Outcome and safety of a surveillance biopsy guided personalized immunosuppression program after liver transplantation.

Graft survival beyond year 1 has not changed after orthotopic liver transplantation (OLT) over the last decades. Likewise, OLT causes comorbidities such as infection, renal impairment and cancer. We evaluated our single-center real-world individualized immunosuppression program after OLT, based on 211 baseline surveillance biopsies (svLbx) without any procedural complications. Patients were classified as low, intermediate and high rejection risk based on graft injury in svLbx and anti-HLA donor-specific antibodies. While 32% of patients had minimal histological inflammation, 57% showed histological inflammation and 23% advanced fibrosis (>F2), which was not predicted by lab parameters. IS was modified in 79% of patients after svLbx. After immunosuppression reduction in 69 patients, only 5 patients showed ALT elevations and three of these patients had a biopsy-proven acute rejection, two of them related to lethal comorbidities. The rate of liver enzyme elevation including rejection was not significantly increased compared to a svLbx control cohort prior to the initiation of our structured program. Immunosuppression reduction led to significantly better kidney function compared to this control cohort. In conclusion, a biopsy guided personalized immunosuppression protocol after OLT can identify patients requiring lower immunosuppression or patients with graft injury in which IS should not be further reduced.

Elevated fractional donor-derived cell-free DNA during subclinical graft injury after liver transplantation.

Personalized immunosuppression (IS) promises to improve the balance of necessary control of alloreactivity and dose-dependent adverse effects of long-term IS such as kidney insufficiency, infections, and malignancies. The majority of liver transplantation (LT) recipients exhibit graft injuries (graft inflammation and/or fibrosis) that are not eligible for an IS reduction according to current Banff criteria, even when liver enzymes are normal or only marginally elevated. This cross-sectional study evaluated the noninvasive prediction of such subclinical graft injuries in surveillance liver biopsies via donor-derived cell-free DNA (dd-cfDNA). Absolute and fractional dd-cfDNA increased stepwise from patients without histological signs of rejection (n = 26) over subclinical graft injury (n = 61), including subclinical T cell-mediated rejection to clinical overt T cell-mediated rejection (n = 21). Thus, fractional plasma dd-cfDNA was significantly elevated paired to surveillance biopsies with relevant subclinical graft injury according to 2016 Banff criteria compared with those with minimal or absent histological graft injury. In contrast, the presence of donor-specific anti-human leukocyte antigen antibodies was not associated with the amount of dd-cfDNA. The sensitivity and specificity of fractional dd-cfDNA to noninvasively predict relevant subclinical graft injury was rather limited with 73% and 52% at the cutoff value of 2.1% fractional dd-cfDNA. The positive predictive value of fractional dd-cfDNA above 2.1% was 76% to noninvasively predict subclinical graft injury, calculated on the prevalence of graft injury in our prospective surveillance biopsy program, whereas the negative predictive values was not predictive (47%). In conclusion, dd-cfDNA has a rather limited diagnostic fidelity in addition to other noninvasive markers for the assessment of subclinical graft injury in personalized IS approaches after LT in a cross-sectional setting.

KIF12 Variants and Disturbed Hepatocyte Polarity in Children with a Phenotypic Spectrum of Cholestatic Liver Disease.

KIF12 has been identified as a cholestasis-associated candidate gene. We describe 6 cases from 4 unrelated families with diverse cholestatic phenotypes carrying 2 different homozygous KIF12 truncating variants. Immunofluorescence investigations of paraffin-embedded liver sections suggest that KIF12-associated impaired functional cell polarity may be the underlying cause.

Anthracyclines induce DNA damage response-mediated protection against severe sepsis.

