Cutaneous Brucellosis unmasked as Aureimonas altamirensis in a wound culture.

Aureimonas altamirensis was isolated from a wound culture and initially misidentified as Brucella melitensis by the VITEK® 2 system. The VITEK-MS did not provide identification whereas the Bruker MALDI-ToF MS system and 16-S sequencing revealed a clear identification, which highlights the importance of inclusion of species in databases for accurate and fast identification of bacteria.

From a case-control survey to a diagnostic viral gastroenteritis panel for testing of general practitioners' patients.

OBJECTIVE: To evaluate the pathogenicity of a broad range of 11 possible gastroenteritis viruses, by means of statistical relationships with cases vs. controls, or Ct-values, in order to establish the most appropriate diagnostic panel for our general practitioner (GP) patients in the Netherlands (2010-2012). METHODS: Archived stool samples from 1340 cases and 1100 controls were retested using internally controlled multiplex real-time PCRs for putative pathogenic gastroenteritis viruses: adenovirus, astrovirus, bocavirus, enterovirus, norovirus GI and GII, human parechovirus, rotavirus, salivirus, sapovirus, and torovirus. RESULTS: The prevalence of any virus in symptomatic cases and asymptomatic controls was 16.6% (223/1340) and 10.2% (112/1100), respectively. Prevalence of astrovirus (adjusted odds ratio (aOR) 10.37; 95% confidence interval (CI) 1.34-80.06) and norovirus GII (aOR 3.10; CI 1.62-5.92) was significantly higher in cases versus controls. Rotavirus was encountered only in cases. We did not find torovirus and there was no statistically significant relationship with cases for salivirus (aOR 1,67; (CI) 0.43-6.54)), adenovirus non-group F (aOR 1.20; CI 0.75-1.91), bocavirus (aOR 0.85; CI 0.05-13.64), enterovirus (aOR 0.83; CI 0.50-1.37), human parechovirus (aOR 1.61; CI 0.54-4.77) and sapovirus (aOR 1.15; CI 0.67-1.98). Though adenovirus group F (aOR 6.37; CI 0.80-50.92) and norovirus GI (aOR 2.22, CI: 0.79-6.23) are known enteropathogenic viruses and were more prevalent in cases than in controls, this did not reach significance in this study. The Ct value did not discriminate between carriage and disease in PCR-positive subjects. CONCLUSIONS: In our population, diagnostic gastroenteritis tests should screen for adenovirus group F, astrovirus, noroviruses GI and GII, and rotavirus. Case-control studies as ours are lacking and should also be carried out in populations from other epidemiological backgrounds.

Distribution and relevance of Dientamoeba fragilis and Blastocystis species in gastroenteritis: results from a case-control study.

The actual role of Dientamoeba fragilis and Blastocystis in patients with gastrointestinal symptoms is still under debate. A multicenter case-control study was performed in The Netherlands to elucidate the clinical relevance of molecular diagnostics results in gastroenteritis (GE). Samples from this case-control study were used to perform a detailed analysis on the presence of D. fragilis and Blastocystis in relation to gastrointestinal symptoms. In the present study, a real-time PCR for Blastocystis was performed on 1374 case samples and 1026 control samples from the multicenter gastroenteritis case-control study previously tested for D. fragilis. Prevalence of both micro-organisms was highest in children under 20 years of age and lowest in the oldest age group. A significantly lower overall detection of D. fragilis and Blastocystis was found in cases (both 25.8%) as compared to controls (37.6% and 40.0%, respectively). The difference for D. fragilis was statistically significant for subjects above 20 years of age. For Blastocystis, the difference was statistically significant in all age groups, except in children less than 5 years of age. A negative relation between D. fragilis-positive cases and diarrhea was found in this study population. More GE symptoms were reported in cases without D. fragilis or Blastocystis. In the present study, prevalence of both D. fragilis and Blastocystis is lower in cases with gastroenteritic symptoms than in controls. Besides, in cases with D. fragilis or Blastocystis, no association is shown between any of the GE symptoms. Interestingly, this suggests that the presence of these protozoans may be considered characteristic of a healthy intestinal microbiome.

Epidemiology of Extended-Spectrum ß-Lactamase-Producing E. coli and Vancomycin-Resistant Enterococci in the Northern Dutch-German Cross-Border Region.

