a-retrograde-implantation-approach-for-peritoneal-dialysis-catheter

A Retrograde Implantation Approach for Peritoneal Dialysis Catheter Placement in Mice

Murine models are employed to probe various aspects of peritoneal dialysis (PD), such as peritoneal inflammation and fibrosis. These events drive ...

Casitas B lymphoma (c-Cbl) is an E3 ubiquitin ligase and a negative regulator of colorectal cancer (CRC). Despite its high expression in immune cells, the effect of c-Cbl on the tumor microenvironment remains poorly understood. Here we demonstrate that c-Cbl alters the tumor microenvironment and suppresses Programmed cell death-1 (PD-1) protein, an immune checkpoint receptor. Using syngeneic CRC xenografts, we observed significantly higher growth of xenografts and infiltrating immune cells in c-Cbl compared to c-Cbl mice. Tumor-associated CD8+ T-lymphocytes and macrophages of c-Cbl mice showed 2-3-fold higher levels of PD-1. Functionally, macrophages from c-Cbl mice showed a 4-5-fold reduction in tumor phagocytosis, which was restored with an anti-PD-1 neutralizing antibody suggesting regulation of PD-1 by c-Cbl. Further mechanistic probing revealed that C-terminus of c-Cbl interacted with the cytoplasmic tail of PD-1. c-Cbl destabilized PD-1 through ubiquitination- proteasomal degradation depending on c-Cbl's RING finger function. This data demonstrates c-Cbl as an E3 ligase of PD-1 and a regulator of tumor microenvironment, both of which were unrecognized components of its tumor suppressive activity. Advancing immune checkpoint and c-Cbl biology, our study prompts for probing of PD-1 regulation by c-Cbl in conditions driven by immune checkpoint abnormalities such as cancers and autoimmune diseases.

c-Cbl targets PD-1 in immune cells for proteasomal degradation and modulates colorectal tumor growth.

Kidney injury molecule-1 (KIM-1) staining has been shown to be very useful in identifying acute proximal tubular injury, but its sensitivity, specificity and predicting values for the recovery of renal function after injury in renal biopsies have not been well established. In the first study, we randomly selected 184 renal biopsies from a wide age range of patients (children to elderly) with various renal diseases. KIM-1 staining scores were significantly correlated with serum creatinine (sCr) levels (P < 0.05) in all age groups. Receiver-operating characteristic curve (ROC) was generated to evaluate true-positive rate (sensitivity) and true-negative rate (1-specificity). The area under the curve (AUC) in pediatric cases was 0.74, which demonstrated KIM-1 was a fair index in correlating with sCr. In adults, the AUC was 0.87, indicating that KIM-1 was an even better index in the adult population in correlating to sCr. The second study was to determine whether KIM-1 could be a potential predictor of the recovery of acute kidney injury (AKI), and 51 indicated native biopsies with acute tubular injury were randomly selected for KIM-1 staining and sCr follow-up over a 6-month period. A higher KIM-1/sCr ratio (0.57 ± 0.06) was significantly and positively associated with a better reduction in sCr over 6 months. In summary, our data demonstrated that KIM-1 staining in renal biopsies is a sensitive and specific marker to identify acute tubular injury and KIM-1/sCr ratio is useful for predicting the recovery of renal function after injury, although some patients' sCr levels cannot return to their baseline levels.

Kidney injury molecule-1, a sensitive and specific marker for identifying acute proximal tubular injury, can be used to predict renal functional recovery in native renal biopsies.