Severe sepsis remains a poorly understood systemic inflammatory condition with high mortality rates and limited therapeutic options in addition to organ support measures. Here we show that the clinically approved group of anthracyclines acts therapeutically at a low dose regimen to confer robust protection against severe sepsis in mice. This salutary effect is strictly dependent on the activation of DNA damage response and autophagy pathways in the lung, as demonstrated by deletion of the ataxia telangiectasia mutated (Atm) or the autophagy-related protein 7 (Atg7) specifically in this organ. The protective effect of anthracyclines occurs irrespectively of pathogen burden, conferring disease tolerance to severe sepsis. These findings demonstrate that DNA damage responses, including the ATM and Fanconi Anemia pathways, are important modulators of immune responses and might be exploited to confer protection to inflammation-driven conditions, including severe sepsis.

CK-18 cell death markers improve the prediction of histological remission in autoimmune hepatitis during biochemical remission.

Incomplete histological remission of autoimmune hepatitis (AIH) is associated with a reduced long-term survival and an increased relapse rate even during biochemical remission (BR). The aim of this international multicentre study was to explore the diagnostic fidelity of cytokeratin-18 cell death markers to noninvasively detect incomplete histological remission. Thereby, cytokeratin-18 cell death marker M65 but not ALT and immunoglobulins was significantly higher in patients with incomplete histological remission (mHAI ≥ 4) compared to those with mHAI ≤ 3. M65 levels > 305 U/L, identified in the training cohort, facilitated the noninvasive detection of incomplete histological remission with a sensitivity of 75% and negative predictive value of 86% in the validation cohort. While BR with M65 < 305 U/L suggested complete histological remission (86%), BR with M65 > 305 U/L reduced the rate of histological remission to 60%. In conclusion, M65 may help to better select patients for or to reduce surveillance liver biopsies in the future.

Dubin-Johnson Syndrome as Differential Diagnosis for Neonatal Cholestasis.

OBJECTIVES: Dubin-Johnson syndrome (DJS) is an autosomal recessive disorder in which multidrug-resistance-associated protein 2 (MRP2) deficiency causes an excretion disorder of conjugated bilirubin from hepatocytes into bile canaliculi. Its clinical presentation as neonatal cholestasis (NC) is rare but represents an important differential diagnosis. We aimed to define DJS-specific characteristics in NC, in particular in contrast to biliary atresia (BA) patients, and to highlight diagnostic tools that can help to avoid invasive diagnostic tests. METHODS: We performed a review of case records from 2006 to 2020 and compared 4 DJS patients to 26 patients with proven BA consecutively diagnosed from 2014 to 2017. DJS was diagnosed by urine coproporphyrin analysis (UCA) and by genetic analysis (GA) for disease-associated ABCC2 variants. RESULTS: Four male patients with NC were diagnosed with DJS by UCA and GA. DJS patients presenting as NC showed significantly lower values for aspartate aminotransferase (AST) (P < 0.001), for alanine aminotransferase (ALT) (P = 0.002) and for gamma-glutamyl transferase (GGT) (P < 0.001) compared with BA patients. Other examinations, however, could not clearly discriminate them (e.g.: stool colour, serum bile acids, total serum bilirubin). CONCLUSIONS: DJS is not only a rare differential diagnosis in NC with a suspicious phenotype (almost normal AST, ALT) but also shows overlapping features with BA. It should, therefore, be considered in every infant with NC and an atypical liver enzyme pattern to protect patients from unnecessary, invasive examinations. For this, UCA is a fast and reliable diagnostic tool. Confirmation based on GA is recommended. DJS patients have a good long-term prognosis.

[A supposed hepatocellular adenoma turns out to be intrahepatic splenosis - a case report]. / Ein vermeintliches hepatozelluläres Adenom stellt sich als intrahepatische Splenose heraus ­ ein Fallbericht.

