| 1. |
- Breimer, Michael, 1951, et al.
(författare)
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Extracorporeal ("ex vivo") connection of pig kidneys to humans. I. Clinical data and studies of platelet destruction.
- 1996
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Ingår i: Xenotransplantation. - : Wiley. - 0908-665X .- 1399-3089. ; 3:4, s. 328-39
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Tidskriftsartikel (refereegranskat)abstract
- The pioneering experiment by Welsh et al. (Immunological Lett 1991:29:167-170) connecting a pig kidney to the human circulation has been repeated in a modified manner. Two volunteer dialysis patients were pretreated by daily plasmapheresis on days -2,-1, and 0 to remove the naturally occurring anti-pig xenoantibodies. The anti-pig lymphocytotoxic liters were reduced from 1:8 to 1:2 in patient 1 and from 1:8 to 1:1 in patient 2. No steroids or immunosuppressive drugs were administrated before or during the experiments. A sterile pig kidney was extracorporeally ("ex vivo") connected to the patients a/v fistula using an arterial and a venous pump similar to a dialysis. The two experiments gave different results. In the first experiment the perfusion pressure was kept at 100 mmHg for the initial 25 min by reducing the pump speed until the minimum blood flow of 30 ml/min was reached. Thereafter, the pressure rose continuously and the experiment was terminated at 65 min at a perfusion pressure of 200 mmHg. The patient did not feel any discomfort during the perfusion. In the second experiment, a stable blood flow of 200 ml/min was reached at a pressure of 100 mmHg after a few minutes. The perfusion was terminated at 15 min when the patient developed chest and abdominal pain, hypotension, and electrocardiographic signs of myocardial ischemia. The patient recovered quickly. In the first experiment, small volumes of clear urine was produced until the pressure rose above 100 mmHg, which resulted in hematuria. In the second experiment clear urine (4 ml/min) was produced. (51)Chromium clearance values were after 15 min <1 ml/min for kidney 1 and 12 ml/min (8 ml/min/100 g) for kidney 2. A drastic reduction in platelet count (128 to 48 and 64 to 8 × 10(9)/1, respectively) during the passage through the kidney was found in blood samples collected simultaneously before and after the organ. No change in hemoglobin values and leucocyte counts were found. Light- and electron-microscopical analysis of the kidney tissues revealed for kidney 1 focal areas with obliteration of the glomerular and peritubular capillaries by platelets and PMN cells and severe damage of the endothelial cells comparable to a picture of a hyperacute rejection. In kidney 2, all vessels were patent but in the capillaries large amount of membrane fragments were detected by electron microscopy and a discrete damage of the endothelial cells were seen in some segments. No intact platelets were present in the vascular tree. These human experiments support the hypothesis that hyperacute rejection of pig to human xenografts is delayed in time by removal of the preformed anti-pig xenoantibodies. A new finding was a very rapid destruction of platelets occurring in the kidney of patient 2 who had very low liters of xenoantibodies. The humoral immune response is described in detail in an accompanying paper (Rydberg et al., this issue).
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| 2. |
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| 3. |
- Bengtsson, Anders, 1954, et al.
(författare)
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Extracorporeal ("ex vivo") connection of pig kidneys to humans. III. Studies of plasma complement activation and complement deposition in the kidney tissue.
