Epstein-Barr Viral Load Monitoring Strategy and the Risk for Posttransplant Lymphoproliferative Disease in Adult Liver Transplantation : A Cohort Study.

BACKGROUND: Primary infection with or reactivation of Epstein-Barr virus (EBV) can occur after liver transplant (LT) and can lead to posttransplant lymphoproliferative disease (PTLD). In pediatric LT, an EBV-DNA viral load (EBV VL) monitoring strategy, including the reduction of immunosuppression, has led to a lower incidence of PTLD. For adult LT recipients with less primary infection and more EBV reactivation, it is unknown whether this strategy is effective. OBJECTIVE: To examine the effect of an EBV VL monitoring strategy on the incidence of PTLD after LT in adults. DESIGN: Cohort study. SETTING: Two university medical centers in the Netherlands. PATIENTS: Adult recipients of first LT in Leiden between September 2003 and January 2017 with an EBV VL monitoring strategy formed the monitoring group (M1), recipients of first LT in Rotterdam between January 2003 and January 2017 without such a strategy formed the contemporary control group (C1), and those who had transplants in Leiden between September 1992 and September 2003 or Rotterdam between 1986 and January 2003 formed the historical control groups (M0 and C0, respectively). MEASUREMENTS: Influence of EBV VL monitoring on incidence of PTLD. RESULTS: After inverse probability of treatment weighting of the 4 groups to achieve a balance among the groups for important patient characteristics, differences within hospitals between the historical and recent era in cumulative incidences-expressed as the number of events per 1000 patients measured at 5-, 10-, and 15-year follow-up-showed fewer events in the contemporary era in both centers. This difference was considerably larger in the monitoring center, whereas the 95% CI included the null value of 0 for point estimates. LIMITATION: Retrospective, low statistical power, and incompletely balanced groups, and non-EBV PTLD cannot be prevented. CONCLUSION: Monitoring EBV VL may reduce PTLD incidence after LT in adults; larger studies are warranted. PRIMARY FUNDING SOURCE: None.

Tactile Perception of the Hand in Children With an Upper Neonatal Brachial Plexus Palsy.

OBJECTIVE: To investigate factors that cause impairment of hand function in children with an upper Neonatal Brachial Plexus Palsy (NBPP), we performed an in-depth analysis of tactile hand sensibility, especially the ability to correctly localize a sensory stimulus on their fingers. DESIGN: A cross-sectional investigation of children with NBPP, compared with healthy controls. The thickest Semmes-Weinstein (SW) monofilament was pressed on the radial or ulnar part of each fingertip (10 regions), while a screen prevented seeing the hand. SETTING: Tertiary referral center for nerve lesions in an academic hospital in The Netherlands. The control group was recruited at their school. PARTICIPANTS: Forty-one children with NBPP (mean age 10.0 y) and 25 controls (mean age 9.5 y; N=41). INTERVENTIONS: Not applicable. MAIN OUTCOMES MEASURES: Correct localization of the applied stimuli was evaluated, per region, per finger, and per dermatome with a test score. The affected side of the NBPP group was compared with the non-dominant hand of the controls. RESULTS: The ability to localize stimuli on the tips of the fingers in children with an upper NBPP was significantly diminished in all fingers, except for the little finger, as compared with healthy controls. Mean localization scores were 6.6 (thumb) and 6.3 (index finger) in the NBPP group and 7.6 in both fingers for controls (maximum score possible is 8.0). Localization scores were significant lower in regions attributed to dermatomes C6 (P<.001) and C7 (P=.001), but not to C8 (P=.115). CONCLUSION: Children with an upper NBPP showed a diminished and incorrect ability to localize sensory stimuli to their fingers. This finding is likely 1 of the factors underlying the impairment of hand function and should be addressed with sensory focused therapy.

Reduced Glomerular Endothelial Thrombomodulin Is Associated with Glomerular Macrophage Infiltration in Diabetic Nephropathy.