We report the case of a 62-year-old Caucasian male patient who presented with epigastric pain to our outpatient clinic. On abdominal ultrasound we detected a 26 mm oval hypoechoic lesion in segment 2 of the left liver lobe. Performing contrast-enhanced ultrasound this lesion showed an arterial hypervascularization with centripetal filling and a spoke wheel pattern. Due to a hyperenhancement during the portal and late phase this lesion led to the diagnosis of a benign liver tumor, probably a hepatocellular adenoma (HCA). As focal nodular hyperplasia (FNH) was still another possible diagnosis, we decided to perform an MRI, which could not differentiate between HCA and hepatocellular carcinoma (HCC). Therefore, we performed liver biopsy of this lesion. Histology and immunohistochemistry led to the final diagnosis of intrahepatic splenosis. Reassessment of patient history revealed an abdominal trauma with splenic rupture 5 years ago. Intrahepatic splenosis should be considered as an important differential diagnosis in patients with unknown liver tumor and a history of splenic trauma.

Generation of focal mutations and large genomic deletions in the pancreas using inducible in vivo genome editing.

Beyond the nearly uniform presence of KRAS mutations, pancreatic cancer is increasingly recognized as a heterogeneous disease. Preclinical in vivo model systems exist, but with the advent of precision oncology, murine models with enhanced genetic flexibility are needed to functionally annotate genetic alterations found in the human malignancy. Here, we describe the generation of focal gene disruptions and large chromosomal deletions via inducible and pancreas-specific expression of Cas9 in adult mice. Experimental mice are derived on demand directly from genetically engineered embryonic stem cells, without the need for further intercrossing. To provide initial validation of our approach, we show that disruption of the E3 ubiquitin ligase Rnf43 accelerates KrasG12D-dependent tumourigenesis. Moreover, we demonstrate that this system can be used to rapidly interrogate the impact of complex cancer-associated alleles through the generation of a previously unstudied 1.2 megabase deletion surrounding the CDKN2A and CDKN2B tumour suppressors. Thus, our approach is capable of reproducibly generating biallelic and precise loss of large chromosomal fragments that, in conjunction with mutant Kras, leads to development of pancreatic ductal adenocarcinoma with full penetrance.

Cholemic Nephropathy Causes Acute Kidney Injury and Is Accompanied by Loss of Aquaporin 2 in Collecting Ducts.

Impairment of renal function often occurs in patients with liver disease. Hepatorenal syndrome is a significant cause of acute kidney injury (AKI) in patients with cirrhosis (HRS-AKI, type 1). Causes of non-HRS-AKI include cholemic nephropathy (CN), a disease that is characterized by intratubular bile casts and tubular injury. As data on patients with CN are obtained primarily from case reports or autopsy studies, we aimed to investigate the frequency and clinical course of CN. We identified 149 patients who underwent kidney biopsy between 2000 and 2016 at the Department of Gastroenterology, Hepatology and Endocrinology at Hannover Medical School. Of these, 79 had a history of liver disease and deterioration of renal function. When applying recent European Association for the Study of the Liver criteria, 45 of 79 patients (57%) presented with AKI, whereas 34 patients (43%) had chronic kidney disease (CKD). Renal biopsy revealed the diagnosis of CN in 8 of 45 patients with AKI (17.8%), whereas none of the patients with CKD was diagnosed with CN. Univariate analysis identified serum bilirubin, alkaline phosphatase, and urinary bilirubin and urobilinogen as predictive factors for the diagnosis of CN. Histological analysis of AKI patients with normal bilirubin, elevated bilirubin, and the diagnosis of CN revealed loss of aquaporin 2 (AQP2) expression in collecting ducts in patients with elevated bilirubin and CN. Biopsy-related complications requiring medical intervention occurred in 4 of 79 patients (5.1%). Conclusion: CN is a common finding in patients with liver disease, AKI, and highly elevated bilirubin. Loss of AQP2 in AKI patients with elevated bilirubin and CN might be the result of toxic effects of cholestasis and in part be responsible for the impairment of renal function.

The FERM protein EPB41L5 regulates actomyosin contractility and focal adhesion formation to maintain the kidney filtration barrier.