- 1998
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Ingår i: Xenotransplantation. - 0908-665X. ; 5:3, s. 176-83
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Tidskriftsartikel (refereegranskat)abstract
- The complement system is one of the important factors involved in the hyperacute rejection of xenografts. This report deals with the activation of the complement system in a clinical trial where pig kidneys were extracorporeally connected to two volunteer dialysis patients who were pretreated with plasmapheresis in order to substantially reduce anti-pig xenoantibodies. The clinical data of the perfusion experiments and the patients humoral immune response to pig xenoantigens have been reported in detail (Xenotransplantation 1996; 3:328-339, 340-353). Three consecutive daily plasmapheresis treatments of the patients reduced the plasma complement protein (C3, C4, and C5) concentrations to 8-27% of the baseline values. The perfusion of the pig kidney connected to patient 1 was terminated at 65 min due to graft rejection and this patient was not hemodynamically affected by the experiment. The second experiment was terminated at 15 min due to an anaphylactic like reaction of the patient. In patient 1 a slight reduction of plasma C3, C4, and C5 and an increase of C5a and SC5b-9 occurred, while C3a decreased during the perfusion. Patient 2 had an increase of all complement parameters, most prominent for C4d and SC5b-9, which occurred concomitant with the appearance of the anaphylactic like side effects. In general, plasma levels of PMN elastase, IL6 and IL8 increased in both patients during the perfusion. Immunohistochemical investigation of the kidney tissues revealed deposition of human complement factors C1q, C4c, and C3c in a congruent pattern with the vasculature of the kidney in patient 1. In kidney 2 only trace amounts of C1q and C3c were found. Both kidneys were negative for properdin. Therefore, in this experimental set up with extracorporeal connection of pig kidneys to the human circulation the human complement cascade is activated mainly through the classical pathway.
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| 4. |
- Breimer, Michael, 1951, et al.
(författare)
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Blood group A and B antigen expression in human kidneys correlated to A1/A2/B, Lewis, and secretor status.
- 2006
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Ingår i: Transplantation. - : Ovid Technologies (Wolters Kluwer Health). - 0041-1337. ; 82:4, s. 479-85
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: In the revived interest in crossing ABO barriers in organ transplantation renal A/B antigen expression has been correlated with donor ABO, Lewis, and secretor subtype to predict antigen expression. METHODS: A/B antigen expression was explored by immunohistochemistry in LD renal biopsies. Donor A1/A2/B, Lewis, and secretor status were determined by serology and polymerase chain reaction. RESULTS: In the renal vascular bed, three distinct A antigen expression patterns with a major, minor, and minimal staining distribution, and intensity (designated as types 3+, 1+ and (+) respectively) were identified. Type 3+ had a strong A antigen expression in the endothelium of arteries, glomerular/peritubular capillaries and veins. The type 1+ showed an overall weaker antigen expression, whereas type (+) had faint staining of peritubular capillaries only. In all cases, distal tubular epithelium was focally stained, whereas proximal tubules were negative. Type 3+ were all from blood group A1 subtype individuals while A2 cases expressed either a 1+ or (+) pattern. The secretor gene did not appear to influence renal A antigen expression. All B kidneys examined showed a B antigen pattern slightly weaker but otherwise similar to A type 3+. CONCLUSION: Renal vascular A antigen expression correlates to donor A1/A2 subtypes, whereas B individuals show one singular antigen pattern. From antigen perspective, A1 and B donors are a "major" and A2 individuals a "minor" antigen challenge in ABO-incompatible renal transplantation.
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| 5. |
- Breimer, Michael, 1951, et al.
(författare)
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Physiological and histological characterisation of a pig kidney in vitro perfusion model for xenotransplantation studies.
