Tissue expression of PD-L1 mediates peripheral T cell tolerance

Programmed death 1 (PD-1), an inhibitory receptor expressed on activated lymphocytes, regulates tolerance and autoimmunity. PD-1 has two ligands: PD-1 ligand 1 (PD-L1), which is expressed broadly on hematopoietic and parenchymal cells, including pancreatic islet cells; and PD-L2, which is restricted to macrophages and dendritic cells. To investigate whether PD-L1 and PD-L2 have synergistic or unique roles in regulating T cell activation and tolerance, we generated mice lacking PD-L1 and PD-L2 (PD-L1/PD-L2−/− mice) and compared them to mice lacking either PD-L. PD-L1 and PD-L2 have overlapping functions in inhibiting interleukin-2 and interferon-γ production during T cell activation. However, PD-L1 has a unique and critical role in controlling self-reactive T cells in the pancreas. Our studies with bone marrow chimeras demonstrate that PD-L1/PD-L2 expression only on antigen-presenting cells is insufficient to prevent the early onset diabetes that develops in PD-L1/PD-L2−/− non-obese diabetic mice. PD-L1 expression in islets protects against immunopathology after transplantation of syngeneic islets into diabetic recipients. PD-L1 inhibits pathogenic self-reactive CD4+ T cell–mediated tissue destruction and effector cytokine production. These data provide evidence that PD-L1 expression on parenchymal cells rather than hematopoietic cells protects against autoimmune diabetes and point to a novel role for PD-1–PD-L1 interactions in mediating tissue tolerance

The Role of TNF-α in Mice with Type 1- and 2- Diabetes

Background: Previously, we have demonstrated that short-term treatment of new onset diabetic Non-obese diabetic (NOD) mice, mice that are afflicted with both type 1 (T1D) and type 2 (T2D) diabetes with either Power Mix (PM) regimen or alpha1 antitrypsin (AAT) permanently restores euglycemia, immune tolerance to self-islets and normal insulin signaling. Methodology and Principal Findings: To search for relevant therapeutic targets, we have applied genome wide transcriptional profiling and systems biology oriented bioinformatics analysis to examine the impact of the PM and AAT regimens upon pancreatic lymph node (PLN) and fat, a crucial tissue for insulin dependent glucose disposal, in new onset diabetic non-obese diabetic (NOD) mice. Systems biology analysis identified tumor necrosis factor alpha (TNF-$\alpha$) as the top focus gene hub, as determined by the highest degree of connectivity, in both tissues. In PLNs and fat, TNF-$\alpha$ interacted with 53% and 32% of genes, respectively, associated with reversal of diabetes by previous treatments and was thereby selected as a therapeutic target. Short-term anti-TNF-$\alpha$ treatment ablated a T cell-rich islet-invasive and beta cell-destructive process, thereby enhancing beta cell viability. Indeed anti-TNF-$\alpha$ treatment induces immune tolerance selective to syngeneic beta cells. In addition to these curative effects on T1D anti-TNF-e33254 treatment restored in vivo insulin signaling resulting in restoration of insulin sensitivity. Conclusions: In short, our molecular analysis suggested that PM and AAT both may act in part by quenching a detrimental TNF-$\alpha$ dependent effect in both fat and PLNs. Indeed, short-term anti-TNF-$\alpha$ mAb treatment restored enduring euglycemia, self-tolerance, and normal insulin signaling

Dysregulated Nephrin in Diabetic Nephropathy of Type 2 Diabetes: A Cross Sectional Study

Podocyte specific proteins are dysregulated in diabetic nephropathy, though the extent of their expression loss is not identical and may be subject to different regulatory factors. Quantifying the degree of loss may help identify the most useful protein to use as an early biomarker of diabetic nephropathy.Protein expression of synaptopodin, podocin and nephrin were quantified in 15 Type 2 diabetic renal biopsies and 12 control patients. We found statistically significant downregulation of synaptopodin (P<0.0001), podocin (P = 0.0002), and nephrin (P<0.0001) in kidney biopsies of diabetic nephropathy as compared with controls. Urinary nephrin levels (nephrinuria) were then measured in 66 patients with Type 2 diabetes and 10 healthy controls by an enzyme-linked immunosorbent assay (Exocell, Philadelphia, PA). When divided into groups according to normo-, micro-, and macroalbuminuria, nephrinuria was found to be present in 100% of diabetic patients with micro- and macroalbuminuria, as well as 54% of patients with normoalbuminuria. Nephrinuria also correlated significantly with albuminuria (rho = 0.89, p<0.001), systolic blood pressure (rho = 0.32, p = 0.007), and correlated negatively with serum albumin (rho = -0.48, p<0.0001) and eGFR (rho = -0.33, p = 0.005).These data suggest that key podocyte-specific protein expressions are significantly and differentially downregulated in diabetic nephropathy. The finding that nephrinuria is observed in a majority of these normoalbuminuric patients demonstrates that it may precede microalbuminuria. If further research confirms nephrinuria to be a biomarker of pre-clinical diabetic nephropathy, it would shed light on podocyte metabolism in disease, and raise the possibility of new and earlier therapeutic targets