Podocytes form the outer part of the glomerular filter, where they have to withstand enormous transcapillary filtration forces driving glomerular filtration. Detachment of podocytes from the glomerular basement membrane precedes most glomerular diseases. However, little is known about the regulation of podocyte adhesion in vivo. Thus, we systematically screened for podocyte-specific focal adhesome (FA) components, using genetic reporter models in combination with iTRAQ-based mass spectrometry. This approach led to the identification of FERM domain protein EPB41L5 as a highly enriched podocyte-specific FA component in vivo. Genetic deletion of Epb41l5 resulted in severe proteinuria, detachment of podocytes, and development of focal segmental glomerulosclerosis. Remarkably, by binding and recruiting the RhoGEF ARGHEF18 to the leading edge, EPB41L5 directly controls actomyosin contractility and subsequent maturation of focal adhesions, cell spreading, and migration. Furthermore, EPB41L5 controls matrix-dependent outside-in signaling by regulating the focal adhesome composition. Thus, by linking extracellular matrix sensing and signaling, focal adhesion maturation, and actomyosin activation EPB41L5 ensures the mechanical stability required for podocytes at the kidney filtration barrier. Finally, a diminution of EPB41L5-dependent signaling programs appears to be a common theme of podocyte disease, and therefore offers unexpected interventional therapeutic strategies to prevent podocyte loss and kidney disease progression.

Potential Role for Urine Polymerase Chain Reaction in the Diagnosis of Whipple's Disease.

BACKGROUND: Whipple's disease (WD) is a rare infection with Tropheryma whipplei that is fatal if untreated. Diagnosis is challenging and currently based on invasive sampling. In a case of WD diagnosed from a kidney biopsy, we observed morphologically-intact bacteria within the glomerular capsular space and tubular lumens. This raised the questions of whether renal filtration of bacteria is common in WD and whether polymerase chain reaction (PCR) testing of urine might serve as a diagnostic test for WD. METHODS: We prospectively investigated urine samples of 12 newly-diagnosed and 31 treated WD patients by PCR. As controls, we investigated samples from 110 healthy volunteers and patients with excluded WD or acute gastroenteritis. RESULTS: Out of 12 urine samples from independent, therapy-naive WD patients, 9 were positive for T. whipplei PCR. In 3 patients, fluorescence in situ hybridization visualized T. whipplei in urine. All control samples were negative, including those of 11 healthy carriers with T. whipplei-positive stool samples. In our study, the detection of T. whipplei in the urine of untreated patients correlated in all cases with WD. CONCLUSIONS: T. whipplei is detectable by PCR in the urine of the majority of therapy-naive WD patients. With a low prevalence but far-reaching consequences upon diagnosis, invasive sampling for WD is mandatory and must be based on a strong suspicion. Urine testing could prevent patients from being undiagnosed for years. Urine may serve as a novel, easy-to-obtain specimen for guiding the initial diagnosis of WD, in particular in patients with extra-intestinal WD.

Assessment of liver ischemia reperfusion injury in mice using hepatic T2 mapping: Comparison with histopathology.

BACKGROUND: Liver ischemia reperfusion injury (IRI) occurs during liver surgery or transplantation resulting in an inflammatory response, tissue damage, and functional impairment of the organ. PURPOSE: To assess the feasibility of T2 mapping for noninvasive quantification of liver edema after partial liver IRI in mice. STUDY TYPE: Prospective, experimental study. ANIMAL MODEL: Partial liver IRI was induced in C57BL/6-mice by transient clamping of the left lateral and median liver lobes for 35 (n = 8), 45 (n = 6), 60 (n = 17), or 90 minutes (n = 5). For comparison, healthy C57BL/6-mice were examined as controls (n = 9). FIELD STRENGTH/SEQUENCE: Functional liver MRI was performed on a 7T scanner using a respiratory-triggered multiecho spin-echo sequence. ASSESSMENT: Healthy control mice and mice with partial liver IRI on day 1 after surgery, and additionally on day 7 in a subgroup with 60 minutes IRI (n = 8) were examined. Maps of T2 relaxation time of liver tissue were used to assess distribution, severity of tissue edema (mean T2 time), and the percentage of edematous liver tissue. STATISTICAL TEST: One-way analysis of variance (ANOVA) with Tukey's honest significant difference (HSD), paired t-tests, Pearson's test for correlation of MRI parameters with levels of liver enzymes, and histopathology, receiver operating characteristic (ROC) analysis. RESULTS: Significant tissue edema induced by liver IRI as compared to the control group was detected by increased mean T2 times in groups with 60 minutes (P < 0.001) and 90 minutes IRI (P < 0.001). The percentage of edematous liver tissue significantly increased with longer ischemia times (controls 3.4 ± 0.4%, 35 minutes 5.3 ± 0.6%, 45 minutes 23.3 ± 7.6%, 60 minutes 39.7 ± 3.6%, 90 minutes 51.3 ± 4.5%). Mean T2 times and the percentage of edematous liver tissue significantly correlated with elevation of liver enzymes (P < 0.001), histological evidence of liver injury (r = 0.80 and r = 0.82, P < 0.001), and neutrophil infiltration (r = 0.70 and r = 0.74, P < 0.001). In the subgroup with follow-up, the severity (P < 0.01) and extent of liver edema decreased significantly over time (P < 0.01). DATA CONCLUSION: T2 mapping allows quantification and follow-up of liver injury in mice. LEVEL OF EVIDENCE: 1 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2018;48:1586-1594.