Objectives: To reveal the prevalence and epidemiology of extended-spectrum ß-lactamase (ESBL)- and/or plasmid AmpC (pAmpC)- and carbapenemase (CP) producing Enterobacteriaceae and vancomycin-resistant enterococci (VRE) across the Northern Dutch-German border region. Methods: A point-prevalence study on ESBL/pAmpC/CP producing Enterobacteriaceae and VRE was carried out in hospitalized patients in the Northern Netherlands (n = 445, 2012-2013) and Germany (n = 242, 2012). Healthy individuals from the Dutch community (n = 400, 2010-2012) were also screened. In addition, a genome-wide gene-by-gene approach was applied to study the epidemiology of ESBL-Escherichia coli and VRE. Results: A total of 34 isolates from 27 patients (6.1%) admitted to Dutch hospitals were ESBL/pAmpC positive and 29 ESBL-E. coli, three pAmpC-E. coli, one ESBL-Enterobacter cloacae, and one pAmpC-Proteus mirabilis were found. In the German hospital, 18 isolates (16 E. coli and 2 Klebsiella pneumoniae) from 17 patients (7.7%) were ESBL positive. In isolates from the hospitalized patients CTX-M-15 was the most frequently detected ESBL-gene. In the Dutch community, 11 individuals (2.75%) were ESBL/pAmpC positive: 10 ESBL-E. coli (CTX-M-1 being the most prevalent gene) and one pAmpC E. coli. Six Dutch (1.3%) and four German (3.9%) hospitalized patients were colonized with VRE. Genetic relatedness by core genome multi-locus sequence typing (cgMLST) was found between two ESBL-E. coli isolates from Dutch and German cross-border hospitals and between VRE isolates from different hospitals within the same region. Conclusion: The prevalence of ESBL/pAmpC-Enterobacteriaceae was similar in hospitalized patients across the Dutch-German border region, whereas VRE prevalence was slightly higher on the German side. The overall prevalence of the studied pathogens was lower in the community than in hospitals in the Northern Netherlands. Cross-border transmission of ESBL-E. coli and VRE seems unlikely based on cgMLST analysis, however continuous monitoring is necessary to control their spread and stay informed about their epidemiology.

Dynamics of nasopharyngeal pneumococcal carriage during the course of viral bronchiolitis.

The effect of viral infection on nasopharyngeal carriage of Streptococcus pneumoniae during childhood is not well known. We studied dynamics of pneumococcal colonization by quantitative PCR during the natural course of viral bronchiolitis. At time of admission, 47 (47%) of 100 patients with bronchiolitis carried pneumococci. In patients with viral bronchiolitis who did not receive antibiotics, pneumococcal load decreased from time of admission to discharge (n = 35, cycle threshold 23 vs. 25, P = 0.0017) and from discharge to follow-up (n = 22, cycle threshold 25 vs. 40, P = 0.003). We conclude that viral respiratory infection is negatively associated with pneumococcal colonization of the upper airways. Pediatr Pulmonol. 2016;51:863-867. © 2016 Wiley Periodicals, Inc.

Brain death induces renal expression of heme oxygenase-1 and heat shock protein 70.

BACKGROUND: Kidneys derived from brain dead donors have lower graft survival and higher graft-function loss compared to their living donor counterpart. Heat Shock Proteins (HSP) are a large family of stress proteins involved in maintaining cell homeostasis. We studied the role of stress-inducible genes Heme Oxygenase-1 (HO-1), HSP27, HSP40, and HSP70 in the kidney following a 4 hour period of brain death. METHODS: Brain death was induced in rats (n=6) by inflating a balloon catheter in the epidural space. Kidneys were analysed for HSPs using RT-PCR, Western blotting, and immunohistochemistry. RESULTS: RT-PCR data showed a significant increase in gene expression for HO-1 and HSP70 in kidneys of brain dead rats. Western blotting revealed a massive increase in HO-1 protein in brain dead rat kidneys. Immunohistochemistry confirmed these findings, showing extensive HO-1 protein expression in the renal cortical tubules of brain dead rats. HSP70 protein was predominantly increased in renal distal tubules of brain dead rats treated for hypotension. CONCLUSION: Renal stress caused by brain death induces expression of the cytoprotective genes HO-1 and HSP70, but not of HSP27 and HSP40. The upregulation of these cytoprotective genes indicate that renal damage occurs during brain death, and could be part of a protective or recuperative mechanism induced by brain death-associated stress.