The endothelial glycoprotein thrombomodulin regulates coagulation, inflammation, and apoptosis. In diabetic mice, reduced thrombomodulin function results in diabetic nephropathy (DN). Furthermore, thrombomodulin treatment reduces renal inflammation and fibrosis. Herein, thrombomodulin expression was examined in human kidney samples to investigate the possibility of targeting thrombomodulin in patients with DN. Glomerular thrombomodulin was analyzed together with the number of glomerular macrophages in 90 autopsied diabetic cases with DN, 55 autopsied diabetic cases without DN, and 37 autopsied cases without diabetes or kidney disease. Thrombomodulin mRNA was measured in glomeruli microdissected from renal biopsies from patients with DN and nondiabetic controls. Finally, glomerular thrombomodulin was measured in diabetic mice following treatment with the selective endothelin A receptor (ETAR) blocker, atrasentan. In diabetic patients, glomerular thrombomodulin expression was increased at the mRNA level, but decreased at the protein level, compared with nondiabetic controls. Reduced glomerular thrombomodulin was associated with an increased glomerular influx of macrophages. Blocking the ETAR with atrasentan restored glomerular thrombomodulin protein levels in diabetic mice to normal levels. The reduction in glomerular thrombomodulin in diabetes likely serves as an early proinflammatory step in the pathogenesis of DN. Thrombomodulin protein may be cleaved under diabetic conditions, leading to a compensatory increase in transcription. The nephroprotective effects of ETAR antagonists in diabetic patients may be attributed to the restoration of glomerular thrombomodulin.

Increased microchimerism in peripheral blood of women with systemic lupus erythematosus: relation with pregnancy.

OBJECTIVES: We aimed to determine the presence, amount and origin of microchimerism in peripheral blood of pregnant and non-pregnant parous women with systemic lupus erythematosus (SLE) as compared to control subjects. METHODS: We performed a comparative study in which peripheral blood was drawn from eleven female non-pregnant SLE-patients and 22 control subjects, and from six pregnant SLE-patients and eleven control subjects during gestation and up to six months postpartum. Quantitative PCR for insertion-deletion polymorphisms and null alleles was used to detect microchimerism in peripheral blood mononuclear cells and granulocytes. RESULTS: Microchimerism was detected more often in non-pregnant SLE-patients than control subjects (54.4% vs. 13.6%, respectively; p=0.03). When present, the median total number of foetal chimeric cells was 5 gEq/106 in patients and 2.5gEq/106 in control subjects (p=0.048). Microchimerism was mostly foetal in origin; maternal microchimerism was detected in one patient and one control subject. In control subjects, microchimerism was always derived from only one source whereas in 50% of patients it originated from multiple sources. The pregnant patients had a significantly higher median number of foetal chimeric cells in the granulocyte fraction just after delivery than control subjects (7.5 gEq/106 vs. 0 gEq/106, respectively; p=0.02). CONCLUSIONS: Just after delivery, SLE-patients had more microchimerism than control subjects. Three months post-partum, microchimerism was no longer detectable, only to reappear many years after the last pregnancy, more often and at higher levels in SLE-patients than in control subjects. This suggests that these chimeric cells may originate from non-circulating foetal chimeric stem cells.

The VEGF Inhibitor Soluble Fms-like Tyrosine Kinase 1 Does Not Promote AKI-to-CKD Transition.

(1) Background: Soluble Fms-like tyrosine kinase 1 (sFLT1) is an endogenous VEGF inhibitor. sFLT1 has been described as an anti-inflammatory treatment for diabetic nephropathy and heart fibrosis. However, sFLT1 has also been related to peritubular capillary (PTC) loss, which promotes fibrogenesis. Here, we studied whether transfection with sFlt1 aggravates experimental AKI-to-CKD transition and whether sFLT1 is increased in human kidney fibrosis. (2) Methods: Mice were transfected via electroporation with sFlt1. After confirming transfection efficacy, mice underwent unilateral ischemia/reperfusion injury (IRI) and were sacrificed 28 days later. Kidney histology and RNA were analyzed to study renal fibrosis, PTC damage and inflammation. Renal sFLT1 mRNA expression was measured in CKD biopsies and control kidney tissue. (3) Results: sFlt1 transfection did not aggravate renal fibrosis, PTC loss or macrophage recruitment in IRI mice. In contrast, higher transfection efficiency was correlated with reduced expression of pro-fibrotic and pro-inflammatory markers. In the human samples, sFLT1 mRNA levels were similar in CKD and control kidneys and were not correlated with interstitial fibrosis or PTC loss. (4) Conclusion: As we previously found that sFLT1 has therapeutic potential in diabetic nephropathy, our findings indicate that sFLT1 can be administered at a dose that is therapeutically effective in reducing inflammation, without promoting maladaptive kidney damage.

Endoglin Is an Important Mediator in the Final Common Pathway of Chronic Kidney Disease to End-Stage Renal Disease.

