling, enabling yes-associated protein 1 (YAP1) nuclear translocation and TEAD-dependent transcription. OPA1-overexpressing cells exhibited robust nuclear YAP1 enrichment, driving stem-like expansion. Critically, clinical analysis established OPA1^High tumors as having elevated growth rates, consolidating mitochondrial hyperfusion as a prognostic determinant. Therapeutically, MYLS22-a first-in-class OPA1 inhibitor-suppressed mitochondrial hyperfusion and reduced stemness in patient-derived organoids. The present work unveils an OPA1-mediated mitochondrial fusion-YAP1 nuclear translocation axis as the cornerstone of AM stemness, proposing OPA1 as a druggable target for this recalcitrant tumor.

Renal fibrosis is the main pathologic change observed with the progression of chronic kidney disease (CKD), which predicts kidney outcomes. The ability to detect fibrosis early in the disease course may be crucial to identify those at the highest risk of CKD progression. Clinical studies have observed increased expression of serum matrix Gla protein (MGP), a potent inhibitor of soft tissue calcification, in patients with CKD. In a cross-sectional study of patients with CKD, serum MGP levels were found to be associated with albuminuria and waist circumference after controlling for kidney function, which modified the association between MGP and albuminuria. To examine the impact of MGP on the onset and progression of CKD, various mouse models were used in the current study. Using Cre-reporter, Rosa^Tomato;Mgp-Cre mice and a new knock-in model expressing hemagglutinin epitope-tagged MGP, it was identified that pericytes in healthy kidneys and myofibroblasts in the folic acid (FA)-injured kidneys are the primary sources of MGP production. FA injection in Mgp^-/- mice induced significantly less renal fibrosis in comparison to the control mice because of a reduced number of pericytes and attenuated Notch signaling. In a complementary experiment, restoration of Mgp expression in myofibroblasts in Mgp^-/- mice leads to renal fibrosis as severe as in control mice. This work suggests that MGP expression in myofibroblasts exacerbates renal fibrosis in FA-injured kidneys.

Ischemic acute kidney injury may accelerate the progression to end-stage renal disease. Megalin has been shown to shuttle stanniocalcin (STC)-1 (which promotes mitochondrial antioxidant defenses) to the mitochondria through the retrograde-early endosomes-to-Golgi pathway; knockout of megalin in cultured cells has been reported to impair glycolysis and mitochondrial respiration. This study sought to determine kidney phenotype after ischemia/reperfusion kidney injury in mice with tubular epithelium-specific deletion of megalin. Mice (on C57B/6 background) with conditional tubular epithelium-specific knockout of megalin (tLrp2KO) and mice with combined conditional tubular epithelium-specific knockout of megalin and overexpression of STC1 (tLrp2KO;tSTC1O) were subjected to ischemia (clamping of renal pedicles), followed by reperfusion for 1, 3, 10, 45, and 90 days. Serum creatinine was measured and kidneys were harvested for analysis. After ischemia/reperfusion (I/R) and compared with control mice, tLrp2KO mice displayed worse acute kidney injury, severe and persistent inflammation, diminished tubular epithelial cell proliferation, up-regulation of TGFβ1 signaling, fibrosis, and accelerated progression to chronic kidney disease. Kidney injury was not rescued in tLrp2KO;tSTC1O mice, consistent with megalin-dependent renal protection by STC1. Freshly isolated proximal tubule fragments from tLrp2KO mice or cultured proximal tubule epithelial cells with megalin knockout displayed activation of TGFβ1 signaling, consistent with modulation of TGFβ1 signaling by megalin. In conclusion, tubular epithelium-specific deletion of megalin aggravates ischemia/reperfusion kidney injury, up-regulates TGFβ1 signaling, and accelerates chronic kidney disease progression.