Influence of Hepatocyte-Rich Liver Cell Mixture and Liver Fibroblasts on Prolonging Graft Islet Survival in Rats without Immunosuppressive Drugs

Abstract

Background: We noted that a liver cell suspension, made up of a mixture of several kinds of hepatic cells, affected allogenic islet survival when it was transplanted into the liver, mixed with the islets or separately.

Aim: To study if this effect was related to a liver cell mixture rich in hepatocytes (hp) or to liver fibroblasts (fb).

Methods: We studied 14 groups of rats: (A) a sham group with saline; (B) a group receiving transplantation with hepatic cells alone; (C) a control group, with islets alone via the portal vein, without hepatic cells (hp or fb). For the other groups, we used a different ratio of cells/islets (100 : 1, 150 : 1 and 200 : 1) and different co-transplantation techniques with both types of cells. For the D, E, J groups, a mixture of hepatocytes (hepatocyte-rich liver cell mixture) or fibroblasts with islets was injected into the portal vein. For the other groups, we used a sequential procedure with a 15 minute interval between a first injection of hp or fb into the portal vein or into the vena cava, and a second injection of islets always into the portal vein; thus, it was a sequential portal/portal procedure with hepatocyte-rich liver cell mixture (hp) (F, G) or fibroblasts (K, L) and a sequential cava/portal with hp (H, I) or fibroblasts (M, N).

Results: Most of the co-transplantation groups showed functional islets (blood glucose < 250 mg/dl) on the first or second day of transplantation; after several days they once again had high glucose levels, though not as high as pre-transplantation. There was statistical significance (p < 0.001) between the presence or not of hepatic cells to obtain prolongation of graft survival (blood glucose < 250 mg/dl). Statistical significance (p < 0.001) was found for several sequential groups with hp (F, I) and fb (K, L). It was also remarkable that 3 rats (37.5 %) from the I group (sequential cava/portal with hp/islets 200 : 1) were euglycemic (blood glucose < 150 mg/dl) for more than 3 months. ANOVA showed a large interaction between the type of transplant performed and the cellular ratio used, with a significance of p < 0.001. Histological studies in rats with prolonged euglycemia, showed insulin-producing cell aggregates in the liver, while there was a remarkable decrease in insulin-producing cells in the remaining islets of pancreatic tissue.

Conclusion: The results showed a marginal prolongation of islet graft survival when they are co-transplanted with a hepatocyte-rich liver cell mixture or with liver fibroblasts. The mechanism does not seem to be a cellular interaction between different hepatic cells and islets, but some kind of cellular interaction or released factor from either two cell types on the immune system, blocking or modulating it, at least temporarily.