Chronic kidney disease (CKD) is a slow-developing, progressive deterioration of renal function. The final common pathway in the pathophysiology of CKD involves glomerular sclerosis, tubular atrophy and interstitial fibrosis. Transforming growth factor-beta (TGF-ß) stimulates the differentiation of fibroblasts towards myofibroblasts and the production of extracellular matrix (ECM) molecules, and thereby interstitial fibrosis. It has been shown that endoglin (ENG, CD105), primarily expressed in endothelial cells and fibroblasts, can function as a co-receptor of TGF signaling. In several human organs, endoglin tends to be upregulated when chronic damage and fibrosis is present. We hypothesize that endoglin is upregulated in renal interstitial fibrosis and plays a role in the progression of CKD. We first measured renal endoglin expression in biopsy samples obtained from patients with different types of CKD, i.e., IgA nephropathy, focal segmental glomerulosclerosis (FSGS), diabetic nephropathy (DN) and patients with chronic allograft dysfunction (CAD). We showed that endoglin is upregulated in CAD patients (p < 0.001) and patients with DN (p < 0.05), compared to control kidneys. Furthermore, the amount of interstitial endoglin expression correlated with eGFR (p < 0.001) and the amount of interstitial fibrosis (p < 0.001), independent of the diagnosis of the biopsies. Finally, we investigated in vitro the effect of endoglin overexpression in TGF-ß stimulated human kidney fibroblasts. Overexpression of endoglin resulted in an enhanced ACTA2, CCN2 and SERPINE1 mRNA response (p < 0.05). It also increased the mRNA and protein upregulation of the ECM components collagen type I (COL1A1) and fibronectin (FN1) (p < 0.05). Our results suggest that endoglin is an important mediator in the final common pathway of CKD and could be used as a possible new therapeutic target to counteract the progression towards end-stage renal disease (ESRD).

What Range of Motion is Achieved 5 Years After External Rotationplasty of the Shoulder in Infants with an Obstetric Brachial Plexus Injury?

BACKGROUND: Obstetric brachial plexus injuries result from traction injuries during delivery, and 30% of these children have persisting functional limitations related to an external rotation deficit of the shoulder. Little is known about the long-term effect of soft-tissue procedures of the shoulder in patients with obstetric brachial plexus injuries. QUESTIONS/PURPOSES: (1) After soft-tissue release for patients with passive external rotation less than 20° and age younger than 2 years and for patients older than 2 years with good external rotation strength, what are the improvements in passive external rotation and abduction arcs at 1 and 5 years? (2) For patients who underwent staged tendon transfer after soft-tissue release, what are the improvements in active external rotation and abduction arcs at 1 and 5 years? (3) For patients with passive external rotation less than 20° and no active external rotation, what are the improvements in active external rotation and abduction arcs at 1 and 5 years? METHODS: This was a retrospective analysis of a longitudinally maintained institutional database. Between 1996 and 2009, 149 children underwent a soft-tissue procedure of the shoulder for an internal rotation contracture. The inclusion criteria were treatment with an internal contracture release and/or tendon transfer, a maximum age of 18 years at the time of surgery, and a minimum follow-up period of 2 years. Six patients were older than 18 years at the time of surgery and 31 children were seen at our clinic until 1 year postoperatively, but because they had good clinical results and lived far away from our center, these children were discharged to physical therapists in their hometown for annual follow-up. Thus, 112 children (59 boys) were available for analysis. Patients with passive external rotation less than 20° and age younger than 2 years and patients older than 2 years with good external rotation strength received soft-tissue release only (n = 37). Of these patients, 17 children did not have adequate active external rotation, and second-stage tendon transfer surgery was performed. For patients with passive external rotation less than 20° with no active external rotation, single-stage contracture release with tendon transfer was performed (n = 68). When no contracture was present (greater than 20° of external rotation) but the patient had an active deficit (n = 7), tendon transfer alone was performed; this group was not analyzed. A functional assessment of the shoulder was performed preoperatively and postoperatively at 6 weeks, 3 months, and annually thereafter and included abduction, external rotation in adduction and abduction, and the Mallet scale. RESULTS: Internal contracture release resulted in an improvement in passive external rotation in adduction and abduction of 29° (95% confidence interval, 21 to 38; p < 0.001) and 17° (95% CI, 10 to 24; p < 0.001) at 1 year of follow-up and 25° (95% CI, 15-35; p < 0.001) and 15° (95% CI, 7 to 24; p = 0.001) at 5 years. Because of insufficient strength of the external rotators after release, 46% of the children (17 of 37) underwent an additional tendon transfer for active external rotation, resulting in an improvement in active external rotation in adduction and abduction at each successive follow-up visit. Patients with staged transfers had improved active function; improvements in active external rotation in adduction and abduction were 49° (95% CI, 28 to 69; p < 0.05) and 45° (95% CI, 11 to 79; p < 0.001) at 1 year of follow-up and 38° (95% CI, 19 to 58; p < 0.05) and 23° (95% CI, -8 to 55; p < 0.001) at 5 years. In patients starting with less than 20° of passive external rotation and no active external rotation, after single-stage contracture release and tendon transfer, active ROM was improved. Active external rotation in adduction and abduction were 75° (95% CI, 66 to 84; p < 0.001) and 50° (95% CI, 43 to 57; p < 0.001) at 1 year of follow-up and 65° (95% CI, 50 to 79; p < 0.001) and 40° (95% CI, 28 to 52; p < 0.001) at 5 years. CONCLUSION: Young children with obstetric brachial plexus injuries who have internal rotation contractures may benefit from soft-tissue release. When active external rotation is lacking, soft-tissue release combined with tendon transfer improved active external rotation in this small series. Future studies on the degree of glenohumeral deformities and functional outcome might give more insight into the level of increase in external rotation. LEVEL OF EVIDENCE: Level III, therapeutic study.

