Biology subjects

Uchimura, K.

Publications and source records attributed to Uchimura, K..

7 recordsLinked to original sources

bioRxiv2025

Amyloid-β fibrils accumulated in preeclamptic placentas suppress syncytialization of cytotrophoblasts

Cerebral deposition of fibrillar amyloid-{beta} (A{beta}) is a pathological hallmark of Alzheimers disease. While A{beta} is present in human placentas and accumulates in preeclamptic placentas characterized by poor placentation, the production and role of A{beta} in the human placenta remain unclear. Because hypoxia in mid-to-late pregnancy is a risk for preeclampsia, we found that levels of hypoxia-inducible factor 1- and {beta}-secretase (BACE-1) increased concurrently with placental A{beta} deposition in late stage preeclamptic placentas. We also found that a human cytotrophoblast (CTB) model, BeWo cells, actually produced A{beta} species, and that hypoxia increased A{beta} production and BACE-1 protein levels. A{beta}42 fibrils inhibited CTB syncytialization, a critical step in maintaining pregnancy, by inducing loss of membrane localization of cell-cell adhesion molecules. Primary human CTBs confirmed these observations. Taken together, our results suggest that increased A{beta} production in CTBs by hypoxia may lead to the formation of A{beta} fibrils, which inhibit syncytiotrophoblast formation and are detrimental to pregnancy. Thus, our results reveal the novel role of A{beta} fibrils in the pathogenesis of preeclampsia.

biochemistry↗

bioRxiv2023

Structural flexibility of apolipoprotein E-derived arginine-rich peptides improves their cell penetration capability

Amphipathic arginine-rich peptide, A2-17, exhibits moderate perturbation of lipid membranes and the highest cell penetration among its structural isomers. We investigated the direct cell-membrane penetration mechanism of the A2-17 peptide. We designed structurally constrained versions of A2-17, stapled (StpA2-17) and stitched (StchA2-17), whose -helical conformations were stabilized by chemical crosslinking. Circular dichroism confirmed that StpA2-17 and StchA2-17 had higher -helix content than A2-17 in aqueous solution. Upon liposome binding, only A2-17 exhibited a coil-to-helix transition. Confocal microscopy revealed that A2-17 had higher cell penetration efficiency than StpA2-17 in HeLa cells. Partitioning into lipid membranes was more prominent for StchA2-17 than for A2-17 or StpA2-17; StchA2-17 remained on the cell membrane without cell penetration. Tryptophan fluorescence analysis suggested that A2-17 and its analogs had similar membrane-insertion positions between the interface and hydrophobic core. Atomic force microscopy demonstrated that A2-17 reduced the mechanical rigidity of liposomes to a greater extent than StpA2-17 and StchA2-17. Finally, electrophysiological analysis showed that A2-17 induced a higher charge influx through transient pores in a planer lipid bilayer than StpA2-17 and StchA2-17. These findings indicate that structural flexibility, which enables diverse conformations of A2-17, leads to a membrane perturbation mode that contributes to cell membrane penetration.

biophysics↗

bioRxiv2021

Extracellular endosulfatase Sulf-2 harbours a chondroitin/dermatan sulfate chain that modulates its enzyme activity

Sulfs represent a class of unconventional sulfatases, which differ from all other members of the sulfatase family by their structures, catalytic features and biological functions. Through their specific endosulfatase activity in extracellular milieu, Sulfs provide an original post-synthetic regulatory mechanism for heparan sulfate complex polysaccharides and have been involved in multiple physiopathological processes, including cancer. However, Sulfs remain poorly characterized enzymes, with major discrepancies regarding their in vivo functions. Here we show that human Sulf-2 (HSulf-2) features a unique polysaccharide post-translational modification. We identified a chondroitin/dermatan sulfate glycosaminoglycan (GAG) chain, attached to the enzyme substrate binding domain. We found that this GAG chain affects enzyme/substrate recognition and tunes HSulf-2 activity in vitro and in vivo using a mouse model of tumorigenesis and metastasis. In addition, we showed that mammalian hyaluronidase acted as a promoter of HSulf-2 activity by digesting its GAG chain. In conclusion, our results highlight HSulf-2 as a unique proteoglycan enzyme and its newly-identified GAG chain as a critical non-catalytic modulator of the enzyme activity. These findings contribute in clarifying the conflicting data on the activities of the Sulfs and introduce a new paradigm into the study of these enzymes.

biochemistry↗

bioRxiv2020

Stepwise transmigration cascade of T and B cells through the perivascular channel in lymph node high endothelial venules