Local renal complement C3 induction by donor brain death is associated with reduced renal allograft function after transplantation.

BACKGROUND: Kidneys derived from brain-dead donors have inferior outcomes after transplantation compared to kidneys from living donors. Strikingly, early and profound serum levels of IL-6 in brain-dead donors are observed. IL-6 is the main regulator of the acute phase response (APR). The aim of this translational study was to investigate the expression of renal acute phase proteins (APPs) following brain death (BD) and to assess the association with renal allograft outcome after transplantation. METHODS: BD was induced in rats by inflating a subdurally placed balloon catheter. Kidney biopsies were obtained from human living and brain-dead donors at donation, after cold preservation and reperfusion. In vitro, renal proximal tubular epithelial cells (HK-2 cells) were stimulated with IL-6. RESULTS: Both in human and rat brain-dead donors, C3 and FBG expression was enhanced at donation compared to living donors and sham-operated animals. In human donors, no additional expression was found after cold ischaemia or reperfusion. C3 expression after reperfusion was independently associated with decreased short-term function after transplantation in grafts from brain-dead donors. In cultured HK-2 cells, C3 production was induced in the presence of IL-6. CONCLUSIONS: In conclusion, BD induces renal C3 and FBG expression. Moreover, C3 expression is associated with a worse allograft function early after transplantation. Therefore, targeting renal APPs in brain-dead donors, especially complement C3, may improve transplant outcome.

Reduction of proteinuria in adriamycin-induced nephropathy is associated with reduction of renal kidney injury molecule (Kim-1) over time.

Tubulointerstitial lesions are important in the progression of proteinuric renal disease. Tubular kidney injury molecule-1 (Kim-1) is induced in acute renal injury and reversible as a natural course. Kim-1 is also present in chronic renal damage; however, the dynamics of Kim-1 in chronic renal damage and effects of antiproteinuric treatment on Kim-1 are unknown. We studied Kim-1 in adriamycin nephrosis (AN) before and after renin-angiotensin system blockade. A renal biopsy was taken 6 wk after adriamycin injection to study renal damage and Kim-1 expression. Subsequently, ACE inhibition (ACEi; n = 23), angiotensin II antagonist (AT(1A); n = 23), or vehicle (n = 10) was given for 6 wk; healthy rats served as controls (CON; n = 8). In AN, renal Kim-1 mRNA was induced 26-fold vs. CON at week 6, with further increase in vehicle to week 12 (40-fold) but was reduced by ACEi and AT(1A) to 10- and 12-fold vs. CON (P < 0.05 vs. week 6). Kim-1 protein was undetectable in CON; in AN, it was present in brush border of dilated tubules in areas with adjacent interstitial lesions. Renal Kim-1 protein levels increased from weeks 6-12 in vehicle and decreased in ACEi- and AT(1A)-treated groups (P < 0.05). In vehicle, urinary Kim-1 was increased (P < 0.05 vs. CON), with a reduction by ACEi and AT(1A) (P < 0.05 vs. vehicle). Renal and urinary Kim-1 correlated with proteinuria and interstitial damage cross-sectionally. Reductions in proteinuria and renal Kim-1 correlated, which was not associated by corresponding changes in tubulointerstitial fibrosis. In conclusion, on longitudinal follow-up during antiproteinuric treatment increased renal Kim-1 expression is reversible in proportion to proteinuria reduction, likely reflecting reversibility of early tubular injury, supporting its potential as a biomarker for tubulointerstitial processes of damage and repair.

The role of bile salt toxicity in the pathogenesis of bile duct injury after non-heart-beating porcine liver transplantation.