Endoglin Promotes Myofibroblast Differentiation and Extracellular Matrix Production in Diabetic Nephropathy.

Diabetic nephropathy (DN) is a complication of diabetes mellitus that can lead to proteinuria and a progressive decline in renal function. Endoglin, a co-receptor of TGF-ß, is known primarily for regulating endothelial cell function; however, endoglin is also associated with hepatic, cardiac, and intestinal fibrosis. This study investigates whether endoglin contributes to the development of interstitial fibrosis in DN. Kidney autopsy material from 80 diabetic patients was stained for endoglin and Sirius Red and scored semi-quantitatively. Interstitial endoglin expression was increased in samples with DN and was correlated with Sirius Red staining (p < 0.001). Endoglin expression was also correlated with reduced eGFR (p = 0.001), increased creatinine (p < 0.01), increased systolic blood pressure (p < 0.05), hypertension (p < 0.05), and higher IFTA scores (p < 0.001). Biopsy samples from DN patients were also co-immunostained for endoglin together with CD31, CD68, vimentin, or α-SMA Endoglin co-localized with both the endothelial marker CD31 and the myofibroblast marker α-SMA. Finally, we used shRNA to knockdown endoglin expression in a human kidney fibroblast cell line. We found that TGF-ß1 stimulation upregulated SERPINE1, CTGF, and ACTA2 mRNA and α-SMA protein, and that these effects were significantly reduced in fibroblasts after endoglin knockdown. Taken together, these data suggest that endoglin plays a role in the pathogenesis of interstitial fibrosis in DN.

Glomerular C4d deposition can precede the development of focal segmental glomerulosclerosis.

Recent studies suggest that complement plays a role in the pathogenesis of focal segmental glomerulosclerosis (FSGS). Moreover, co-localization of IgM and C3 deposits with FSGS lesions has frequently been reported. Here, we investigated whether glomerular complement deposition precedes the development of FSGS and whether it represents local complement activation. Renal biopsies from 40 patients with primary FSGS, 84 patients with minimal change disease, and 10 healthy individuals were stained for C4d, C1q, and mannose-binding lectin. C4d deposits were also measured in renal allograft biopsies from 34 patients with native primary FSGS, 18 of whom subsequently developed recurrent FSGS. Lastly, we measured C4d deposits in the Munich Wistar Frömter rat model of FSGS. The prevalence of C4d-positive glomeruli was significantly higher among patients with FSGS (73%) compared to patients with minimal change disease (21%) and healthy individuals (10%). Moreover, segmental sclerosis was absent in 42% of C4d-positive glomeruli. Glomerular C1q was significantly more prevalent in FSGS compared to minimal change disease or healthy individuals, while mannose-binding lectin was infrequently observed. C4d deposition was significantly more prevalent in recurrent FSGS (72%) before the development of sclerotic lesions compared to control transplant samples (27%). Finally, at the onset of albuminuria but before the development of FSGS lesions, Munich Wistar Frömter rats had a significantly higher percentage of C4d-positive glomeruli (31%) compared to control rats (4%). Thus, glomerular C4d deposition can precede the development of FSGS, suggesting that complement activation may play a pathogenic role in the development of FSGS.

Complement-mediated microangiopathy in IgA nephropathy and IgA vasculitis with nephritis.