- 1996
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Ingår i: Scandinavian journal of urology and nephrology. - 0036-5599. ; 30:3, s. 213-21
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Tidskriftsartikel (refereegranskat)abstract
- A pig kidney perfusion model aimed for use in immunological and physiological xenotransplantation research has been developed. Organ viability was characterised by clearance studies, functional response to hormones/diureticum and by light microscopical examination. The pig kidney was perfused in a specially designed plexiglass chamber, using a roller pump and a small membrane oxygenator (O2/CO2, 95/5). The recirculating perfusate used was autologous pig blood diluted by Tyrodes solution to a hematocrit of 30%, at a total starting volume of 600-650 ml. The temperature was 37 degrees C. It was crucial for good organ function that the nephrectomy operating time, as well as the warm (1-2 min) and cold ischemia (average 43 min) times were minimized. The average total perfusion time was 151 minutes. Physiological parameters were measured during 10-15 minute periods at average times of 40, 63, 88 and 142 minutes. The clearance values of inulin in these periods were 54 +/- 13, 59 +/- 15, 48 +/- 23, 27 +/- 5 and for PAH; 103 +/- 14, 121 +/- 14, 106 +/- 30, 114 +/- 34 ml/min/100 g tissue weight. The plasma flows were 123 +/- 12, 155 +/- 17, 136 +/- 36 and 206 +/- 57 ml/min/100 g. The injection of 0.5 micrograms of alpha ANP to the perfusate resulted in a significant decrease in vascular resistance, and increase in urine production (+107%), as well as sodium (+112%) and potassium (+46%) excretion. Ten mg furosemide doubled the urine production and sodium excretion, while potassium excretion increased marginally. The number of leucocytes decreased by 39% during the perfusion, while the platelet count was unaffected. Light microscopy of the renal tissue after termination of the experiments revealed endothelial damage to variable extent. Loss of endothelial cells was most obvious at the level of arcuate and interlobular arteries, while the endothelium was intact in larger arteries and veins. Accumulation of polymorphonuclear granulocytes was found predominantly in the peritubular vessels, and to a lesser degree in the cortical venules. In the tubular cells, only minimal epithelial swelling and irregular cytoplasmic vacuolisation was found. Thus, a good functional viability can be maintained during 2 hours in vitro perfusion, although a decline in function as well as structural damage can be seen at the end of the experiment.
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| 6. |
- Freese, P M, et al.
(författare)
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Renal allograft glomerulopathy and the value of immunohistochemistry.
- 2004
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Ingår i: Clinical nephrology. - 0301-0430. ; 62:4, s. 279-86
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Tidskriftsartikel (refereegranskat)abstract
- Studies of late renal allograft biopsies focus on chronic damage investigated by light microscopy (LM). We evaluated the use of immunohistochemistry (IH) as applied in the routine study of transplant glomerulopathies. Among renal transplants in 1985 - 1997, 129 were identified where a graft biopsy had been obtained 6 months or more after transplantation, studied by LM and IH and the original renal disease was known. IH results were evaluated in relation to glomerular LM findings and the original diagnosis. The risk of graft loss in relation to recurrent and de novo glomerulopathy was evaluated. By LM, 69 biopsies (53%) showed glomerulopathy, mesangial sclerosis only in 26, proliferative changes in 15, membranous in 15 and combined membranous and proliferative in 13. By IH, 46 biopsies (36%) stained positive with IgM and/or complement only and 24 with immune complexes including IgA and/or IgG. Seven biopsies (5.4%) showed glomerular disease by IH in spite of normal LM. Recurrence was diagnosed in 22 grafts; 12 had IgA nephropathy, 3 had SLE, 6 other immune complex nephritides and 1 systemic vasculitis. Twenty-eight biopsies (22%) with proliferative and/or membranous glomerulopathy lacked clear connection to the original renal disorder. More than half of these had deposits of IgM and C3 only. The further graft survival was significantly reduced in the presence of de novo glomerulopathy by LM, relative risk 2.0 (confidence interval 1.1 - 3.8) in a Cox-proportional hazards analysis also including serum creatinine and Banff chronic allograft nephropathy (CAN) grade, p = 0.03. In conclusion, transplant glomerulopathy should be separated from recurrence. De novo glomerulopathy is frequent and ominous.
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| 7. |
- Mölne, Johan, 1958, et al.
(författare)
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Glomerular C3c deposition and intravascular macrophage accumulation in early humoral renal allograft rejection signifies a poor short-term outcome.