Patients with malignancy are at 4- to 7-fold higher risk of venous thromboembolism (VTE), a potentially fatal, yet preventable complication. Although general mechanisms of thrombosis are enhanced in these patients, malignancy-specific triggers and their therapeutic implication remain poorly understood. Here we examined a colon cancer-specific VTE model and probed a set of metabolites with prothrombotic propensity in the inferior vena cava (IVC) ligation model. Athymic mice injected with human colon adenocarcinoma cells exhibited significantly higher IVC clot weights, a biological readout of venous thrombogenicity, compared with the control mice. Targeted metabolomics analysis of plasma of mice revealed an increase in the blood levels of kynurenine and indoxyl sulfate (tryptophan metabolites) in xenograft-bearing mice, which correlated positively with the increase in the IVC clot size. These metabolites are ligands of aryl hydrocarbon receptor (AHR) signaling. Accordingly, plasma from the xenograft-bearing mice activated the AHR pathway and augmented tissue factor (TF) and plasminogen activator inhibitor 1 (PAI-1) levels in venous endothelial cells in an AHR-dependent manner. Consistent with these findings, the endothelium from the IVC of xenograft-bearing animals revealed nuclear AHR and upregulated TF and PAI-1 expression, telltale signs of an activated AHR-TF/PAI-1 axis. Importantly, pharmacological inhibition of AHR activity suppressed TF and PAI-1 expression in endothelial cells of the IVC and reduced clot weights in both kynurenine-injected and xenograft-bearing mice. Together, these data show dysregulated tryptophan metabolites in a mouse cancer model, and they reveal a novel link between these metabolites and the control of the AHR-TF/PAI-1 axis and VTE in cancer.

Metabolites in a mouse cancer model enhance venous thrombogenicity through the aryl hydrocarbon receptor-tissue factor axis.

As the COVID-19 pandemic continues to spread, investigating the processes underlying the interactions between SARS-CoV-2 and its hosts is of high importance. Here, we report the identification of CD209L/L-SIGN and the related protein CD209/DC-SIGN as receptors capable of mediating SARS-CoV-2 entry into human cells. Immunofluorescence staining of human tissues revealed prominent expression of CD209L in the lung and kidney epithelium and endothelium. Multiple biochemical assays using a purified recombinant SARS-CoV-2 spike receptor binding domain (S-RBD) or S1 encompassing both NTB and RBD and ectopically expressed CD209L and CD209 revealed that CD209L and CD209 interact with S-RBD. CD209L contains two glycosylation sequons, at sites N92 and N361, but we determined that only site N92 is occupied. Removal of the -glycosylation at this site enhances the binding of S-RBD with CD209L. CD209L also interacts with ACE2, suggesting a role for heterodimerization of CD209L and ACE2 in SARS-CoV-2 entry and infection in cell types where both are present. Furthermore, we demonstrate that human endothelial cells are permissive to SARS-CoV-2 infection and interference with CD209L activity by knockdown strategy or with soluble CD209L inhibits virus entry. Our observations demonstrate that CD209L and CD209 serve as alternative receptors for SARS-CoV-2 in disease-relevant cell types, including the vascular system. This property is particularly important in tissues where ACE2 has low expression or is absent, and may have implications for antiviral drug development.

CD209L/L-SIGN and CD209/DC-SIGN act as receptors for SARS-CoV-2.

COVID-19 infection has protean systemic manifestations. Experience from previous coronavirus outbreaks, including the current SARS-CoV-2, has shown an augmented risk of thrombosis of both macrovasculature and microvasculature. The former involves both arterial and venous beds manifesting as stroke, acute coronary syndrome and venous thromboembolic events. The microvascular thrombosis is an underappreciated complication of SARS-CoV-2 infection with profound implications on the development of multisystem organ failure. The telltale signs of perpetual on-going coagulation and fibrinolytic cascades underscore the presence of diffuse endothelial damage in the patients with COVID-19. These parameters serve as strong predictors of mortality. While summarizing the alterations of various components of thrombosis in patients with COVID-19, this review points to the emerging evidence that implicates the prominent role of the extrinsic coagulation cascade in COVID-19-related coagulopathy. These mechanisms are triggered by widespread endothelial cell damage (endotheliopathy), the dominant driver of macro- and micro-vascular thrombosis in these patients. We also summarize other mediators of thrombosis, clinically relevant nuances such as the occurrence of thromboembolic events despite thromboprophylaxis (breakthrough thrombosis), current understanding of systemic anticoagulation therapy and its risk-benefit ratio. We conclude by emphasizing a need to probe COVID-19-specific mechanisms of thrombosis to develop better risk markers and safer therapeutic targets.

