Assessment of the role of neutrophils on the antitumor effect of TNFalpha in an in vivo isolated limb perfusion model in sarcoma-bearing brown Norway rats.
BACKGROUND
Isolated limb perfusion (ILP) with TNFalpha in combination with melphalan and IFNgamma has resulted in an immediate and dramatic tumor response in patients. Such an effect was also noted following ILP in a rat sarcoma model. This model enables us to investigate several factors responsible for the TNFalpha-induced tumor responses. We applied total body irradiation (TBI) to reduce white blood cell count, to investigate the contribution of leukocytes to the anti-tumor effect of TNFalpha.


METHODS
Small fragments of the nonimmunogenic BN 175 sarcoma were implanted sc in the lower hind leg. A 5 Gy TBI was performed before ILP at a tumor diameter of approximately 15 mm. The hind limbs of 63 rats were perfused and were divided into 6 groups: group 1, sham perfusion, n = 9; group 2, TBI + sham perfusion, n = 6; group 3, TNFalpha 50 microgram, n = 9; group 4, melphalan 40 microgram, n = 9; group 5, TNFalpha 50 microgram + melphalan 40 microgram, n = 22; group 6, TBI + TNFalpha + melphalan ILP, n = 8. In addition, 10 rats were perfused for histological analysis at 24 h post-ILP.


RESULTS
We observed in Group 1: 9/9 progressive disease (PD); Group 2: 6/6 PD; Group 3: 9/9 PD; Group 4: 9/9 no change (NC) of tumor diameter for at least 4 days; Group 5: 6/22 NC, 16/22 complete remission (CR), 12/16 of which showed skin necrosis at the tumor site; and Group 6: 7/8 NC and 1/8 CR (without skin necrosis). After TBI, WBC reduction of 80-95% was observed, while the number of platelets was not significantly reduced and platelet aggregation was maintained at 72 %. Histological analysis revealed decreased hemorrhagic necrosis associated with the absence of PMN infiltration at the tumor margins in the TBI rats.


CONCLUSIONS
TBI and the associated reduction in WBC count decreased the tumor response by TNFalpha and melphalan significantly and abrogated the immediate response of skin necrosis at the tumor site, as found in rats treated with TNFalpha and melphalan without TBI. These data strongly suggest that leukocytes play an important role in the hemorrhagic effects of TNFalpha.
Manusama
E R
ER
Department of Surgery, University Hospital Rotterdam-Dr Daniel den Hoed Cancer Center, Rotterdam, The Netherlands.
Nooijen
P T
PT
Stavast
J
J
de Wilt
J H
JH
Marquet
R L
RL
Eggermont
A M
AM
D000818:Animals; D005121:Extremities / Q000098:blood supply*; D007958:Leukocyte Count; D008297:Male; D008558:Melphalan / Q000494:pharmacology; D009336:Necrosis; D009389:Neovascularization, Pathologic / Q000188:drug therapy / Q000473:pathology; D009504:Neutrophils / Q000276:immunology*; D009924:Organ Culture Techniques; D010974:Platelet Aggregation; D051381:Rats; D011914:Rats, Inbred BN; D012513:Sarcoma, Experimental / Q000098:blood supply / Q000188:drug therapy* / Q000473:pathology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D014409:Tumor Necrosis Factor-alpha; D008558:Melphalan