The contribution of podocytes to chronic allograft nephropathy.

BACKGROUND: Progressive proteinuria and glomerulosclerosis characterize chronic allograft nephropathy. However, the causes are not fully elucidated. Podocytes function to prevent proteinuria; injury to this glomerular cell leads to glomerulosclerosis. The potential role of podocytes in the failing transplanted kidney is unknown. A rat model of kidney transplantation, characterized by proteinuria and glomerulosclerosis, was utilized to examine the potential role of podocytes. METHODS: Archival tissue was examined from allografts (Dark Agouti kidneys transplanted into operationally tolerant Albino Surgery rats), isografts (Dark Agouti) and controls (Dark Agouti: age-matched or after unilateral nephrectomy). The number of podocytes (by WT-1 staining) as well as the podocyte proteins podocin, nephrin and synaptopodin (by immunostaining) were measured at days 0, 2, 6 and at 6 months after transplantation. Changes in these parameters were compared between groups and correlated with urinary protein excretion. RESULTS: At 6 months, podocyte number was reduced in allografted kidneys, accompanied by a decrease in nephrin and synaptopodin, but not podocin staining. Remnant kidneys in the uninephrectomized rats also showed a decreased podocyte number but no change in podocyte protein staining. Podocyte loss in allografts was established on day 6, whereas a decrease in nephrin and synaptopodin was not evident until 6 months. In contrast, podocyte number and protein staining was decreased but not significantly so in remnant and isografted kidneys. CONCLUSION: A decrease in the slit diaphragm proteins, nephrin and synaptopodin, is a component of chronic allograft pathology.

Beyond operational tolerance: effect of ischemic injury on development of chronic damage in renal grafts.

BACKGROUND: The induction of operational tolerance is the holy grail of clinical transplantation. However, in animal models with operational tolerance, long- term grafts still develop chronic damage. The elucidation of the impact of allogenic versus nonallogeneic factors in such a model is important. This study examined the effect of a clinically relevant combination of warm ischemia and cold preservation in the absence of allogeneic response (isografts) and in the context of operational tolerance. METHODS: Dark Agouti (DA) rat kidneys were transplanted into DA recipients (isografts) or Albino Surgery recipients (allografts) tolerized by two transfusions of DA blood, under cover of cyclosporin A. Grafts were subjected to minimal cold preservation or to 30 mins warm ischemia followed by 24 hrs cold preservation. RESULTS: After an initial peak of renal dysfunction, serum creatinine concentration returned to normal in isografts and nonischemic allografts, but remained significantly elevated in ischemic allografts (P<0.0002) throughout 6 months follow-up. Both allograft groups developed proteinuria. At 6 months, ischemic isografts and nonischemic allografts demonstrated very mild tubular atrophy and interstitial fibrosis. Tubulointerstitial injury was significantly more severe in ischemic allografts (P<0.01 vs. nonischemic allografts) and was associated with increased infiltrating monocyte/macrophages and NK cells (P<0.05). Moderate glomerulosclerosis was a feature of both allograft groups (P<0.05). CONCLUSIONS: The modified allogeneic response in operationally tolerant recipients acts in synergy with ischemia/reperfusion injury in the development of chronic damage. Strategies to limit or modify the initial ischemia/reperfusion injury may ameliorate chronic tubulointerstitial damage. Progressive glomerular damage and proteinuria in allografts may require other pharmacological intervention.

The location of insulin receptors in bovine retina and isolated retinal cells.

PURPOSE: The binding of insulin to its cell-surface receptor is the sole means by which the hormone influences cellular activity. The location of insulin receptors in bovine retina and on isolated retinal cells was investigated to determine the specific cells sensitive to insulin. METHODS: Insulin receptors were located in frozen retinal sections prepared from enucleated bovine eyes, with polyclonal anti-insulin receptor antibodies using an immuno-peroxidase method. Isolated cells were obtained by enzymatic and physical dispersion of bovine retinal tissue. Insulin receptors on isolated cells were located by a monoclonal anti-insulin receptor antibody using an immunogold silver staining technique. RESULTS: Insulin receptors demonstrated a widespread distribution throughout the bovine retina, being present in all retinal layers. A particular association with the plexiform layers and Müller cells was identified in the frozen sections. Consistent with these findings, insulin receptors were predominantly located on dendritic processes of isolated retinal neurones and on Müller cells. CONCLUSIONS: The widespread distribution of retinal insulin receptors in the bovine retina supports the hypothesis that insulin has a role in regulating retinal activity. Insulin receptors associated with plexiform regions suggests that insulin may influence neural activity, while receptors on Müller cells indicate that insulin may have a role in metabolic or functional mechanisms in bovine retina.

Pretransplant rinse of hearts preserved with colloid-free UW solution and more effective heart preservation: studies in a rat abdominal heart transplant model.

BACKGROUND: University of Wisconsin solution (UW) provides effective heart preservation under hypothermic conditions, but it can be deleterious at warmer temperatures. Re-warming during the implantation of the graft may be a problem. This study examined the damaging effect of peri-operative warm ischemia in a transplant setting and recovery from such damage. The amelioration of damage by rinsing the graft before re-warming and transplantation was also examined. METHODS: Rat donor hearts were preserved for 2 hr (0 degrees C) as follows: Series A was preserved with colloid-free UW (MUW), St. Thomas' solution (ST), or calcium-supplemented MUW (MUW+Ca) followed by either transplantation or warming (22 degrees C) for 10 min before transplantation. Series B was preserved with MUW, rinsed with fresh MUW, ST, MUW+Ca, or low-potassium MUW before warming and transplantation. All heart isografts were transplanted heterotopically with an indwelling left intraventricular balloon-tipped catheter. Graft function was measured 1 and 7 days after transplantation. RESULTS: Grafts re-warmed rapidly during implantation. Function (left ventricular developed pressure, contractility, and relaxation) was significantly and persistently diminished in MUW-preserved grafts subjected to additional warming before transplantation. Preservation with ST was less effective than MUW despite being unaffected by warming. Preservation with MUW+Ca and rinsing with fresh MUW or ST before re-warming allowed recovery of function within 7 days despite significantly diminished function on day 1. CONCLUSION: This study demonstrated that an increase in the peri-transplant warm ischemic period was detrimental when hearts were preserved with MUW. Preservation with calcium-supplemented MUW or rinsing the heart with fresh MUW or ST before transplantation ameliorated this damage.