Podocyturia as a Diagnostic Marker for Preeclampsia amongst High-Risk Pregnant Patients

Urinary podocyte (podocyturia) has been studied as a diagnostic marker for preeclampsia. We sought to validate its use in preeclampsia and in differentiating it from other high risk pregnancy states. We studied an obstetric population at high risk to develop preeclampsia (study group) and uncomplicated pregnancies (control group) by analyzing their urine sediment for podocytes within 24 hours of delivery. Podocytes were identified by immunohistochemistry using the podocyte-specific protein synaptopodin. Of the 56 patients who were enrolled, 29 patients were diagnosed with preeclampsia, 9 patients had hypertensive conditions such as chronic and gestational hypertension, 6 patients had Type I/II and gestational diabetes mellitus, 3 patients were classified as others, and 9 patients exhibited uncomplicated pregnancies. Podocyturia was identified in 11 out of 29 (38%) of patients with preeclampsia/eclampsia, 3 out of 9 (33%) with gestational and chronic hypertension, and 3 out of 6 (50%) with Type I/II and gestational diabetes mellitus. None of the 9 patients (0%) with uncomplicated pregnancies demonstrated podocyturia. The sensitivity and specificity of podocyturia for preeclampsia were found to be 38% and 70%. Our study showed that podocyturia does not appear to be a sensitive nor a specific marker to diagnose preeclampsia

A comparison of podocyturia, albuminuria and nephrinuria in predicting the development of preeclampsia: a prospective study.

Preeclampsia, a hypertensive multisystem disease that complicates 5-8% of all pregnancy, is a major cause for maternal and fetal mortality and morbidity. The disease is associated with increased spontaneous and evoked preterm birth and remote cardio-renal disorders in the mother and offspring. Thus the ability to predict the disease should lead to earlier care and decreased morbidity. This has led to fervent attempts to identify early predictive biomarkers and research endeavors that have expanded as we learn more regarding possible causes of the disease. As preeclampsia is associated with specific renal pathology including podocyte injury, early urinary podocyte (podocyturia), or the podocyte specific proteinuria nephrin in the urine (nephrinuria), as well as the more easily measured urinary albumin (albuminuria), have all been suggested as predictive markers. We performed a prospective study recruiting 91 pregnant women (78 of whom were high risk) and studied the predictive ability of these three urinary biomarkers. The subjects were recruited between 15-38 weeks of gestation. Fourteen patients, all in the high-risk obstetric group, developed preeclampsia. The levels of podocyturia, nephrinuria, and albuminuria were variably higher in the high-risk pregnant patients who developed preeclampsia. The sensitivities and specificities for podocyturia were 70% and 43%, for albuminuria were 36% and 96%, and for nephrinuria were 57% and 58%, respectively. Also, abnormal nephrinuria (69%) and podocyturia (38%) were detected in low risk women who had uncomplicated gestations; none of these women exhibited albuminuria. In our study, none of the three urinary markers achieved the minimum predictive values required for clinical testing. The lack of excessive albuminuria, however, may indicate a preeclampsia-free gestation. Given a discrepant literature, further studies with larger sample size should be considered

Dot plots of albumin-to-creatinine ratios.

<p>A) Urine albumin-to-creatinine ratio measurements in 2^nd and 3^rd trimester recruited patients who developed PE vs. patients who did not develop PE B) Urine albumin-to-creatinine ratio measurements in 2^rd trimester recruited patients who developed PE vs. patients who did not develop PE C) Urine albumin-to-creatinine ratio measurements in 3^rd trimester recruited patients who developed PE vs. patients who did not develop PE.</p

Dot plots of urine nephrin-to-creatinine ratios.

<p>A) Urine nephrin-to-creatinine ratio measurements in 2^nd and 3^rd trimester recruited patients who developed PE vs. patients who did not develop PE B) Urine nephrin-to-creatinine ratio measurements in 2^rd trimester recruited patients who developed PE vs. patients who did not develop PE C) Urine nephrin-to-creatinine ratio measurements in 3^rd trimester recruited patients who developed PE vs. patients who did not develop PE.</p

Patient Characteristics According to Preeclampsia Outcome.

<p>Data median (interquartile range) or <b>(#/%)</b>.</p>1<p>SBP: systolic blood pressure.</p>2<p>DBP: diastolic blood pressure.</p>3<p>BUN: blood urea nitrogen.</p

Dot plots of urine podocyte-to-creatinine ratios.

<p>A) Urine podocyte-to-creatinine ratio measurements in 2^nd and 3^rd trimester recruited patients who developed PE vs. patients who did not develop PE B) Urine podocyte-to-creatinine ratio measurements in 2^rd trimester recruited patients who developed PE vs. patients who did not develop PE C) Urine podocyte-to-creatinine ratio measurements in 3^rd trimester recruited patients who developed PE vs. patients who did not develop PE.</p