Islet allografts - cotransplantation - hepatocytes - fibroblasts - rats

References

• 1 Alvarez S, Soto M L, Martínez B, Arranz A, Martín M C, Jara A. Diabetes reversion after allotransplantation of islets previous cultured in RPMI for 7 - 8 days compared with 1 - 2 days.  Endocrinologia. 1997;  44 119-121
• 2 Chiang Y J, Lu L, Fung J J, Qian S. Liver-derived dendritic cells induce donor-specific hyporesponsiveness: use of a sponge implant as a cell transplant model.  Cell Transplant. 2001;  10 343-350
• 3 Dafoe D C, Wang X, Tafra L, Berezniak R, Lloyd R V. Studies of composite grafts of fetal pancreas (FP) and fetal liver (FL) in the streptozotocin-induced diabetic rat.  Adv Exp Med Biol. 1992;  321 171-177
• 4 Del Río R, Soto M L, Alvarez S, Martínez B, Arranz A, Martin-Scapa M C, Jara A. Study of the number and purity of pancreatic rat islet cells in vivo and in vitro in more than 500 isolations.  Avances en Diabetología. 1997;  13 139-143
• 5 Ikebukuro K, Adachi Y, Yamada Y, Fujimoto S, Seino Y, Oyaizu H, Hioki K, Ikehara S. Treatment of streptozotocin-induced diabetes mellitus by transplantation of islet cells plus bone marrow cells via portal vein in rats.  Transplantation. 2002;  73 512-518
• 6 Jara-Albarrán A, Soto-Montenegro L, del Rio R, Alvarez S, Martin-Scapa C. Influence of hepatic cells on allogeneic islet transplantation in rats without immunosuppressive drugs.  Horm Metab Res. 2001;  33 30-33
• 7 Jara A, Soto M L, Del Río R, Zugasti A, Malvetti V. Induction of tolerance to pancreatic islet transplantation with different hepatic cell subpopulations. without immunosuppressive drugs.  Acta Chirurgica Austriaca. 2001;  33 (Suppl) 70
• 8 Jara-Albarrán A, Soto-Montenegro M L, Zugasti A, Rollán E, Alvarez Y, Alvarez G. Hepatic cells via cava vein can influence allogenic islet rat transplantation.  Cell Transplant. 2003;  12 891-896
• 9 Kaufmann P M, Fiegel H C, Kneser U, Pollok J M, Kluth D, Rogiers X. Influence of pancreatic islets on growth and differentiation on hepatocytes in co-culture.  Tissue Eng. 1999;  5 583-596
• 10 Kang S M, Braat D, Schneider D B, O'Rourke R W, Lin Z, Ascher N L, Dichet D A, Baekkeskov S, Stock P G. A non-cleavable mutant of Fas ligand does not prevent neutrophilic destruction of islet transplants.  Transplantation. 2000;  69 1813-1817
• 11 Korbutt G S, Elliott J F, Rajotte R V. Cotransplantation of allogeneic islets with allogeneic testicular cell aggregates allows long-term graft survival without systemic immunosuppression.  Diabetes. 1997;  46 317-322
• 12 Lake S P, Anderson J, Chamberlain J, Gardner S J, Bell P R, James R F. Bovine serum albumin density gradient isolation of rat pancreatic islets.  Transplantation. 1987;  43 805-808
• 13 Latif Z A, Noel J, Alejandro R. A simple method of staining fresh and cultured islets.  Transplantation. 1988;  45 827-830
• 14 Lau H T, Yu M, Fontana A, Stoeckert Jr C J. Prevention of islet allograft rejection with engineered myoblasts expressing FasL in mice.  Science. 1996;  273 109-112
• 15 Legrelle M, Chapa O, Race J M, Mallegol S, Capron F, Altman J J. Islet and hepatocyte cotransplantation in rats.  Transplant Proc. 1993;  25 981
• 16 Markees T G, Appel M C, Noelle R J, Mordes J P, Greiner D L, Rossini A A. Tolerance to islet xenografts induced by dual manipulation of antigen presentation and co-stimulation.  Transplant Proc. 1996;  28 814-815
• 17 Ricordi C, Flye M W, Lacy P E. Renal subcapsular transplantation of clusters of hepatocytes in conjunction with pancreatic islets.  Transplantation. 1988;  45 1148-1151
• 18 Ricordi C, Hering B J, London N JM, Rajotte R V, Gray D WR, Sutherland D ER. Islet isolation assessment. Ricordi C Pancreatic Islet Cell Transplantation. Austin, TX; Landes Co. 1992: 132-142
• 19 Ricordi C, Murase N, Rastellini C, Behboo R, Demetris A J, Starzl T E. Indefinite survival of rat islet allografts following infusion of donor bone marrow without cytoablation.  Cell Transplant. 1996;  5 53-55
• 20 Sandbichler P, Erhart R, Herbst P, Schmid T, Vogel W, Thaler W, Totsch M, Klima G, Margreiter R. Simultaneous transplantation of isolated hepatocytes reduces rejection of small bowel allografts and improves survival in the rat.  Eur Surg Res. 1996;  28 222-226
• 21 Sawada T, Asanuma Y, Furuya T, Saito K, Koyama K. Tolerance of islet allografts induced by orthotopic liver transplantation.  Surg Res. 1999;  85 209-216
• 22 Seglen P O. Preparation of isolated rat liver cells.  Methods Cell Biol. 1976;  13 29-83
• 23 Selawry H, Fojaco R, Whittington K. Extended survival of MHC-compatible islet grafts from diabetes-resistant donors in spontaneously diabetic BB/W rat.  Diabetes. 1987;  36 1061-1067
• 24 Selawry H P, Cameron D F. Sertoli cell-enriched fractions in successful islet cell transplantation.  Cell Transplant. 1993;  2 123-129
• 25 Sun J, Wang X, Wang C, Sheil A G. Sequential transplantation induces islet allograft tolerance.  Microsurgery. 2001;  21 148-152
• 26 Tafra L, Berezniak R, Dafoe D C. Beneficial effects of fetal liver tissue on pancreatic transplantation.  Surgery. 1990;  108 734-740
• 27 Takeda Y, Gotoh M, Dono K, Nishihara M, Grochowiecki T, Kimura F, Yoshida T, Ohta Y, Ota H, Ohzato H, Umeshita K, Takeda T, Matsuura N, Sakon M, Kayagaki N, Yagita H, Okumura K, Miyasaka M, Monden M. Protection of islet allografts transplanted together with Fas ligand expressing testicular allografts.  Diabetologia. 1998;  41 315-321
• 28 Wang C, Sun J, Li L, Wang L, Dolan P, Sheil A G. Conversion of pancreas allograft rejection to acceptance by liver transplantation.  Transplantation. 1998;  65 188-192
• 29 Wang X Y, Sun J, Wang C, France M, Sheil A G. Effect of liver transplantation on islet allografts: up-regulation of Fas ligand and apoptosis of T lymphocytes are associated with islet graft tolerance.  Transplantation. 2001;  71 102-111
• 30 Yang L, Li S, Hatch H, Ahrens K, Cornelius J G, Petersen B E, Peck A B. In vitro trans-differentiation of adult hepatic stem cells into pancreatic endocrine hormone-producing cells.  Proc Natl Acad Sci USA. 2002;  99 8078-8083
• 31 Yoo-Ott K A, Schiller H, Fandrich F, Oswald H, Richter K, Xhu X F, Kampen W U, Kronke M, Zavazava N. Co-transplantation of donor-derived hepatocytes induces long-term tolerance to cardiac allografts in a rat model.  Transplantation. 2000;  69 2538-2546