Marius C. van den Heuvel

Expression of the Gastrin-Releasing Peptide Receptor, the Prostate Stem Cell Antigen and the Prostate-Specific Membrane Antigen in Lymph Node and Bone Metastases of Prostate Cancer

Cell membrane antigens like the gastrin‐releasing peptide receptor (GRPR), the prostate stem cell antigen (PSCA), and the prostate‐specific membrane antigen (PSMA), expressed in prostate cancer, are attractive targets for new therapeutic and diagnostic applications. Therefore, we investigated in this study whether these antigens are expressed in metastasized prostate cancer.

The angiogenic makeup of human hepatocellular carcinoma does not favor vascular endothelial growth factor/angiopoietin-driven sprouting neovascularization.

Quantitative data on the expression of multiple factors that control angiogenesis in hepatocellular carcinoma (HCC) are limited. A better understanding of the mechanisms underlying angiogenesis in HCC will improve the rational choice of anti‐angiogenic treatment. We quantified gene and protein expression of members of the vascular endothelial growth factor (VEGF) and angiopoietin systems and studied localization of VEGF, its receptors VEGFR‐1 and VEGFR‐2, Angiopoietin (Ang)‐1 and Ang‐2, and their receptor, in HCC in noncirrhotic and cirrhotic livers. We employed real‐time reverse transcription polymerase chain reaction (RT‐PCR), western blot, and immunohistology, and compared the outcome with highly angiogenic human renal cell carcinoma (RCC). HCC in noncirrhotic and cirrhotic livers expressed VEGF and its receptors to a similar extent as normal liver, although in cirrhotic background, VEGFR‐2 levels in both tumor and adjacent tissue were decreased. Ang‐1 expression was slightly increased compared with normal liver, whereas Tie‐2 was strongly down‐regulated in the tumor vasculature. Ang‐2 messenger RNA (mRNA) levels were also low in HCCs of both noncirrhotic and cirrhotic livers, implying that VEGF‐driven angiogenic sprouting accompanied by angiopoietin‐driven vascular destabilization is not pronounced. In RCC, VEGF‐A levels were one order of magnitude higher. At the same time, endothelially expressed Ang‐2 was over 30‐fold increased compared with expression in normal kidney, whereas Ang‐1 expression was decreased. Conclusion: In hepatocellular carcinoma, tumor vascularization is not per se VEGF/angiopoietin driven. However, increased CD31 expression and morphological changes representative of sinusoidal capillarization in tumor vasculature indicate that vascular remodeling is taking place. This portends that therapeutic intervention of HCC at the level of the vasculature is optional, and that further studies into the molecular control thereof are warranted. (HEPATOLOGY 2008.)

Predictive factors for portal fibrosis in pediatric liver transplant recipients

Background. Recent histopathological studies showed an unexpected high incidence of pathological changes in asymptomatic survivors after pediatric liver transplantation. The aim of this study was to analyze the occurrence of histological abnormalities, to assess the clinical significance, and to identify predictive factors for these pathological changes. Methods. The first annual protocol graft biopsies of 84 consecutive liver transplants were analyzed and correlated with concomitant liver function tests. Identification of predictive factors for the histological abnormalities in the biopsies was performed by a multivariate logistic regression analysis. Results. The incidence of portal fibrosis (PF) was 31%. Liver function tests showed except for the albumin level, an increase in the PF group compared with the group without PF. Mean values of alkaline phosphatase and direct bilirubin were 264 U/liter and 3 &mgr;mol/liter, respectively, in the normal group, and 435 U/liter and 23 &mgr;mol/liter, respectively, in the PF group (P =0.043 and 0.037). Eight of 19 univariantly tested variables were entered into a logistic regression model: cold ischemia time, preservation solution, type of allograft, cytomegalovirus recipient status, type of biliary reconstruction, biliary complications, graft complications, and rejection. A significant positive correlation with PF was found for cold ischemia time, biliary complications, and cytomegalovirus status. Acute rejection showed a negative correlation. Conclusions. The incidence of PF within 1 year post liver transplantation was 31%. This finding was accompanied by cholestatic liver function test abnormalities. Factors predisposing to PF were a prolonged cold ischemia time, biliary complications, and a positive cytomegalovirus recipient status. Acute rejection seemed to prevent for PF.

