ISSUES RAISED BY HUMAN CLONING RESEARCH HEARING ...

41With animals representing 5 different mammalian species now having been producedby somatic cell nuclear transfer, cloning has been proposed as a tool for assistedreproduction in humans i.e. a means for producing a human baby. Experimentsfrom our laboratory and others provide strong evidence that the current proceduresused for mammalian cloning are not safe and many times result in abnormaldevelopment. This can ultimately lead to death of the cloned offspring and thesurrogate mother. Based on these observations and evidence from studies in micewhich demonstrate incompatibilities between nucleus and cytoplasm from differentstrains, cloning as an approach to human assisted reproduction is at present bothrisky and extremely irresponsible.Although animals can be cloned by nuclear transplantation using somatic cells asnucleus donors, the efficiency of the technique is still extremely low. In cattle wherethe majority of the work has been completed, problems with early embryonic developmentdo not seem to be a major factor affecting the efficiency of cloning, as developmentrates to the blastocyst stage in vitro are similar to those of normal embryosproduced by in vitro fertilization. Maternal recognition and the establishment ofpregnancy as indicated by pregnancy rates at 35 days of gestation are also similarbetween normal embryos and those produced by nuclear transplantation. However,after 35 days of gestation, pregnancy loss is dramatic and very few fetuses surviveto term. Approximately 90% of the pregnancies are lost and abort between days 35and 90 of gestation (the first trimester). The most common developmental malformationobserved to date is aberrant placentation. Of those calves that do survive, mostexhibit placental edema and a reduced number of enlarged placentomes. These placentalabnormalities pose serious health risks not only to the developing fetus andoffspring but also to the surrogate mothers carrying the pregnancies. In severalcases involving cattle, both the surrogate mother and the bovine fetuses have diedduring late gestation due to a variety of complicated health issues related to the abnormalpregnancy. Moreover, even if the cloned offspring survive to term, many ofthe resulting calves exhibit developmental abnormalities and die at birth or shortlythereafter, normally a result of cardiopulmonary abnormalities. In general, regardlessof the species, only 1%-5% of cloned embryos survive to term.In our laboratory we have utilized nuclear transfer to try and reproduce thegenotypes of several different animals, selected for cloning based on their inherentgenetic value. Results we have obtained to date are similar to those reported byother laboratories regardless of the species involved. The first case involved a Brahmansteer named ‘‘Chance’’, known to be at least 21 years old. Adult fibroblastswere obtained from a skin biopsy and expanded in culture using standard methodsfor tissue culture prior to being frozen and stored in liquid nitrogen. When nucleartransfer was performed using the fibroblast cells derived from Chance, 28% of thefused couplets (53 of 190) developed into blastocysts in culture. Twenty-six of thesewere transferred into 11 recipient cows resulting in 6 pregnancies. Three of thesecontinued to develop through 90 days of gestation but only one survived to term.‘‘Second Chance’’ is now over a year old and appears normal and healthy for his age.However during the first week of life he required intensive monitoring and therapyto treat lung dysmaturity and pulmonary hypertension. At 7 days of age he was alsodiagnosed and treated for Type 1 insulin-dependent diabetes, which is extremelyrare in cattle. He also lacked the expression of an important T-cell antigen CD45,indicating his immune system was in some way abnormal (Hill et al, 2000).The second and third attempts at reproducing desired genotypes by cloning involvedtwo middle-aged cows, one Brangus and one Charolais. These were selectedbased on being top performers in the herd. Fibroblasts were again obtained fromskin biopsies. Development rate to the blastocyst stage following nuclear transferand embryo culture averaged 16%. Thirty-seven blastocysts derived from theCharolais cow were transferred into 13 recipients. Six of these were diagnosed aspregnant at 30 days of gestation but only 4 remained pregnant through 60 days.One of these pregnancies was subsequently lost. In two cases the fetus was removedfor research purposes. The final pregnancy was allowed to proceed to term resultingin twin heifers. However, both calves died between 7-10 days after birth due to complicationsrelated to the cloning procedure. Forty-three blastocysts derived from theBrangus cow were transferred into 14 recipients resulting in 3 pregnancies. Howevernone of these survived past 90 days of gestation.Our most recent attempt at cloning a specific animal has involved a deceasedBlack Angus bull previously shown to be naturally (genetically) resistant to Brucellosis.Of the oocyte-fibroblast couplets fused and cultured, 44% developed to theblastocyst stage. Thirty-nine blastocysts were transferred into 20 recipients resultingin 10 pregnancies at 35 days of gestation. One of these survived to approxi-VerDate 11-MAY-2000 07:46 May 24, 2001 Jkt 000000 PO 00000 Frm 00045 Fmt 6633 Sfmt 6621 71495.TXT HCOM2 PsN: HCOM2