Complement factor C4d was recently observed in renal biopsies from patients who had IgA nephropathy and a poor prognosis. We previously reported that C4d is a common denominator in microangiopathies. In this retrospective cohort study, we investigated whether C4d is a marker of microangiopathy in both IgA nephropathy and IgA vasculitis with nephritis, and whether patients with C4d and microangiopathy have poor renal outcome. We examined 128 renal biopsies from adult and pediatric patients, including normotensive and hypertensive patients, who presented with IgA nephropathy or IgA vasculitis with nephritis. Biopsies were re-evaluated in accordance with the Oxford classification, scored for additional lesions, and stained for complement proteins using immunohistochemistry, including C4d and C5b-9. Clinical data were collected with a mean (±SD) follow-up period of 51 ± 39 months. Changes in estimated glomerular filtration rate over time were compared using linear mixed-effects models. Renal survival was analyzed using multivariable Cox regression. Microangiopathic lesions were present in 20% of all biopsies (23% and 9% of patients with IgA nephropathy and IgA vasculitis with nephritis, respectively). Microangiopathy was associated with C4d and C5b-9 deposits, a higher number of chronic lesions, and hypertension (all p < 0.05). Patients with C4d and microangiopathic lesions had significantly poorer renal survival than patients without these findings, corrected for hypertension (p < 0.01). In conclusion, patients with IgA nephropathy or IgA vasculitis with nephritis with a combination of C4d positivity and microangiopathy comprise a clinical subgroup with an increased number of chronic lesions, lower estimated glomerular filtration rate, and poorer renal survival, even when corrected for hypertension. These data suggest that complement activation is involved in the development of microangiopathy in patients with IgA nephropathy and IgA vasculitis with nephritis, and that complement-mediated microangiopathy contributes to disease progression.

Endoglin Mediates Vascular Endothelial Growth Factor-A-Induced Endothelial Cell Activation by Regulating Akt Signaling.

In diabetic nephropathy, differential expression of growth factors leads to vascular changes, including endothelial cell activation, monocyte infiltration, and inflammation. Endoglin plays an important role in endothelial function and is also associated with inflammation. In the kidney, vascular endoglin expression is increased in animal models of renal injury, where it contributes to disease severity, possibly by promoting endothelial cell activation and inflammation. Herein, we investigated whether endoglin expression is associated with diabetic nephropathy. In addition, we examined whether reducing endothelial endoglin expression in vitro affects endothelial cell activation and monocyte adhesion and, if so, which intracellular pathways are involved. Finally, we analyzed whether glomerular endoglin expression is correlated with endothelial cell activation in patients with diabetic nephropathy. Endoglin levels were significantly increased in mice with type 1 diabetes compared with control mice. Reducing endoglin expression in cultured endothelial cells significantly impaired the vascular endothelial growth factor-A-induced up-regulation of activation markers and monocyte adhesion. This was mediated by increased phosphorylation of Akt, thereby inhibiting activating transcription factor 2 phosphorylation, which regulates vascular cell adhesion molecule-1 (VCAM1) gene transcription in these cells. Last, endoglin colocalized with VCAM-1 in the glomeruli of diabetic patients, glomerular VCAM-1 expression was significantly increased in these patients, and this increase in VCAM-1 expression was correlated with increased glomerular endoglin expression. Thus, targeting endoglin function may have therapeutic value in patients at risk for diabetic nephropathy.

Etanercept in patients with inflammatory hand osteoarthritis (EHOA): a multicentre, randomised, double-blind, placebo-controlled trial.

OBJECTIVE: Hand osteoarthritis is a prevalent disease with limited treatment options. Since joint inflammation is often present, we investigated tumour necrosis factor (TNF) as treatment target in patients with proven joint inflammation in a proof-of-concept study. METHODS: This 1-year, double-blind, randomised, multicentre trial (NTR1192) enrolled patients with symptomatic erosive inflammatory hand osteoarthritis. Patients flaring after non-steroidal anti-inflammatory drug washout were randomised to etanercept (24 weeks 50 mg/week, thereafter 25 mg/week) or placebo. The primary outcome was Visual Analogue Scale (VAS) pain at 24 weeks. Secondary outcomes included clinical and imaging outcomes (radiographs scored using Ghent University Scoring System (GUSS, n=54) and MRIs (n=20)). RESULTS: Of 90 patients randomised to etanercept (n=45) or placebo (n=45), respectively, 12 and 10 discontinued prematurely. More patients on placebo discontinued due to inefficacy (6 vs 3), but fewer due to adverse effects (1 vs 6). The mean between-group difference (MD) in VAS pain was not statistically significantly different (-5.7 (95% CI -15.9 to 4.5), p=0.27 at 24 weeks; - 8.5 (95% CI -18.6 to 1.6), p=0.10 at 1 year; favouring etanercept). In prespecified per-protocol analyses of completers with pain and inflammation at baseline (n=61), MD was -11.8 (95% CI -23.0 to -0.5) (p=0.04) at 1 year. Etanercept-treated joints showed more radiographic remodelling (delta GUSS: MD 2.9 (95% CI 0.5 to 5.4), p=0.02) and less MRI bone marrow lesions (MD -0.22 (95% CI -0.35 to -0.09), p = 0.001); this was more pronounced in joints with baseline inflammation. CONCLUSION: Anti-TNF did not relieve pain effectively after 24 weeks in erosive osteoarthritis. Small subgroup analyses showed a signal for effects on subchondral bone in actively inflamed joints, but future studies to confirm this are warranted.