High endothelial venules (HEVs) effectively recruit circulating lymphocytes from the blood to lymph nodes. HEVs have endothelial cells (ECs) and perivascular sheaths consisting of fibroblastic reticular cells (FRCs). Many studies have characterized the multiple steps of lymphocyte migration interacting with ECs at the luminal side of HEVs. However, post-luminal migration steps are not well elucidated. Herein, we performed intravital imaging to investigate post-luminal T and B cell migration, consisting of trans-EC migration, crawling in the perivascular channel (a narrow space between ECs and FRCs) and trans-FRC migration. The post-luminal migration of T cells occurred in a PNAd-dependent manner. Remarkably, we found hot spots for the trans-EC and trans-FRC migration of T and B cells. Interestingly, T and B cells preferentially shared trans-FRC migration hot spots but not trans-EC migration hot spots. Furthermore, the trans-FRC T cell migration was confined to fewer sites than trans-EC T cell migration, and trans-FRC migration of T and B cells preferentially occurred at FRCs covered by CD11c+ dendritic cells in HEVs. These results suggest that HEV ECs and FRCs with perivascular DCs delicately regulate T and B cell entry into lymph nodes.

immunology↗

bioRxiv2020

Cell profiling of mouse acute kidney injury reveals conserved cellular responses to injury

After acute kidney injury (AKI), patients either recover or alternatively develop fibrosis and chronic kidney disease. Interactions between injured epithelia, stroma and inflammatory cells determine whether kidneys repair or undergo fibrosis, but the molecular events that drive these processes are poorly understood. Here, we use single nucleus RNA sequencing of a mouse model of AKI to characterize cell states during repair from acute injury. We identify a distinct proinflammatory and profibrotic proximal tubule cell state that fails to repair. Deconvolution of bulk RNA-seq datasets indicates that this "failed-repair proximal tubule cell" or FR-PTC, state can be detected in other models of kidney injury, increasing in the aging rat kidney and over time in human kidney allografts. We also describe dynamic intercellular communication networks and discern transcriptional pathways driving successful vs. failed repair. Our study provides a detailed description of cellular responses after injury and suggests that the FR-PTC state may represent a therapeutic target to improve repair. Significance StatementSingle nucleus RNA sequencing revealed gene expression changes during repair after acute kidney injury. We describe a small population of proximal tubule cells that fail to repair (FR-PTC). Since this subpopulation expresses abundant pro-inflammatory and profibrotic genes, it may represent a new therapeutic target to improve repair and reduce fibrosis after AKI.

genomics↗

bioRxiv2019

The Single Cell Transcriptomic Landscape of Early Human Diabetic Nephropathy

Diabetic nephropathy is characterized by damage to both the glomerulus and tubulointerstitium, but relatively little is known about accompanying cell-specific changes in gene expression. We performed unbiased single nucleus RNA sequencing (snRNAseq) on cryopreserved human diabetic kidney samples to generate 23,980 single nucleus transcriptomes from three control and three early diabetic nephropathy samples. All major cell types of the kidney were represented in the final dataset. Side by side comparison demonstrated cell-type-specific changes in gene expression that are important for ion transport, angiogenesis, and immune cell activation. In particular, we show that the diabetic loop of Henle, late distal convoluted tubule, and principal cells all adopt a gene expression signature consistent with increased potassium secretion, including alterations in Na-K+-ATPase, WNK1, mineralocorticoid receptor and NEDD4L expression, as well as decreased paracellular calcium and magnesium reabsorption. We also identify strong angiogenic signatures in glomerular cell types, proximal convoluted tubule, distal convoluted tubule and principal cells. Taken together, these results suggest that increased potassium secretion and angiogenic signaling represent early kidney responses in human diabetic nephropathy.\n\nSignificance StatementSingle nucleus RNA sequencing revealed gene expression changes in early diabetic nephropathy that promote urinary potassium secretion and decreased calcium and magnesium reabsorption. Multiple cell types exhibited angiogenic signatures, which may represent early signs of aberrant angiogenesis. These alterations may help to identify biomarkers for disease progression or signaling pathways amenable to early intervention.\n\nO_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

genomics↗

bioRxiv2017

Comparative analysis of kidney organoid and adult human kidney single cell and single nucleus transcriptomes

Kidney organoids differentiated from human pluripotent stem cells hold great promise for understanding organogenesis, modeling disease and ultimately as a source of replacement tissue. Realizing the full potential of this technology will require better differentiation strategies based upon knowledge of the cellular diversity and differentiation state of all cells within these organoids. Here we analyze single cell gene expression in 45,227 cells isolated from 23 organoids differentiated using two different protocols. Both generate kidney organoids that contain a diverse range of kidney cells at differing ratios as well as non-renal cell types. We quantified the differentiation state of major organoid kidney cell types by comparing them against a 4,259 single nucleus RNA-seq dataset generated from adult human kidney, revealing immaturity of all kidney organoid cell types. We reconstructed lineage relationships during organoid differentiation through pseudotemporal ordering, and identified transcription factor networks associated with fate decisions. These results define impressive kidney organoid cell diversity, identify incomplete differentiation as a major roadblock for current directed differentiation protocols and provide a human adult kidney snRNA-seq dataset against which to benchmark future progress.

developmental biology↗