BACKGROUND: Intrahepatic bile duct strictures are a serious complication after non-heart-beating (NHB) liver transplantation. Bile salt toxicity has been identified as an important factor in the pathogenesis of bile duct injury and cholangiopathies. The role of bile salt toxicity in the development of biliary strictures after NHB liver transplantation is unclear. METHODS: In a porcine model of NHB liver transplantation, we studied the effect of different periods of warm ischemia in the donor on bile composition and subsequent bile duct injury after transplantation. After induction of cardiac arrest in the donor, liver procurement was delayed for 0 min (group A), 15 min (group B), or more or equal to 30 min (group C). Livers were subsequently transplanted after 4 hr of cold preservation. In the recipients, bile flow was measured, and bile samples were collected daily to determine the bile salt-to-phospholipid ratio. Severity of bile duct injury was semiquantified by using a histologic grading scale. RESULTS: Posttransplantation survival was directly related to the duration of warm ischemia in the donor. The bile salt-to-phospholipid ratio in bile produced early after transplantation was significantly higher in group C, compared with group A and B. Histopathologic condition showed the highest degree of bile duct injury in group C. CONCLUSION: Prolonged warm ischemia in NHB donors is associated with the formation of toxic bile after transplantation, with a high biliary bile salt-to-phospholipid ratio. These data suggest that bile salt toxicity contributes to the pathogenesis of bile duct injury after NHB liver transplantation.

Complement and renal transplantation: from donor to recipient.

Long-term kidney graft survival is affected by different variables including donor condition, ischemia-reperfusion injury, and graft rejection during the transplantation process. The complement system is an important mediator of renal ischemia-reperfusion injury and in rejecting allografts. However, donor complement C3 seems to be crucial in renal transplantation-related injury as renal injury is attenuated in C3 deficient kidney grafts. Interestingly, before ischemia-reperfusion induced C3 expression, C3 is already induced in donors suffering from brain death. Therefore, strategies targeting complement activation in the brain-dead donor may increase graft viability and transplant outcome.

Early events in kidney donation: progression of endothelial activation, oxidative stress and tubular injury after brain death.

Cerebral injury leading to brain death (BD) causes major physiologic derangements in potential organ donors, which may result in vascular-endothelial activation and affect posttransplant graft function. We investigated the kinetic of pro-coagulatory and pro-inflammatory endothelial activation and the subsequent oxidative stress and renal tubular injury, early after BD declaration. BD was induced by slowly inflating a balloon-catheter inserted in the extradural space over a period of 30 min. Rats (n = 30) were sacrificed 0.5, 1, 2 or 4 h after BD-induction and compared with sham-controls. This study demonstrates immediate pro-coagulatory and pro-inflammatory activation of vascular endothelium after BD in kidney donor rats, proportional with the duration of BD. E- and P-Selectins, Aalpha/Bbeta-fibrinogen mRNA were abruptly and progressively up-regulated from 0.5 h BD onwards; P-Selectin membrane protein expression was increased; fibrinogen was primarily visualized in the peritubular capillaries. Plasma von Willebrand factor was significantly higher after 2 h and 4 h BD. Urine heart-fatty-acid-binding-protein and N-acetyl-glucosaminidase, used as new specific and sensitive markers of proximal and distal tubular damage, were found significantly increased after 0.5 h, with a maximum at 4 h. Unexpectedly, oxidative stress was detectable only late, after the installation of tubular injury, suggesting only a secondary role for hypoxia in triggering these injuries.

Static cold storage preservation of ischemically damaged kidneys. a comparison between IGL-1 and UW solution.

Especially in damaged organs, adequate organ preservation is critically important to maintain viability. Institut Georges Lopez-1 (IGL-1) is a new preservation solution, with an extracellular sodium/potassium ratio and polyethylene glycol as a colloid. The influence of warm and cold ischemia was evaluated in a rat Lewis-Lewis transplant model with a follow up of 14 days. Eight groups of donation after cardiac death donor kidneys were studied with warm ischemia of 0 and 15 min followed by 0- or 24-h cold storage (CS) preservation in IGL-1 or UW-CSS. Blood was collected daily during the first week and at day 14. Recipients were placed in metabolic cages at day 4 and 14 after transplantation allowing urine collection and adequate measurement of glomerular filtration rate. Focussing on inflammation, reactive oxygen species production, proximal tubule damage, proteinuria, histology, and renal function after transplantation we could not show any relevant difference between IGL-1 and UW-CSS. Furthermore, the combination of 15-min warm ischemia and by 24-h cold ischemia did not result in life sustaining kidney function after transplantation, irrespective of the used solution. In the present experiment, static CS preservation of ischemically damaged rat kidneys in either IGL-1 or UW-CSS rendered equal results after transplantation.