The Epstein-Barr Virus Glycoprotein gp150 Forms an Immune-Evasive Glycan Shield at the Surface of Infected Cells.

Cell-mediated immunity plays a key role in host control of viral infection. This is exemplified by life-threatening reactivations of e.g. herpesviruses in individuals with impaired T-cell and/or iNKT cell responses. To allow lifelong persistence and virus production in the face of primed immunity, herpesviruses exploit immune evasion strategies. These include a reduction in viral antigen expression during latency and a number of escape mechanisms that target antigen presentation pathways. Given the plethora of foreign antigens expressed in virus-producing cells, herpesviruses are conceivably most vulnerable to elimination by cell-mediated immunity during the replicative phase of infection. Here, we show that a prototypic herpesvirus, Epstein-Barr virus (EBV), encodes a novel, broadly acting immunoevasin, gp150, that is expressed during the late phase of viral replication. In particular, EBV gp150 inhibits antigen presentation by HLA class I, HLA class II, and the non-classical, lipid-presenting CD1d molecules. The mechanism of gp150-mediated T-cell escape does not depend on degradation of the antigen-presenting molecules nor does it require gp150's cytoplasmic tail. Through its abundant glycosylation, gp150 creates a shield that impedes surface presentation of antigen. This is an unprecedented immune evasion mechanism for herpesviruses. In view of its likely broader target range, gp150 could additionally have an impact beyond escape of T cell activation. Importantly, B cells infected with a gp150-null mutant EBV displayed rescued levels of surface antigen presentation by HLA class I, HLA class II, and CD1d, supporting an important role for iNKT cells next to classical T cells in fighting EBV infection. At the same time, our results indicate that EBV gp150 prolongs the timespan for producing viral offspring at the most vulnerable stage of the viral life cycle.

New function of the myostatin/activin type I receptor (ALK4) as a mediator of muscle atrophy and muscle regeneration.

Skeletal muscle fibrosis and impaired muscle regeneration are major contributors to muscle wasting in Duchenne muscular dystrophy (DMD). Muscle growth is negatively regulated by myostatin (MSTN) and activins. Blockage of these pathways may improve muscle quality and function in DMD. Antisense oligonucleotides (AONs) were designed specifically to block the function of ALK4, a key receptor for the MSTN/activin pathway in skeletal muscle. AON-induced exon skipping resulted in specific Alk4 down-regulation, inhibition of MSTN activity, and increased myoblast differentiation in vitro Unexpectedly, a marked decrease in muscle mass (10%) was found after Alk4 AON treatment in mdx mice. In line with in vitro results, muscle regeneration was stimulated, and muscle fiber size decreased markedly. Notably, when Alk4 was down-regulated in adult wild-type mice, muscle mass decreased even more. RNAseq analysis revealed dysregulated metabolic functions and signs of muscle atrophy. We conclude that ALK4 inhibition increases myogenesis but also regulates the tight balance of protein synthesis and degradation. Therefore, caution must be used when developing therapies that interfere with MSTN/activin pathways.-Pasteuning-Vuhman, S., Boertje-van der Meulen, J. W., van Putten, M., Overzier, M., ten Dijke, P., Kielbasa, S. M., Arindrarto, W., Wolterbeek, R., Lezhnina, K. V., Ozerov, I. V., Aliper, A. M., Hoogaars, W. M., Aartsma-Rus, A., Loomans, C. J. M. New function of the myostatin/activin type I receptor (ALK4) as a mediator of muscle atrophy and muscle regeneration.

Donor-specific anti-HLA antibodies are not associated with nonanastomotic biliary strictures but both are independent risk factors for graft loss after liver transplantation.