Gd-EOB-DTPA-enhanced MRI for quantitative assessment of liver organ damage after partial hepatic ischaemia reperfusion injury: correlation with histology and serum biomarkers of liver cell injury.

OBJECTIVE: To evaluate Gd-EOB-DTPA-enhanced MRI for quantitative assessment of liver organ damage after hepatic ischaemia reperfusion injury (IRI) in mice. METHODS: Partial hepatic IRI was induced in C57Bl/6 mice (n = 31) for 35, 45, 60 and 90 min. Gd-EOB-DTPA-enhanced MRI was performed 1 day after surgery using a 3D-FLASH sequence. A subgroup of n = 9 animals with 60 min IRI underwent follow-up with MRI and histology 7 days after IRI. The total liver volume was determined by manual segmentation of the entire liver. The volume of functional, contrast-enhanced liver parenchyma was quantified by a region growing algorithm (visual threshold) and an automated segmentation (Otsu's method). The percentages of functional, contrast-enhanced and damaged non-enhanced parenchyma were calculated according to these volumes. MRI data was correlated with serum liver enzyme concentrations and histologically quantified organ damage using periodic acid-Schiff (PAS) staining. RESULTS: The percentage of functional (contrasted) liver parenchyma decreased significantly with increasing ischaemia times (control, 94.4 ± 3.3%; 35 min IRI, 89.3 ± 4.1%; 45 min IRI, 87.9 ± 3.3%; 60 min IRI, 68 ± 10.5%, p < 0.001 vs. control; 90 min IRI, 55.9 ± 11.5%, p < 0.001 vs. control). The percentage of non-contrasted liver parenchyma correlated with histologically quantified liver organ damage (r = 0.637, p < 0.01) and serum liver enzyme elevations (AST r = 0.577, p < 0.01; ALT r = 0.536, p < 0.05). Follow-up MRI visualized recovery of functional liver parenchyma (71.5 ± 8.7% vs. 84 ± 2.1%, p < 0.05), consistent with less histological organ damage on day 7. CONCLUSION: We demonstrated the feasibility of Gd-EOB-DTPA-enhanced MRI for non-invasive quantification of damaged liver parenchyma following IRI in mice. This novel methodology may refine the characterization of liver disease and could have application in future studies targeting liver organ damage. KEY POINTS: • Prolonged ischaemia times in partial liver IRI increase liver organ damage. • Gd-EOB-DTPA-enhanced MRI at hepatobiliary phase identifies damaged liver volume after hepatic IRI. • Damaged liver parenchyma quantified with MRI correlates with histological liver damage. • Hepatobiliary phase Gd-EOB-DTPA-enhanced MRI enables non-invasive assessment of recovery from liver injury.

Targeting mTOR Signaling Can Prevent the Progression of FSGS.