Induction of kidney injury molecule-1 in homozygous Ren2 rats is attenuated by blockade of the renin-angiotensin system or p38 MAP kinase.

Kidney injury molecule-1 (Kim-1) is associated with ischemic and proteinuric tubular injury; however, whether dysregulation of the renin-angiotensin system (RAS) can also induce Kim-1 is unknown. We studied Kim-1 expression in homozygous Ren2 rats, characterized by renal damage through excessive RAS activation. We also investigated whether antifibrotic treatment (RAS blockade or p38 MAP kinase inhibition) would affect Kim-1 expression. At 7 wk of age, homozygous Ren2 rats received a nonhypotensive dose of candesartan (0.05 mg x kg(-1) x day(-1) sc) or the p38 inhibitor SB-239063 (15 mg x kg(-1) x day(-1) sc) for 4 wk; untreated Ren2 and Sprague-Dawley (SD) rats served as controls. Kim-1 mRNA and protein expression were determined by quantitative PCR and immunohistochemistry, respectively, and related to markers of prefibrotic renal damage. Urinary Kim-1 was measured in 8-wk-old Ren2 and SD rats with and without angiotensin-converting enzyme inhibition (ramipril, 1 mg x kg(-1) x day(-1) in drinking water for 4 wk). Untreated Ren2 rats showed a >20-fold increase in renal Kim-1 mRNA (expressed as Kim-1-to-GAPDH ratio): 75.5 +/- 43.6 vs. 3.1 +/- 1.0 in SD rats (P < 0.01). Candesartan and SB-239063 strongly reduced Kim-1 mRNA: 3.1 +/- 1.5 (P < 0.01) and 9.8 +/- 4.2 (P < 0.05), respectively. Kim-1 protein expression in damaged tubules paralleled mRNA expression. Kim-1 expression correlated with renal osteopontin, alpha-smooth muscle actin, and collagen III expression and with tubulointerstitial fibrosis. Damaged tubular segments expressing activated p38 also expressed Kim-1. Urinary Kim-1 was increased in Ren2 vs. SD (458 +/- 70 vs. 27 +/- 2 pg/ml, P < 0.01) rats and abolished in Ren2 rats treated with ramipril (33 +/- 5 pg/ml, P < 0.01). Kim-1 is associated with development of RAS-mediated renal damage. Antifibrotic treatment through RAS blockade or p38 MAP kinase inhibition reduced Kim-1 in the homozygous Ren2 model.

Spatiotemporal expression of heme oxygenase-1 detected by in vivo bioluminescence after hepatic ischemia in HO-1/Luc mice.

Upregulation of heme oxygenase-1 (HO-1) has been proposed as a critical mechanism protecting against cellular stress during liver transplantation, providing a potential target for new therapeutic interventions. We investigated the feasibility of in vivo bioluminescence imaging (BLI) to noninvasively quantify the spatiotemporal expression of HO-1 after warm hepatic ischemia in living animals. Luciferase activity was measured by BLI as an index of HO-1 transcription in transgenic reporter mice (Ho1-luc) at standardized time points after 60 minutes of warm hepatic ischemia. HO-1 mRNA levels were measured in postischemic livers of mice sacrificed at the same time points in separate experiments. Bioluminescent signals from postischemic liver lobes were first detected at 3 hours after reperfusion. Peak levels were reached at 9 hours, after which bioluminescent activity declined and returned to baseline values at 48 hours after reperfusion. Upregulation of HO-1 as detected by in vivo BLI was preceded by increased HO-1 mRNA expression and confirmed by enhanced immunohistochemical staining of hepatocytes. In conclusion, this study shows that in vivo BLI allows a sensitive assessment of HO-1 expression after hepatic ischemia in living animals. The capability of whole-body temporal imaging of HO-1 expression provides a valuable tool in the development of novel strategies to modulate HO-1 expression in liver transplantation.

Matrix metalloproteinases as profibrotic factors in terminal ileum in Crohn's disease.