Donor-specific alloantibodies (DSA) have been associated with rejection and shorter graft survival after orthotopic liver transplantation (OLT). We examined the role of DSA in nonanastomotic biliary strictures (NAS) after OLT. Patients receiving first OLT who developed NAS (n = 68) and a control group without NAS (n = 83), with pre-OLT and 12 months post-OLT serum samples, were included. DSA were specified using the Luminex single antigen test. Risk factors for NAS and graft survival were analyzed. The presence of preformed DSA was not significantly different between patients with NAS and controls (P = .89). After 12 months, 26.5% of NAS patients and 16.9% of controls had generated de novo DSA (P = .15). Neither de novo class I DSA nor de novo class II DSA were associated with NAS. De novo DSA generally developed after the diagnosis of NAS. Time-dependent regression analysis identified both NAS (aHR 8.05, CI 3.28 - 19.77, P < .01) and de novo class II DSA (aHR 2.84, CI 1.38 - 5.82, P < .01) as independent risk factors for graft loss. Preformed or de novo DSA were not associated with the development of NAS. However, NAS as well as de novo class II DSA were independent risk factors for graft loss after OLT.

Low wintertime pre-diagnostic vitamin D status is associated with an increased risk of internal malignancies in kidney transplant recipients.

Low serum 25-hydroxyvitamin D (25OHD) concentrations have been associated with increased cancer risk, but the relative importance of seasonality, i.e. high summer concentrations versus low winter concentrations, is unclear. We investigated this issue in a high risk group: kidney transplant recipients with known increased risk of cancer and low vitamin D statuses. We examined the relationship between registered concentrations of 25OHD binned by quarter and subsequent risk of internal malignancy or cutaneous squamous cell carcinoma in 1112 kidney transplant recipients. Hazard ratios for internal malignancies were significantly increased with lower pre-diagnostic 25OHD concentrations in the first quarter of the year (January-March); a 1.4 fold increase (95%CI 1.1;1.7) per 10 nmol L-1 decrease in 25OHD. Except for women in April-June (1.3 (1.01;1.7) per 10 nmol L-1 decrease) pre-diagnostic 25OHD concentrations in the other quarters were not statistically significantly associated with internal malignancies. Higher 25OHD concentrations tended to be associated with the development of cutaneous squamous cell carcinomas, independent of the time of the year. Our study indicates that low wintertime 25OHD concentrations are associated with an increased risk of internal malignancies and that transplant recipients may benefit from wintertime vitamin D supplementation. Our findings need further corroboration, but suggest that the lowest concentrations of vitamin D, which occur in winter, are important for the risk of internal malignancies.

Apolipoprotein C-I plays a role in the pathogenesis of glomerulosclerosis.

Diabetic nephropathy is the leading cause of end-stage renal disease. Diabetic patients have increased plasma concentrations of apolipoprotein C-I (apoCI), and meta-analyses found that a polymorphism in APOC1 is associated with an increased risk of developing nephropathy. To investigate whether overexpressing apoCI contributes to the development of kidney damage, we studied renal tissue and peritoneal macrophages from APOC1 transgenic (APOC1-tg) mice and wild-type littermates. In addition, we examined renal material from autopsied diabetic patients with and without diabetic nephropathy and from autopsied control subjects. We found that APOC1-tg mice, but not wild-type mice, develop albuminuria, renal dysfunction, and glomerulosclerosis with increased numbers of glomerular M1 macrophages. Moreover, compared to wild-type macrophages, stimulated macrophages isolated from APOC1-tg mice have increased cytokine expression, including TNF-alpha and TGF-beta, both of which are known to increase the production of extracellular matrix proteins in mesangial cells. These results suggest that APOC1 expression induces glomerulosclerosis, potentially by increasing the cytokine response in macrophages. Furthermore, we detected apoCI in the kidneys of diabetic patients, but not in control kidneys. Moreover, patients with diabetic nephropathy have significantly more apoCI present in glomeruli compared to diabetic patients without nephropathy, suggesting that apoCI could be involved in the development of diabetic nephropathy. ApoCI co-localized with macrophages. Therefore, apoCI is a promising new therapeutic target for patients at risk of developing nephropathy. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Inclination but not anatomical reconstruction is related with higher cobalt levels in MoM hip arthroplasty.