Donor and Recipient Contribution to Transplant Vasculopathy in Chronic Renal Transplant Dysfunction

Background. Chronic transplant dysfunction is the leading cause of long-term renal allograft loss. One of the histologic hallmarks of chronic transplant dysfunction is transplant vasculopathy characterized by accumulation of smooth muscle cells (SMCs) in the arterial subendothelial space, leading to ischemic graft failure. Currently, no therapy is available for transplant vasculopathy, and knowledge of the origin (donor vs. recipient) of neointimal cells may contribute to develop adequate strategies. Methods. Origin of neointimal SMCs, endothelial, and tubular cells was determined in four nephrectomy samples from male recipients transplanted with a female kidney. Recipient-derived cells were detected using X- and Y-chromosome-specific fluorescent in situ hybridization combined with immunofluorescent staining. Specificity and sensitivity of fluorescent in situ hybridization were determined with corresponding controls. Results. No Y-chromosome-positive cells were detected in the female to female graft, whereas approximately 31% of nucleated cells in male to male grafts had a detectable Y-chromosome. In female to male grafts, a recipient-derived population of neointimal &agr;-smooth muscle actin-positive SMCs were detected (6%, range 3%–11%). Percentages of recipient-derived arterial endothelial cells, glomerular endothelial cells, and tubular epithelial cells were 14% (range 4%–32%), 19% (range 7%–31%) and 3% (range 2%–5%), respectively. Conclusions. Both donor- and recipient-derived cells contribute to vascular remodeling in clinical renal transplantation. The presence of &agr;-smooth muscle actin in donor- and recipient-derived cells supports a constructive role for these cells in neointimal formation. However, the predominance of donor-derived cells in the neointima points to these cells as the likely therapeutic target.

Impaired regeneration of biliary cells in human chronic liver allograft rejection. Special emphasis on the role of the finest branches of the biliary tree

Severe loss of bile ducts is a hallmark of chronic liver rejection. We hypothesize that loss of the finest branches of the biliary tree, including the intralobular segments, contributes to an impaired regenerative response of bile ducts in chronic rejection. The number and proliferative response of bile ducts, intraportal ductules, and extraportal biliary cells were studied in graft biopsies of 12 chronic‐rejection patients. Twenty‐two long‐term‐surviving patients who experienced acute rejection without chronic rejection served as controls. Reperfusion, 1‐week, 1‐month, and 1‐year biopsies were studied. Monochlonal antibody Ki67 was applied to assess proliferation, and cytokeratin 7 antibody to identify bile ducts, intraportal ductules, and extraportal biliary cells. Progressive loss of bile ducts, intraportal ductules, and extraportal biliary cells was observed in the chronic‐rejection group. In controls, all of these structures remained well preserved. Additionally, a significant increase of intraportal ductules was present at 1 week in controls, which was not the case in the chronic‐rejection group. Proliferative activity of intraportal ductules and extraportal biliary cells was significantly increased at 1 week in controls. This proliferative activity was higher in the intraportal ductules of controls, compared with the chronic‐rejection group. After 1 week, proliferative activity was virtually absent in both groups. In conclusion, our results showed that a deficient proliferative response of the finer branches of the biliary tree, including its intralobular segments, might contribute to bile‐duct loss in chronic rejection. This finding supports the postulation that these structures represent a regenerative compartment of the biliary unit in the liver. (Liver Transpl 2004;10:28–35.)

How Normal Is the Liver in Which the Inflammatory Type Hepatocellular Adenoma Develops

The inflammatory type hepatocellular adenoma (IHCA) is a subtype of HCA which is a benign liver tumor, predominantly occurring in young women in an otherwise normal liver. IHCA contains either a mutation of gp130 or STAT3. Both mutations lead to a similar morphologic phenotype and to increased expression of C-reactive protein (CRP) and/or serum amyloid-A (SAA). IHCA comprised about 40% of all HCAs and is associated with obesity. We investigated the histomorphological and immunophenotypical changes of the nontumorous liver of 32 resected IHCA specimens. Similar types of changes are present in samples taken adjacent to tumor and distant ones. The lobular architecture is well preserved. Mild/moderate steatosis is found in a high frequency which is in accordance with the median BMI of 32 in our cases. Of note are the regular findings of sinusoidal dilatation, single arteries, and minute CRP foci which are all features of HCA. These distinct CRP foci are mostly found in cases of multiple IHCA which indicates that the remnant liver may also contain IHCA foci. These findings show that the nonlesional liver in IHCA does contain abnormalities, and this may have consequences for the followup, especially since it is known that obesity may stimulate malignant growth.