BACKGROUND: Returning stenosis in Crohn's disease (CD) patients is poorly understood. After resection, newly developed strictures are seen within 10 years in 50% to 70%. Matrix metalloproteinases (MMPs) are involved in matrix-turnover processes. This study analyzes spatial expression of MMP-1, MMP-3, MMP-9, tissue inhibitor of MMP-1, and collagen III to get better insight in tissue remodeling of terminal ileum of CD patients. METHODS: Expressions were analyzed on mRNA and the protein level (MMP-1, MMP-3) in segments from resected terminal ileum from CD and control patients. In CD, macroscopic distinction was made between proximal resection margin, prestenotic, and stenotic tissue. Immunohistochemistry allowed for expression analyses transmurally. RESULTS: MMP-1 and MMP-3 gene expression was up-regulated (P < 0.05) in both prestenotic and stenotic tissue. MMP-1 protein was significantly up-regulated in submucosal and muscular tissue of prestenotic parts and in muscular tissue of stenotic Crohn samples. MMP-3 protein was significantly up-regulated in all layers of prestenotic and stenotic Crohn samples. Even in submucosa of proximal resection margin tissue, MMP-3 expression was significantly higher than in controls. CONCLUSION: Surprisingly, in proximal resection margin tissue up-regulated MMP-3 was seen. This suggests that in nonresected terminal ileum, in which anastomosis is made, tissue turnover is present, which may account for the high recurrence of intestinal strictures.

Tubular kidney injury molecule-1 in protein-overload nephropathy.

Kim-1, a recently discovered membrane protein, is undetectable in normal kidneys but markedly induced in proximal tubules after ischemic and toxic injury. The function of Kim-1 is unclear, but it is implicated in damage/repair processes. The Kim-1 ectodomain is cleaved by metalloproteinases and detectable in urine. We studied Kim-1 in a nontoxic, nonischemic, model of tubulointerstitial damage caused by acute proteinuria. Uninephrectomized (NX) rats received daily (ip) injections of 2 g BSA (NX+BSA, n = 12) or saline (NX, n = 6) for 3 wk. Kidneys were stained for various damage markers by immunohistochemistry (IHC). Kim-1 mRNA (RT-PCR, in situ hybridization), protein (IHC, Western blotting), and urinary Kim-1 (Luminex) were determined. Spatial relations between Kim-1 and other damage markers were studied by double labeling IHC. NX+BSA rats developed massive proteinuria (1,217 +/- 313 vs. 18 +/- 2 mg/day in NX, P < 0.001) and significant renal damage. Kim-1 mRNA was upregulated eightfold in NX+BSA (ratio Kim-1/beta-actin, 4.08 +/- 2.56 vs. 0.52 +/- 0.64 in NX, P < 0.001) and localized to damaged tubules. Kim-1 protein expression was markedly induced in NX+BSA (2.46 +/- 1.19 vs. 0.39 +/- 0.10% staining/field in NX, P < 0.001). Urinary Kim-1 was significantly elevated in NX+BSA (921 +/- 592 vs. 87 +/- 164 pg/ml in NX, P < 0.001) and correlated with tissue Kim-1 expression (r = 0.66, P =0.02). Kim-1 protein was found at the apical membrane of dilated nephrons. Kim-1 expression was limited to areas with inflammation (MØ), fibrosis (alpha-smooth muscle actin), and tubular damage (osteopontin), and only occasionally with tubular dedifferentiation (vimentin). These results implicate involvement of Kim-1 in the pathogenesis of proteinuria-induced renal damage/repair. Urinary Kim-1 levels may serve as a marker of proteinuria-induced renal damage.

Distinct transcriptional changes in donor kidneys upon brain death induction in rats: insights in the processes of brain death.

