Effects of immunomodulator " Cyclosporine " on regenerative capacity of amputated hindlimbs in larval stages of the Egyptian toad , Bufo regularis

The regeneration of the amputed hindlimbs of stage (54) of tadpole larvae of Bufo regularis was studied after amputing their hind limbs at the mid shank level. The immunomodulator Cyclosporine A (CsA) was selected to determine its effect on regeneration. Two kinds of experiments were desigened. The first of experimental design is by immersing the tadpole larvae in a solution of CsA (0.02, 0.2 and 0.5 mg/mL) for five hours daily for three consecutive days, starting the treatment twelve hours after amputation. The second one is by intraperitoneal injection of CsA solution (0.02 and 0.2 mg/mL) administering 1 μ of CsA solution every other day starting from one day preamputation to day eleven post amputation. Regarding the effect of CsA on regeneration on stage 54, CsA significantly promoted regeneration in both kinds of experiments (immersion and injection), and this effect was dose-dependent. The regerative ability increases with increase of CsA concentations. Significant number of cases regenerated complete hind limbs with five or four toes, 86.3%, 66.7% and 100% of CsA treated cases (0.02, 0.2 and 0.5 mg/mL respectively) versus 45% of control cases when treated by immersion. In the other experiment, which is by injection, 90.5% and 93.3% of CsA treated cases (0.02 and 0.2 mg/mL respectively) restored hind limbs with five or four toes versus 45% of control cases. Histologically, CsA was associated with acceleration in the processes of dedifferentiation, differentiation and histogenesis in comparison with their control counterparts. Moreover, CsA treatment delayed the dermal differentiation underneath the epithelium. An observation which certainly favors regeneration as the absence of dermis allows direct communication between the above epithelium and the underlying cells for better blastema formation. CsA treatment in both experiments was also associated with early dedifferentiation of mesenchyme cells forming a well-formed blastema, and early differentiation of skeletal elements of the regenerates in comparison with their control counterparts. Adminstration of CsA to the tadepole larvae (54 stage) by injection resulted in better regenerative outcomes than adminstration by immersion. CsA significantly promoted and accelerated regeneration of their hind limbs after ambutation at the mid shank level. In addition, CsA effect was dose-dependent. We also consider that these data will give us deeper insights on the mechanisms taking place in higher vertebrates and in human. [Gehan Hassan Fahmy, Kawakeb Ali Omran and Eman Abbas Moussa: Effects of immunomodulator "Cyclosporine" on regenerative capacity of amputated hindlimbs in larval stages of the Egyptian toad, Bufo regularis. Life Sci J 2014;11(7):791-802]. (ISSN:1097-8135). http://www.lifesciencesite. com. 115 Abbreviations of figures 2-15: AsC: astragalus and calcanum, BL: blastema cells, BM: basement membrane, CP: condylar cap. , D: dermis, DP: digit primordia, M: muscle fibers, ML: melanophores, P: periosteum, PH: phalanges, TF: tibiofibula and UG:unicellular glands