Although the sheep has been the first animal to be cloned by somatic cells nucleartransfer (SCNT), post natal survival of cloned lambs seems to be lower than the otheranimal cloned so far. We present the results of a large scale cloning experiment designed toexplore the causes of peri- and post-natal mortality of cloned lambs. Blastocysts (n. 93)obtained by nuclear transfer of somatic cells (granulosa cells), were transferred into 41recipient ewes, and pregnancies were monitored by ultrasound scanning. In vitro derived,fertilized embryos (IVF, n=123) were also transferred as a control. Pre and early postimplantation development was comparable between cloned and control IVF embryos;however, dramatic losses occurred from the first month of pregnancy onward, with only 12out of 93 (13%) clones reaching full-term development, compared to 51 out of 123 (41.6%)lambs born from the IVF control embryos. Blood samples collected from the cloned lambsafter birth, revealed a wide range of abnormalities (hyperkalemia -16 mEq/l in clones, 4.7in controls -; hypernatriemia: -189 mEq/l in clones, 139 in controls). In addition, serumchemistry revealed high levels of ammonia, creatine, urea and a low protein content (inaverage 68 mg/dl, 21 mg/dl 46 mg/dl 2.0 g/dl clones; 38 mg/dl, 2 mg/dl 16 mg/dl 7.0g/dl in controls). These findings are indicative of kidney and liver dysfunction consequent toplacental hypotrophy. In fact, macroscopical and histopathological examination of theplacentae revealed a marked reduction in vascularization, particularly at the apex of thevillous processes, as well as a loss of differentiation of the trophoblastic epithelium. Ourresults strongly suggest that post mortality of 8 of the cloned lambs was primarily due toplacental abnormalities, for no overt, or histological lesions were detected in the clonedlambs. These results provide further evidence for defective function of extraembryonic tissuein somatic cells clones. It is widely accepted that defective development of clones resultsfrom incomplete reprogramming of somatic cells leading to abnormal expression ofimprinted and non imprinted genes. The pictures that results from our, and other companionarticle on other species, is that abnormal reprogramming regards mainly placental genes,which, as know, are mostly regulated by paternally expressed genes. We speculate that thegenome of a differentiated cells undergoes a sort of “asymmetric reprogramming” by theoocyte, which leave unaffected the paternal side of the genome. Our preliminary data onDNA methylation profile of monoparental embryos (androgenetic and parthenogenetic) andsomatic cells clones (Loi P. Beaujean N, unpublished) apparently support this hypothesis.[...]
Frequency and reasons of post natal mortality in sheep somatic cell clones
LOI, Pasqualino;PALMIERI, CHIARA;DELLA SALDA, Leonardo;PTAK, Grazyna
2005-01-01
Abstract
Although the sheep has been the first animal to be cloned by somatic cells nucleartransfer (SCNT), post natal survival of cloned lambs seems to be lower than the otheranimal cloned so far. We present the results of a large scale cloning experiment designed toexplore the causes of peri- and post-natal mortality of cloned lambs. Blastocysts (n. 93)obtained by nuclear transfer of somatic cells (granulosa cells), were transferred into 41recipient ewes, and pregnancies were monitored by ultrasound scanning. In vitro derived,fertilized embryos (IVF, n=123) were also transferred as a control. Pre and early postimplantation development was comparable between cloned and control IVF embryos;however, dramatic losses occurred from the first month of pregnancy onward, with only 12out of 93 (13%) clones reaching full-term development, compared to 51 out of 123 (41.6%)lambs born from the IVF control embryos. Blood samples collected from the cloned lambsafter birth, revealed a wide range of abnormalities (hyperkalemia -16 mEq/l in clones, 4.7in controls -; hypernatriemia: -189 mEq/l in clones, 139 in controls). In addition, serumchemistry revealed high levels of ammonia, creatine, urea and a low protein content (inaverage 68 mg/dl, 21 mg/dl 46 mg/dl 2.0 g/dl clones; 38 mg/dl, 2 mg/dl 16 mg/dl 7.0g/dl in controls). These findings are indicative of kidney and liver dysfunction consequent toplacental hypotrophy. In fact, macroscopical and histopathological examination of theplacentae revealed a marked reduction in vascularization, particularly at the apex of thevillous processes, as well as a loss of differentiation of the trophoblastic epithelium. Ourresults strongly suggest that post mortality of 8 of the cloned lambs was primarily due toplacental abnormalities, for no overt, or histological lesions were detected in the clonedlambs. These results provide further evidence for defective function of extraembryonic tissuein somatic cells clones. It is widely accepted that defective development of clones resultsfrom incomplete reprogramming of somatic cells leading to abnormal expression ofimprinted and non imprinted genes. The pictures that results from our, and other companionarticle on other species, is that abnormal reprogramming regards mainly placental genes,which, as know, are mostly regulated by paternally expressed genes. We speculate that thegenome of a differentiated cells undergoes a sort of “asymmetric reprogramming” by theoocyte, which leave unaffected the paternal side of the genome. Our preliminary data onDNA methylation profile of monoparental embryos (androgenetic and parthenogenetic) andsomatic cells clones (Loi P. Beaujean N, unpublished) apparently support this hypothesis.[...]I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.