Molecular Characterization of the Vascular Features of Focal Nodular Hyperplasia and Hepatocellular Adenoma: A Role for Angiopoietin-1

Focal nodular hyperplasia (FNH) and hepatocellular adenoma (HCA) are two hepatic nodular lesions of different etiologies. FNH, a polyclonal lesion, is assumed to be a regenerative reaction following a vascular injury, whereas HCA is a monoclonal, benign neoplastic lesion. In addition to features that are predominantly found in either FNH or HCA (e.g., dystrophic vessels in FNH and single arteries in HCA), FNH and HCA share morphological vascular abnormalities such as dilated sinusoids. We hypothesized that these anomalous vascular features are associated with altered expression of growth factors involved in vascular remodeling. This was based on reports of morphologically abnormal hepatic vasculature and nodular lesions in transgenic models of hepatocytic overexpression of angiopoietin‐1 (Ang‐1), a member of the angiopoietin family, which is crucially involved in vascular morphogenesis and homeostasis. We investigated gene and protein expression of members of the angiopoietin system and vascular endothelial growth factor A (VEGF‐A) and its receptors in 9 FNH samples, 13 HCA samples, and 9 histologically normal livers. In comparison with normal samples, a significant increase in Ang‐1 was found in FNH (P < 0.01) and HCA (P < 0.05), whereas no significant changes in Ang‐2, receptor tyrosine kinase with immunoglobulin‐like and EGF‐like domains 2, VEGF‐A, or vascular endothelial growth factor receptor 2 (VEGFR‐2) were observed. Conclusion: Because of the different etiological contexts of a preceding vascular injury in FNH and a neoplastic growth in HCA, Ang‐1 might exert different effects on the vasculature in these lesions. In FNH, it could predominantly stimulate recruitment of myofibroblasts and result in dystrophic vessels, whereas in HCA, it may drive vascular remodeling that produces enlarged vessels and arterial sprouting that generates single arteries. Hepatology 2010

The renal angiopoietin/Tie2 system in lethal human sepsis

Sepsis-induced multi-organ dysfunction syndrome (MODS) still has a high mortality. Improvements await a better understanding of the pathophysiological mechanisms. The angiopoietin (Ang)1/2 and Tie2 (tyrosine kinase with immunoglobulin and epidermal growth factor homology domains 2) ligand/receptor system is an important regulator of endothelial cell responses to severe insults. Plasma Ang2 levels are prognostic in sepsis, but data on Ang/Tie responses in organs in humans are lacking.. We hypothesized that, in kidneys of patients who died of sepsis with acute kidney injury (AKI), the Ang/Tie signaling system is changed in such a way that microvessels become destabilized.

Hypothermic oxygenated machine perfusion reduces bile duct reperfusion injury after transplantation of donation after circulatory death livers

Dual hypothermic oxygenated machine perfusion (DHOPE) of the liver has been advocated as a method to reduce ischemia/reperfusion injury (IRI). This study aimed to determine whether DHOPE reduces IRI of the bile ducts in donation after circulatory death (DCD) liver transplantation. In a recently performed phase 1 trial, 10 DCD livers were preserved with DHOPE after static cold storage (SCS; www.trialregister.nl NTR4493). Bile duct biopsies were obtained at the end of SCS (before DHOPE; baseline) and after graft reperfusion in the recipient. Histological severity of biliary injury was graded according to an established semiquantitative grading system. Twenty liver transplantations using DCD livers not preserved with DHOPE served as controls. Baseline characteristics and the degree of bile duct injury at baseline (end of SCS) were similar between both groups. In controls, the degree of stroma necrosis (P = 0.002) and injury of the deep peribiliary glands (PBG; P = 0.02) increased after reperfusion compared with baseline. In contrast, in DHOPE‐preserved livers, the degree of bile duct injury did not increase after reperfusion. Moreover, there was less injury of deep PBG (P = 0.04) after reperfusion in the DHOPE group compared with controls. In conclusion, this study suggests that DHOPE reduces IRI of bile ducts after DCD liver transplantation. Liver Transplantation 24 655–664 2018 AASLD.

Tubular iron deposition and iron handling proteins in human healthy kidney and chronic kidney disease

Iron is suggested to play a detrimental role in the progression of chronic kidney disease (CKD). The kidney recycles iron back into the circulation. However, the localization of proteins relevant for physiological tubular iron handling and their potential role in CKD remain unclear. We examined associations between iron deposition, expression of iron handling proteins and tubular injury in kidney biopsies from CKD patients and healthy controls using immunohistochemistry. Iron was deposited in proximal (PT) and distal tubules (DT) in 33% of CKD biopsies, predominantly in pathologies with glomerular dysfunction, but absent in controls. In healthy kidney, PT contained proteins required for iron recycling including putative iron importers ZIP8, ZIP14, DMT1, iron storage proteins L- and H-ferritin and iron exporter ferroportin, while DT only contained ZIP8, ZIP14, and DMT1. In CKD, iron deposition associated with increased intensity of iron importers (ZIP14, ZIP8), storage proteins (L-, H-ferritin), and/or decreased ferroportin abundance. This demonstrates that tubular iron accumulation may result from increased iron uptake and/or inadequate iron export. Iron deposition associated with oxidative injury as indicated by heme oxygenase-1 abundance. In conclusion, iron deposition is relatively common in CKD, and may result from altered molecular iron handling and may contribute to renal injury.