Hyperthrombotic Milieu in COVID-19 Patients.

Conventional light sheet fluorescence microscopy (LSFM), or selective plane illumination microscopy (SPIM), enables high-resolution 3D imaging over a large volume by using two orthogonally aligned objective lenses to decouple excitation and emission. The recent development of oblique plane microscopy (OPM) simplifies LSFM design with only one single objective lens, by using off-axis excitation and remote focusing. However, most reports on OPM have a limited microscopic field of view (FOV), typically within 1×1 mm. Our goal is to overcome the limitation with a new variant of OPM to achieve a mesoscopic FOV.

Wide field-of-view volumetric imaging by a mesoscopic scanning oblique plane microscopy with switchable objective lenses.

As the COVID-19 pandemic continues to spread, investigating the processes underlying the interactions between SARS-CoV-2 and its hosts is of high importance. Here, we report the identification of CD209L/L-SIGN and the related protein CD209/DC-SIGN as receptors capable of mediating SARS-CoV-2 entry into human cells. Immunofluorescence staining of human tissues revealed prominent expression of CD209L in the lung and kidney epithelia and endothelia. Multiple biochemical assays using a purified recombinant SARS-CoV-2 spike receptor-binding domain (S-RBD) or S1 encompassing both N termal domain and RBD and ectopically expressed CD209L and CD209 revealed that CD209L and CD209 interact with S-RBD. CD209L contains two -glycosylation sequons, at sites N92 and N361, but we determined that only site N92 is occupied. Removal of the -glycosylation at this site enhances the binding of S-RBD with CD209L. CD209L also interacts with ACE2, suggesting a role for heterodimerization of CD209L and ACE2 in SARS-CoV-2 entry and infection in cell types where both are present. Furthermore, we demonstrate that human endothelial cells are permissive to SARS-CoV-2 infection, and interference with CD209L activity by a knockdown strategy or with soluble CD209L inhibits virus entry. Our observations demonstrate that CD209L and CD209 serve as alternative receptors for SARS-CoV-2 in disease-relevant cell types, including the vascular system. This property is particularly important in tissues where ACE2 has low expression or is absent and may have implications for antiviral drug development.

CD209L/L-SIGN and CD209/DC-SIGN Act as Receptors for SARS-CoV-2.

Chronic kidney disease (CKD) imposes a strong and independent risk for peripheral artery disease (PAD). While solutes retained in CKD patients (uremic solutes) inflict vascular damage, their role in PAD remains elusive. Here, we show that the dietary tryptophan-derived uremic solutes including indoxyl sulfate (IS) and kynurenine (Kyn) at concentrations corresponding to those in CKD patients suppress β-catenin in several cell types, including microvascular endothelial cells (ECs), inhibiting Wnt activity and proangiogenic Wnt targets in ECs. Mechanistic probing revealed that these uremic solutes downregulated β-catenin in a manner dependent on serine 33 in its degron motif and through the aryl hydrocarbon receptor (AHR). Hindlimb ischemia in adenine-induced CKD and IS solute-specific mouse models showed diminished β-catenin and VEGF-A in the capillaries and reduced capillary density, which correlated inversely with blood levels of IS and Kyn and AHR activity in ECs. An AHR inhibitor treatment normalized postischemic angiogenic response in CKD mice to a non-CKD level. In a prospective cohort of PAD patients, plasma levels of tryptophan metabolites and plasma's AHR-inducing activity in ECs significantly increased the risk of future adverse limb events. This work uncovers the tryptophan metabolite/AHR/β-catenin axis as a mediator of microvascular rarefaction in CKD patients and demonstrates its targetability for PAD in CKD models.