Metal on Metal total hip arthroplasty is associated with elevated serum cobalt levels. In this study we investigate if there is a relation between the inclination and anteversion angle of the cup and the anatomical reconstruction of the hip on the serum cobalt level. Postoperative cobalt serum levels were measured in 250 patients with the M2a-38 cup and Taperloc stem combination. On standardized radiographs inclination and anteversion angle, lower limb length, lateral offset and center of rotation distance were evaluated. A difference of more than 5 millimeter compared to the preoperative situation was considered as a non-anatomical reconstruction. For every 10 degrees increase in inclination the cobalt level increased 14% (p = 0.036). Women with the same cup inclination angle showed 34% higher cobalt levels than men (p = 0.013). No relation was found between the anteversion angle, anatomical reconstruction and the serum cobalt levels. A higher inclination of the cup leads to higher serum cobalt levels, but a non-anatomical reconstruction has no influence on serum cobalt levels.

The VEGF-A inhibitor sFLT-1 improves renal function by reducing endothelial activation and inflammation in a mouse model of type 1 diabetes.

AIMS/HYPOTHESIS: Animal models of diabetic nephropathy show increased levels of glomerular vascular endothelial growth factor (VEGF)-A, and several studies have shown that inhibiting VEGF-A in animal models of diabetes can prevent albuminuria and glomerular hypertrophy. However, in those studies, treatment was initiated before the onset of kidney damage. Therefore, the aim of this study was to investigate whether transfecting mice with the VEGF-A inhibitor sFlt-1 (encoding soluble fms-related tyrosine kinase 1) can reverse pre-existing kidney damage in a mouse model of type 1 diabetes. In addition, we investigated whether transfection with sFlt-1 can reduce endothelial activation and inflammation in these mice. METHODS: Subgroups of untreated 8-week-old female C57BL/6J control (n = 5) and diabetic mice (n = 7) were euthanised 5 weeks after the start of the experiment in order to determine the degree of kidney damage prior to treatment with sFLT-1. Diabetes was induced with three i.p. injections of streptozotocin (75 mg/kg) administered at 2 day intervals. Diabetic nephropathy was then investigated in diabetic mice transfected with sFlt-1 (n = 6); non-diabetic, non-transfected control mice (n = 5); non-diabetic control mice transfected with sFlt-1(n = 10); and non-transfected diabetic mice (n = 6). These mice were euthanised at the end of week 15. Transfection with sFlt-1 was performed in week 6. RESULTS: We found that transfection with sFlt-1 significantly reduced kidney damage by normalising albuminuria, glomerular hypertrophy and mesangial matrix content (i.e. glomerular collagen type IV protein levels) (p < 0.001). We also found that transfection with sFlt-1 reduced endothelial activation (p < 0.001), glomerular macrophage infiltration (p < 0.001) and glomerular TNF-α protein levels (p < 0.001). Finally, sFLT-1 decreased VEGF-A-induced endothelial activation in vitro (p < 0.001). CONCLUSIONS/INTERPRETATION: These results suggest that sFLT-1 might be beneficial in treating diabetic nephropathy by inhibiting VEGF-A, thereby reducing endothelial activation and glomerular inflammation, and ultimately reversing kidney damage.

Effect of rituximab on malignancy risk in patients with ANCA-associated vasculitis.

OBJECTIVES: Patients with antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) treated with cyclophosphamide have an increased malignancy risk compared with the general population. We investigated whether treatment with rituximab instead of cyclophosphamide has decreased the malignancy risk in patients with AAV. METHODS: The study included patients with AAV treated at a tertiary vasculitis referral centre between 2000 and 2014. The malignancy incidence in these patients was compared with the incidence in the general population by calculating standardised incidence ratios (SIRs), adjusted for sex, age and calendar year. Malignancy incidence was compared between rituximab-treated and cyclophosphamide-treated patients. RESULTS: Of the 323 included patients, 33 developed a total of 45 malignancies during a mean follow-up of 5.6 years. This represented a 1.89-fold increased (95% CI 1.38 to 2.53) malignancy risk, and a non-significantly increased risk if non-melanoma skin cancer was excluded (SIR, 1.09; 95% CI 0.67 to 1.69). The risk of non-melanoma skin cancer was 4.58-fold increased (95% CI 2.96 to 6.76). Cyclophosphamide-treated patients had an increased malignancy risk compared with the general population (SIR, 3.10; 95% CI 2.06 to 4.48). In contrast, rituximab-treated patients had a malignancy risk similar to the general population (SIR, 0.67; 95% CI 0.08 to 2.43). The malignancy risk in cyclophosphamide-treated patients was 4.61-fold higher (95% CI 1.16 to 39.98) than in rituximab-treated patients. CONCLUSIONS: The malignancy risk in patients with AAV was lower in rituximab-treated patients than in cyclophosphamide-treated patients. Notably, rituximab treatment was not associated with an increased malignancy risk compared with the general population. Rituximab could therefore be a safe alternative to cyclophosphamide in the treatment of AAV.