Mammalian target of rapamycin (mTOR) signaling is involved in a variety of kidney diseases. Clinical trials administering mTOR inhibitors to patients with FSGS, a prototypic podocyte disease, led to conflicting results, ranging from remission to deterioration of kidney function. Here, we combined complex genetic titration of mTOR complex 1 (mTORC1) levels in murine glomerular disease models, pharmacologic studies, and human studies to precisely delineate the role of mTOR in FSGS. mTORC1 target genes were significantly induced in microdissected glomeruli from both patients with FSGS and a murine FSGS model. Furthermore, a mouse model with constitutive mTORC1 activation closely recapitulated human FSGS. Notably, the complete knockout of mTORC1 by induced deletion of both Raptor alleles accelerated the progression of murine FSGS models. However, lowering mTORC1 signaling by deleting just one Raptor allele ameliorated the progression of glomerulosclerosis. Similarly, low-dose treatment with the mTORC1 inhibitor rapamycin efficiently diminished disease progression. Mechanistically, complete pharmacologic inhibition of mTOR in immortalized podocytes shifted the cellular energy metabolism toward reduced rates of oxidative phosphorylation and anaerobic glycolysis, which correlated with increased production of reactive oxygen species. Together, these data suggest that podocyte injury and loss is commonly followed by adaptive mTOR activation. Prolonged mTOR activation, however, results in a metabolic podocyte reprogramming leading to increased cellular stress and dedifferentiation, thus offering a treatment rationale for incomplete mTOR inhibition.

Unraveling the role of podocyte turnover in glomerular aging and injury.

Podocyte loss is a major determinant of progressive CKD. Although recent studies showed that a subset of parietal epithelial cells can serve as podocyte progenitors, the role of podocyte turnover and regeneration in repair, aging, and nephron loss remains unclear. Here, we combined genetic fate mapping with highly efficient podocyte isolation protocols to precisely quantify podocyte turnover and regeneration. We demonstrate that parietal epithelial cells can give rise to fully differentiated visceral epithelial cells indistinguishable from resident podocytes and that limited podocyte renewal occurs in a diphtheria toxin model of acute podocyte ablation. In contrast, the compensatory programs initiated in response to nephron loss evoke glomerular hypertrophy, but not de novo podocyte generation. In addition, no turnover of podocytes could be detected in aging mice under physiologic conditions. In the absence of podocyte replacement, characteristic features of aging mouse kidneys included progressive accumulation of oxidized proteins, deposits of protein aggregates, loss of podocytes, and glomerulosclerosis. In summary, quantitative investigation of podocyte regeneration in vivo provides novel insights into the mechanism and capacity of podocyte turnover and regeneration in mice. Our data reveal that podocyte generation is mainly confined to glomerular development and may occur after acute glomerular injury, but it fails to regenerate podocytes in aging kidneys or in response to nephron loss.

Hantavirus infection with severe proteinuria and podocyte foot-process effacement.

Nephropathia epidemica, a zoonosis caused by Hantavirus infection (most commonly subtype Puumala) is associated with flu-like symptoms and acute kidney failure. Kidney manifestations are characterized predominantly by tubulointerstitial nephritis, hemorrhage into medullary tissues, interstitial edema, and tubular cell necrosis. Kidney failure is accompanied by proteinuria, and in some cases, nephrotic-range proteinuria may occur. However, the cellular mechanisms of proteinuria remain to be elucidated. We describe a Hantavirus (Puumala) infection in a 27-year-old man with acute kidney failure and severe and rapidly reversible proteinuria. Light microscopy of a kidney biopsy specimen showed only minor changes of glomeruli. However, transmission electron microscopy revealed podocyte foot-process effacement. Immunofluorescence staining of the slit diaphragm protein podocin and the tight junction protein ZO-1 revealed a partial mislocalization of these proteins. Together, these findings highlight that Hantavirus infection may perturb podocyte integrity, resulting in glomerular proteinuria. These alterations of podocytes and consequently the glomerular filtration barrier may be transient and resolve within weeks.

aPKCλ/ι and aPKCζ contribute to podocyte differentiation and glomerular maturation.