- 2006
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Ingår i: APMIS : acta pathologica, microbiologica, et immunologica Scandinavica. - : Wiley. - 0903-4641. ; 114:10, s. 700-11
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Tidskriftsartikel (refereegranskat)abstract
- C4d deposition in the walls of peritubular capillaries is considered the key phenomenon in the histopathological diagnosis of humoral, i.e. antibody-mediated, allograft rejection. We have earlier proposed that deposition of C3c in glomerular capillaries and simultaneous intravascular accumulation of macrophages in allografts with immediate or early humoral rejection indicates a potentially serious condition with very poor prognosis. The clinical outcome of 45 cadaveric grafts with this phenomenon among 1960 renal allografts transplanted at our centre during 1984-1999, and the recipients of the contralateral kidneys, was retrospectively evaluated. Graft failure occurred in 44/45 grafts within 3 weeks, with graft loss in 33/45 (77%) within 4 months and 37/45 (82%) within 1 year. From the contralateral kidneys, 5/33 (15%) were lost within 1 year. In a recent series of early biopsies, we recognised that of 13 cases showing C4d positivity in peritubular capillaries but lacking C3c in glomeruli, 10/13 (77%) were still functioning after 4 months. The mean number of CD68(+) macrophages per glomerular profile, i.e. the glomerular macrophage index, shows a significant difference between C4d(+)C3c(-) and C4d(+)C3c(+) cases (p<0.001). Our results indicate the existence of a clinically important subgroup of early humoral rejection with particular morphological features.
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| 8. |
- Mölne, Johan, 1958, et al.
(författare)
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Immunoperoxidase versus immunofluorescence in the assessment of human renal biopsies.
- 2005
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Ingår i: American journal of kidney diseases : the official journal of the National Kidney Foundation. - 1523-6838. ; 45:4, s. 674-83
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: For half a century, immunofluorescence (IF) on frozen sections has been the gold standard for immunohistochemical evaluation of renal biopsy specimens. In routine diagnostic immunohistopathologic evaluation, traditional IF has been replaced to a large extent by immunoperoxidase (IP) methods applied to paraffin sections of formaldehyde-fixed tissue. This is caused in part by the practical disadvantages inherent in the IF method, eg, separate tissue specimen and handling, UV microscopy, fading and impermanence of the label-making archiving, and difficult later investigation. Our aim for the present study is to evaluate IP as an alternative to IF in the diagnostic assessment of renal biopsy specimens. METHODS: Proteolytic antigen retrieval, antibodies effective on deparaffinized sections, a sensitive detection system (Dako EnVision HRP; Dako, Copenhagen, Denmark), and a standardized and rigorously controlled procedure were applied to a series of renal biopsy specimens (n = 81) previously classified by means of light microscopy (LM) and IF. Staining for immunoglobulin G (IgG), IgA, IgM, C1q, and C3c were recorded as positive or negative for IF and IP in paired proportions, presuming that IF was the test standard. RESULTS: Concordant observations were 71% for all (282 of 398 observations), 82% for IgG (65 of 79 observations), and 89% for IgA (72 of 81 observations). The majority of discordant observations (74 of 116 observations) were positive by means of IP, with mesangial deposits of IgM and C1q that were not found by IF. Statistically, there was no significant difference in outcomes between IF and IP for IgG, IgA, and C3c ( P > 0.2). In addition, IP staining allowed simultaneous evaluation of tissue by LM and therefore correlation between tissue structure and immune deposits not readily attained by IF. CONCLUSION: In the present study, it is documented that for the detection of IgG, IgA, and C3c, IP applied to protease-digested deparaffinized sections of formaldehyde-fixed renal tissue is, with few exceptions, equal to IF on frozen sections. The EnVision HRP method used here is several times more effective in terms of primary antibody dilution than earlier existing IP methods, and because the avidin-biotin system is not involved, very little nonspecific background staining will occur. Discordant observations (116 of 398 observations; 29%) were in the majority (91 of 116 observations) due to positive IP findings of IgM and C1q, which deserve additional investigation.
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| 9. |
- Nordén, Gunnela, 1945, et al.
(författare)
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ABO-incompatible live donor renal transplantation using blood group A/B carbohydrate antigen immunoadsorption and anti-CD20 antibody treatment.