Tryptophan metabolites suppress the Wnt pathway and promote adverse limb events in chronic kidney disease.

Endothelial cell injury is a common nidus of renal injury in patients and consistent with the high prevalence of AKI reported during the coronavirus disease 2019 pandemic. This cell type expresses integrin 5 (ITGA5), which is essential to the Tie2 signaling pathway. The microRNA miR-218-5p is upregulated in endothelial progenitor cells (EPCs) after hypoxia, but microRNA regulation of Tie2 in the EPC lineage is unclear.

Integrin 5 Is Regulated by miR-218-5p in Endothelial Progenitor Cells.

Aberrant hyperactivation of Wnt signaling, driven by nuclear β-catenin in the colonic epithelium, represents the seminal event in the initiation and progression of colorectal cancer (CRC). Despite its established role in CRC tumorigenesis, clinical translation of Wnt inhibitors remains unsuccessful. Late SV40 factor (LSF; encoded by TFCP2) is a transcription factor and a potent oncogene. We identified a chemotype, named factor quinolinone inhibitors (FQIs), that specifically inhibits LSF DNA-binding, partner protein-binding, and transactivation activities. We examined the role of LSF and FQIs in CRC tumor growth. Herein, we show that LSF and β-catenin interacted in several CRC cell lines irrespective of their mutational profile, which was disrupted by FQI2-34. FQI2-34 suppressed Wnt activity in CRC cells in a dose-dependent manner. Leveraging both allogeneic and syngeneic xenograft models, we show that FQI2-34 suppressed CRC tumor growth, significantly reduced nuclear β-catenin, and down-regulated Wnt targets, such as AXIN-2 and SRY-box transcription factor 9, in the xenograft cells. FQI2-34 suppressed the proliferation of xenograft cells. Adenocarcinomas from a series of stage IV CRC patients revealed a positive correlation between LSF expression and Wnt targets (AXIN-2 and SRY-box transcription factor 9) within the CRC cells. Collectively, this study uncovers the Wnt inhibitory and CRC growth-suppressive effects of these LSF inhibitors in CRC cells, revealing a novel target in CRC therapeutics.

Article	Year
c-Cbl targets PD-1 in immune cells for proteasomal degradation and modulates colorectal tumor growth. Scientific reports\| PubMed ID: 31882749	2019
Kidney injury molecule-1, a sensitive and specific marker for identifying acute proximal tubular injury, can be used to predict renal functional recovery in native renal biopsies. International urology and nephrology\| PubMed ID: 31646460	2019
Metabolites in a mouse cancer model enhance venous thrombogenicity through the aryl hydrocarbon receptor-tissue factor axis. Blood\| PubMed ID: 31877217	2019
CD209L/L-SIGN and CD209/DC-SIGN act as receptors for SARS-CoV-2. bioRxiv : the preprint server for biology\| PubMed ID: 32607506	2021
Hyperthrombotic Milieu in COVID-19 Patients. Cells\| PubMed ID: 33142844	2020
Wide field-of-view volumetric imaging by a mesoscopic scanning oblique plane microscopy with switchable objective lenses. Quantitative imaging in medicine and surgery\| PubMed ID: 33654671	2021
CD209L/L-SIGN and CD209/DC-SIGN Act as Receptors for SARS-CoV-2. ACS central science\| PubMed ID: 34341769	2021
Tryptophan metabolites suppress the Wnt pathway and promote adverse limb events in chronic kidney disease. The Journal of clinical investigation\| PubMed ID: 34752422	2022
Integrin 5 Is Regulated by miR-218-5p in Endothelial Progenitor Cells. Journal of the American Society of Nephrology : JASN\| PubMed ID: 35091451	2022
Pharmacologic Manipulation of Late SV40 Factor Suppresses Wnt Signaling and Inhibits Growth of Allogeneic and Syngeneic Colon Cancer Xenografts. The American journal of pathology\| PubMed ID: 35710032	2022

Wenqing Yin

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