Precise positioning of the highly complex interdigitating podocyte foot processes is critical to form the normal glomerular filtration barrier, but the molecular programs driving this process are unknown. The protein atypical protein kinase C (aPKC)--a component of the Par complex, which localizes to tight junctions and interacts with slit diaphragm proteins--may play a role. Here, we found that the combined deletion of the aPKCλ/ι and aPKCζ isoforms in podocytes associated with incorrectly positioned centrosomes and Golgi apparatus and mislocalized molecules of the slit diaphragm. Furthermore, aPKC-deficient podocytes failed to form the normal network of foot processes, leading to defective glomerular maturation with incomplete capillary formation and mesangiolysis. Our results suggest that aPKC isoforms orchestrate the formation of the podocyte processes essential for normal glomerular development and kidney function. Defective aPKC signaling results in a dramatically simplified glomerular architecture, causing severe proteinuria and perinatal death.

Vps34 deficiency reveals the importance of endocytosis for podocyte homeostasis.

The molecular mechanisms that maintain podocytes and consequently, the integrity of the glomerular filtration barrier are incompletely understood. Here, we show that the class III phosphoinositide 3-kinase vacuolar protein sorting 34 (Vps34) plays a central role in modulating endocytic pathways, maintaining podocyte homeostasis. In mice, podocyte-specific conditional knockout of Vps34 led to early proteinuria, glomerular scarring, and death within 3-9 weeks of age. Vps34-deficient podocytes exhibited substantial vacuolization and foot process effacement. Although the formation of autophagosomes and autophagic flux were impaired, comparisons between podocyte-specific Vps34-deficient mice, autophagy-deficient mice, and doubly deficient mice suggested that defective autophagy was not primarily responsible for the severe phenotype caused by the loss of Vps34. In fact, Rab5-positive endosomal compartments, endocytosis, and fluid-phase uptake were severely disrupted in Vps34-deficient podocytes. Vps34 deficiency in nephrocytes, the podocyte-like cells of Drosophila melanogaster, resulted in a block between Rab5- and Rab7-positive endosomal compartments. In summary, these data identify Vps34 as a major regulator of endolysosomal pathways in podocytes and underline the fundamental roles of endocytosis and fluid-phase uptake for the maintenance of the glomerular filtration barrier.

Association of torque teno virus viremia with liver fibrosis in the first year after liver transplantation.

Introduction: Torque teno virus (TTV) replication is controlled by immune status, mirroring a degree of immunosuppression after solid organ transplantation. TTV viraemia (TTVv) was associated with acute cellular rejection and infection within the first year after liver transplantation (LT). Long-term data on TTV after LT and correlation with graft injury from protocol biopsies are limited. Methods: One hundred plasma samples paired with graft biopsies from a prospective single-center biorepository were analyzed. Results: The median time post-LT was 23 months (range, 2-298). TTVv was detectable in 97%. TTVv decreased over time after LT and showed a significant decline from year 1 to later time points. Hence, TTVv correlated negatively with histologic liver fibrosis (liver allograft fibrosis and Ishak scores) and positively with the overall immunosuppression degree quantified by an immunosuppression score in the first year after LT. There was no association with dosages or trough levels of single immunosuppressants. The pharmacodynamic marker TTVv did not correlate with pharmacokinetic assessments of immunosuppression degree [calcineurin inhibitor (CNI) trough levels or immunosuppressant dosages]-our clinical gold standards to guide immunosuppressive therapy. TTVv was independently associated with histologically proven liver fibrosis after LT in the first year after LT in multivariate analysis. Discussion: The independent association of histological graft fibrosis with lower TTVv in year 1 underscores that a pharmacodynamic marker would be preferable to individualize immunosuppression after LT. However, a high variability of TTVv at the low immunosuppression doses given after the first year precludes TTV as a clinically useful marker after LT in the long-term liver transplant recipients.