Brain death affects hormone regulation, inflammatory reactivity and hemodynamic stability. In transplant models, donor organs retrieved from brain dead (BD) rats suffer from increased rates of primary non-function and lower graft survival. To unravel the mechanisms behind brain death we have performed DNA microarray studies with kidney-derived RNA from normo- and hypotensive BD rats, corresponding with optimal and marginal BD donors, respectively. In kidneys from normotensive donors 63 genes were identified as either up- (55) or down-regulated (8), while 90 genes were differentially expressed (67 up-regulated) in hypotensive BD donor kidneys. Most genes were categorized in different functional groups: metabolism/transport (including the down-regulated water channel Aqp-2), inflammation/coagulation (containing the largest number (16) of up-regulated genes including selectins, Il-6, alpha- and beta-fibrinogen), cell division/fibrosis (including KIM-1 involved in tubular regeneration) and defense/repair (with the cytoprotective genes HO-1, Hsp70, MnSOD2). Also, genes encoding transcription factors (including immediate early genes as Atf-3, Egr-1) and proteins involved in signal transduction (Pik3r1) were identified. Summarizing, the use of DNA microarrays has clarified parts of the process of brain death: Brain-death-induced effects ultimately lead, via activation of transcription factors and signal transduction cascades, to differential expression of different "effector" genes. Not only deleterious processes such as inflammation and fibrosis occur in brain dead donor kidneys but genes involved in protection and early repair processes are activated as well. These findings can be used to introduce specific cytoprotective interventions in the brain dead donor to better maintain or even increase organ viability.

Effect of brain death on gene expression and tissue activation in human donor kidneys.

BACKGROUND: After kidney transplantation, decreased graft survival is seen in grafts from brain dead (BD) donors compared with living donors. This might result partly from a progressive nonspecific inflammation in the graft. In this study, we focused on the effects of BD on inflammatory response (adhesion molecules, leukocyte invasion, gene expression) and stress-related heat shock proteins in the human kidney. Research outcomes and clinical donor parameters were then linked to outcome data after transplantation. METHODS: Kidney biopsy specimens and serum were obtained during organ retrieval from BD and living organ donor controls. Immunohistochemistry and semiquantitative reverse transcriptase-polymerase chain reaction were performed on the biopsy specimens. Clinical and laboratory parameters from BD donors were recorded and connected to outcome data of the recipients of the kidneys studied. RESULTS: After brain death, immunohistochemistry showed an increase of E-selectin (P<0.01) and interstitial leukocyte invasion (P<0.05) compared with controls. Also, reverse transcriptase-polymerase chain reaction showed a threefold increased heme oxygenase-1 (P<0.05) and Hsp70 (P<0.01) gene expression after BD. Levels of monocyte chemotactic protein-1 and transforming growth factor-beta were twice as high after brain death but did not reach significance. Transplantation outcome was influenced by several donor variables: positively most notably by donor treatment with desmopressin and negatively by high serum urea levels during brain death and by high intercellular adhesion molecule and vascular cell adhesion molecule expression in the kidney. Heme oxygenase-1 proved to have a protective function, but only in kidneys from living donors. CONCLUSIONS: The presence of interstitial leukocytes and the early adhesion molecule E-selectin in BD donor kidneys indicates an early-phase inflammatory process during organ retrieval. Elevated levels of monocyte chemotactic protein-1 and transforming growth factor-beta suggest a role for monocytes/macrophages in this phase. We suggest that BD causes a stress-related response against which protective heat shock proteins are formed in the future graft. This stress response may be too severe to be fully counteracted by elevated heat shock proteins. Which systemic and/or local factors trigger brain death-related graft injury is currently under investigation.

Causes of limited survival of microencapsulated pancreatic islet grafts.

Successful transplantation of pancreatic tissue has been demonstrated to be an efficacious method of restoring glycemic control in type 1 diabetic patients. To establish graft acceptance patients require lifelong immunosuppression, which in turn is associated with severe deleterious side effects. Microencapsulation is a technique that enables the transplantation of pancreatic islets in the absence of immunosuppression by protecting the islet tissue through a mechanical barrier. This protection may even allow for the transplantation of animal tissue, which opens the perspective of using animal donors as a means to solve the problem of organ shortage. Microencapsulation is not yet applied in clinical practice, mainly because encapsulated islet graft survival is limited. In the present review we discuss the principal causes of microencapsulated islet graft failure, which are related to a lack of biocompatibility, limited immunoprotective properties, and hypoxia. Next to the causes of encapsulated islet graft failure we discuss possible improvements in the encapsulation technique and additional methods that could prolong encapsulated islet graft survival. Strategies that may well support encapsulated islet grafts include co-encapsulation of islets with Sertoli cells, the genetic modification of islet cells, the creation of an artificial implantation site, and the use of alternative donor sources. We conclude that encapsulation in combination with one or more of these additional strategies may well lead to a simple and safe transplantation therapy as a cure for diabetes.