- 2006
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Ingår i: Xenotransplantation. - : Wiley. - 0908-665X .- 1399-3089. ; 13:2, s. 148-53
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Blood group ABO-incompatible live donor (LD) renal transplantation may provide a significant source of organs. We report the results of our first 14 cases of ABO-incompatible LD renal transplantation using specific anti-A/B antibody (Ab) immunoadsorption (IA) and anti-CD20 monoclonal Ab (mAb) treatment. PATIENTS AND TREATMENT PROTOCOL: Recipients were blood group O (n = 12), A (n = 1) and B (n = 1). Donors were A1 (n = 2), A2 (n = 3), A2B (n = 1) and B (n = 8), and all were secretor positive. Anti-human leukocyte antigen (HLA) Ab panel reactivity was negative in all recipients except one. All recipients were pre-treated with 3 to 6 IA sessions, using A or B carbohydrate antigen columns, until their anti-A1/B RBC panel indirect antiglobulin test (IAT) titers were < or =8. CDC crossmatch was negative in all cases. Recipients received preoperative mycophenolic acid, and steroids/tacrolimus were started at transplantation. No splenectomy was performed. Eight recipients received one dose of anti-CD20 mAb (rituximab, 375 mg/m2) pre-operatively and 11 recipients had postoperative protocol IA. RESULTS: In the initial protocol, anti-CD20 mAbs were used only for recipients receiving A1 grafts. One B graft (HLA-identical donor, 84% panel reactivity) was lost in a severe anti-B Ab-mediated acute rejection. Subsequently, the protocol included anti-CD20 for recipients of both A1 and B grafts and postoperative protocol IA to all recipients. The subsequent 10 grafts had excellent function, giving a total graft survival of 13/14 (observation range 2 to 41 months). At 1 yr, mean serum creatinine was 113 micromol/l (n = 8) and mean glomerular filtration rate was 55 ml/min/1.73 m2 (range 24 to 77). In the remaining five cases, with less than 1 yr follow up, mean serum creatinine was 145 micromol/l at 2 to 9 months follow up. Pre-IA anti-A/B titers were in the range of 2 to 32 (NaCl technique) and 16 to 512 (IAT). More than 90 IA sessions were performed in 14 recipients without any significant side effects. Recipient anti-A/B titers returned after transplantation to pre-IA levels or slightly lower. Postoperative renal biopsies were performed in 10 patients. In the 13 patients with long-term function, one patient experienced cellular rejection (Banff IIB) at 3 months without anti-B titer rise. This rejection was concomitant with low tacrolimus plasma levels and was easily reversed by steroids. In 8 of 10 cases, C4d staining was positive in peritubular capillaries. CONCLUSION: Blood group ABO-incompatible LD renal transplantation using A and B carbohydrate-specific IA and anti-CD20 mAbs has excellent graft survival and function.
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| 10. |
- Rydberg, Lennart, 1944, et al.
(författare)
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Characterization of the humoral immune response in two paediatric patients transplanted with split livers from ABO-incompatible living-related donors: appearance of cytomegalovirus-induced ABO antibodies
- 2005
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Ingår i: Transfus Med. - 0958-7578. ; 15:2, s. 137-44
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Tidskriftsartikel (refereegranskat)abstract
- Two blood group O paediatric patients, 12 and 6 months old, were transplanted with liver segments from their blood group A2Le (a(-)b+) Se and blood group A1Le (a(-)b+) Se fathers, respectively. Recipient anti-A antibody titres were reduced prior to transplantation by blood exchange. Both patients had rejection episodes in the post-transplant period that were reversed by anti-rejection therapy. No anti-A antibody titre rise occurred concomitant with these rejections. Postoperatively both patients had cytomegalovirus (CMV) infections, and simultaneous with these infections, a strong increase in anti-A antibody titres was seen, but no rejection occurred. The anti-A antibody titre increase seemed to be specific for A antigens, because the anti-B and anti-alphaGal (anti-pig) antibody titres did not show any changes. CMV infection is a serious cause of morbidity and mortality in immunosuppressed patients, and the virus can influence glycosylation of infected cells. Whether this can explain the importance of the infection in relation to the increase in titre remains to be elucidated.
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